Journal articles on the topic 'Calpain'
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1
Chhabra, A., H. Fernando, R. E. Mansel, and W. G. Jiang. "Pattern of expression of calpain subunits (large and small) in human breast cancer and the prognostic significance." Journal of Clinical Oncology 25, no. 18_suppl (June 20, 2007): 21078. http://dx.doi.org/10.1200/jco.2007.25.18_suppl.21078.
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21078 Background: Calpains belongs to family of non-lysosomal calcium dependent cysteine proteases which are known to regulate cellular migration, apoptosis and cell cycle progression in normal and tumour cells. Calpain has also been indicated in the metastatic process of certain tumours like RCC and identified as a potential tumor suppressor for gastric cancer. Little is known about the expression pattern of calpain in human breast cancer. The current study investigated the expression pattern of the large (calpainL) and small (calpainS) subunits of calpain which are encoded by different genes, in human breast cancer and attempted to correlate the expression with clinical outcome. Methods: RNA was extracted from frozen sections of breast tissue (n=120, median clinical follow up of the patients - 72 months) for gene amplification. The expression of both subunits of calpain was determined by using RT-PCR and quantitative RT-PCR. Statistical analysis was carried out using Mann- Whitney U test and the Kruskal- Wallis test. Results: We found significantly higher level of both calpain subunits in tumour tissue (n=120) as compared to normal non-neoplastic mammary tissues (calpainL, p=0.022 and calpainS, p=0.039). The expression of calpainL was significantly lower in patients with poor clinical outcome (with metastasis, p=0.024, with local recurrence, p=0.024 and who died of breast cancer, p=0.028), than those who were disease free. In contrast, the levels of calpainS were high in patients with poor prognosis, metastasis and who died of breast cancer but were statistically not significant. The expression of calpainL was marginally lower in node positive tumours than the node negative tumours. CalpainS transcripts were higher in node positive than node negative tumours (p=0.23). Similarly, calpainL showed a low level in high grade tumours whereas calpainS displayed a contrasting high level in these tumours. Conclusions: The large and small subunits of calpain have a distinct pattern of expression in human breast cancer. A decrease expression of calpainL but an increase expression of calpainS may be associated with poor prognostic tumours, thus indicating the intimate nature of the molecular pair in the disease progression of human breast cancer. No significant financial relationships to disclose.
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Upla, Paula, Varpu Marjomäki, Liisa Nissinen, Camilla Nylund, Matti Waris, Timo Hyypiä, and Jyrki Heino. "Calpain 1 and 2 Are Required for RNA Replication of Echovirus 1." Journal of Virology 82, no. 3 (November 21, 2007): 1581–90. http://dx.doi.org/10.1128/jvi.01375-07.
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ABSTRACT Calpains are calcium-dependent cysteine proteases that degrade cytoskeletal and cytoplasmic proteins. We have studied the role of calpains in the life cycle of human echovirus 1 (EV1). The calpain inhibitors, including calpeptin, calpain inhibitor 1, and calpain inhibitor 2 as well as calpain 1 and calpain 2 short interfering RNAs, completely blocked EV1 infection in the host cells. The effect of the inhibitors was not specific for EV1, because they also inhibited infection by other picornaviruses, namely, human parechovirus 1 and coxsackievirus B3. The importance of the calpains in EV1 infection also was supported by the fact that EV1 increased calpain activity 3 h postinfection. Confocal microscopy and immunoelectron microscopy showed that the EV1/caveolin-1-positive vesicles also contain calpain 1 and 2. Our results indicate that calpains are not required for virus entry but that they are important at a later stage of infection. Calpain inhibitors blocked the production of EV1 particles after microinjection of EV1 RNA into the cells, and they effectively inhibited the synthesis of viral RNA in the host cells. Thus, both calpain 1 and calpain 2 are essential for the replication of EV1 RNA.
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Fontenele, Marcio, Bomyi Lim, Danielle Oliveira, Márcio Buffolo, David H. Perlman, Trudi Schupbach, and Helena Araujo. "Calpain A modulates Toll responses by limited Cactus/IκB proteolysis." Molecular Biology of the Cell 24, no. 18 (September 15, 2013): 2966–80. http://dx.doi.org/10.1091/mbc.e13-02-0113.
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Calcium-dependent cysteine proteases of the calpain family are modulatory proteases that cleave their substrates in a limited manner. Among their substrates, calpains target vertebrate and invertebrate IκB proteins. Because proteolysis by calpains potentially generates novel protein functions, it is important to understand how this affects NFκB activity. We investigate the action of Calpain A (CalpA) on the Drosophila melanogaster IκB homologue Cactus in vivo. CalpA alters the absolute amounts of Cactus protein. Our data indicate, however, that CalpA uses additional mechanisms to regulate NFκB function. We provide evidence that CalpA interacts physically with Cactus, recognizing a Cactus pool that is not bound to Dorsal, a fly NFκB/Rel homologue. We show that proteolytic cleavage by CalpA generates Cactus fragments lacking an N-terminal region required for Toll responsiveness. These fragments are generated in vivo and display properties distinct from those of full-length Cactus. We propose that CalpA targets free Cactus, which is incorporated into and modulates Toll-responsive complexes in the embryo and immune system.
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SORIMACHI, Hiroyuki, Shoichi ISHIURA, and Koichi SUZUKI. "Structure and physiological function of calpains." Biochemical Journal 328, no. 3 (December 15, 1997): 721–32. http://dx.doi.org/10.1042/bj3280721.
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For a long time now, two ubiquitously expressed mammalian calpain isoenzymes have been used to explore the structure and function of calpain. Although these two calpains, μ- and m-calpains, still attract intensive interest because of their unique characteristics, various distinct homologues to the protease domain of μ- and m-calpains have been identified in a variety of organisms. Some of these ‘novel’ calpain homologues are involved in important biological functions. For example, p94 (also called calpain 3), a mammalian calpain homologue predominantly expressed in skeletal muscle, is genetically proved to be responsible for limb-girdle muscular dystrophy type 2A. Tra-3, a calpain homologue in nematodes, is involved in the sex determination cascade during early development. PalB, a key gene product involved in the alkaline adaptation of Aspergillus nidulans, is the first example of a calpain homologue present in fungi. These findings indicate various important functional roles for intracellular proteases belonging to the calpain superfamily.
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Murphy, Robyn M., Rodney J. Snow, and Graham D. Lamb. "μ-Calpain and calpain-3 are not autolyzed with exhaustive exercise in humans." American Journal of Physiology-Cell Physiology 290, no. 1 (January 2006): C116—C122. http://dx.doi.org/10.1152/ajpcell.00291.2005.
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μ-calpain and calpain-3 are Ca2+-dependent proteases found in skeletal muscle. Autolysis of calpains is observed using Western blot analysis as the cleaving of the full-length proteins to shorter products. Biochemical assays suggest that μ-calpain becomes proteolytically active in the presence of 2–200 μM Ca2+. Although calpain-3 is poorly understood, autolysis is thought to result in its activation, which is widely thought to occur at lower intracellular Ca2+ concentration levels ([Ca2+]i; ∼1 μM) than the levels at which μ-calpain activation occurs. We have demonstrated the Ca2+-dependent autolysis of the calpains in human muscle samples and rat extensor digitorum longus (EDL) muscles homogenized in solutions mimicking the intracellular environment at various [Ca2+] levels (0, 2.5, 10, and 25 μM). Autolysis of calpain-3 was found to occur across a [Ca2+] range similar to that for μ-calpain, and both calpains displayed a seemingly higher Ca2+ sensitivity in human than in rat muscle homogenates, with ∼15% autolysis observed after 1-min exposure to 2.5 μM Ca2+ in human muscle and almost none after 1- to 2-min exposure to the same [Ca2+]i level in rat muscle. During muscle activity, [Ca2+]i may transiently peak in the range found to autolyze μ-calpain and calpain-3, so we examined the effect of two types of exhaustive cycling exercise (30-s “all-out” cycling, n = 8; and 70% V̇o2 peak until fatigue, n = 3) on the amount of autolyzed μ-calpain or calpain-3 in human muscle. No significant autolysis of μ-calpain or calpain-3 occurred as a result of the exercise. These findings have shown that the time- and concentration-dependent changes in [Ca2+]i that occurred during concentric exercise fall near but below the level necessary to cause autolysis of calpains in vivo.
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Theopold, U., M. Pintér, S. Daffre, Y. Tryselius, P. Friedrich, D. R. Nässel, and D. Hultmark. "CalpA, a Drosophila calpain homolog specifically expressed in a small set of nerve, midgut, and blood cells." Molecular and Cellular Biology 15, no. 2 (February 1995): 824–34. http://dx.doi.org/10.1128/mcb.15.2.824.
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Calpains are calcium-dependent proteases believed to participate in calcium-regulated signal pathways in cells. Ubiquitous calpains as well as tissue-specific calpains have been found in vertebrates. We isolated cDNA clones for a highly tissue-specific calpain gene from Drosophila melanogaster, CalpA, at 56C-D on the second chromosome. The expression of the CalpA gene product was monitored by using a specific antiserum directed against the product expressed by one cDNA clone. The encoded protein is found in a few neurons in the central nervous system, in scattered endocrine cells in the midgut, and in blood cells. In the blood cell line mbn-2, calpain is associated with a granular component in the cytoplasm. The expression of this protein is more restricted than that of the corresponding transcripts, which are widely distributed in the central nervous system, digestive tract, and other tissues. The sequence of CalpA is closely related to that of vertebrate calpains, but an additional segment is inserted in the calmodulin-like carboxy-terminal domain. This insert contains a hydrophobic region that may be involved in membrane attachment of the enzyme. Differential splicing also gives rise to a minor transcript that lacks the calmodulin-like domain.
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Covington, Marisa D., David D. Arrington, and Rick G. Schnellmann. "Calpain 10 is required for cell viability and is decreased in the aging kidney." American Journal of Physiology-Renal Physiology 296, no. 3 (March 2009): F478—F486. http://dx.doi.org/10.1152/ajprenal.90477.2008.
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Aging is associated with abnormalities in kidney function, but the exact mechanisms are unknown. We examined calpains 1, 2, and 10 protein levels in kidneys from rats, mice, and humans of various ages and determined whether calpain 10 is required for cell viability. Calpain 10 protein expression decreased in the kidney, but not in the liver, of aging Fischer 344 rats, and this decrease was attenuated with caloric restriction. There was no change in calpains 1 or 2 levels in the kidney or liver in control and caloric-restricted aging rats. Aging mice also exhibited decreased calpain 10 protein levels. Calpain 10 protein and mRNA levels decreased linearly in human kidney samples with age in the absence of changes in calpains 1 or 2. Our laboratory previously found calpain 10 to be expressed in both the cytosol and mitochondria of rabbit renal proximal tubular cells (RPTC). Adenoviral-delivered shRNA to rabbit RPTC decreased mitochondrial calpain 10 expression below detectable levels by 3 days while cytosolic calpain 10 levels remained unchanged at 3 days and decreased to ∼20% of control by 5 days. Knockdown of mitochondrial calpain 10 resulted in nuclear condensation and cleaved procaspase 3, markers of apoptosis. In summary, mitochondrial calpain 10 is required for cell viability and calpain 10 levels specifically decrease in aging rat, mice, and human kidney tissues when renal function decreases, suggesting that calpain 10 is required for renal function and is a biomarker of the aging kidney.
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Arora, A. S., P. de Groen, Y. Emori, and G. J. Gores. "A cascade of degradative hydrolase activity contributes to hepatocyte necrosis during anoxia." American Journal of Physiology-Gastrointestinal and Liver Physiology 270, no. 2 (February 1, 1996): G238—G245. http://dx.doi.org/10.1152/ajpgi.1996.270.2.g238.
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Calpain proteases contribute to hepatocyte necrosis during anoxia. Our aim was to ascertain the mechanism causing calpain activation during anoxia. In rat hepatocytes, a twofold increase in calpain activity occurred despite the lack of an increase in cytosolic Ca2+ concentration ([Ca2+]i). The increase in calpain activity was not associated with an increase in calpain mRNA or a decrease in calpastatin mRNA expression. Because phospholipid degradation products generated by phospholipases can activate calpains at physiological [Ca2+]i, we determined the effect of phospholipase inhibitors and activators on calpain activity. Pretreatment of hepatocytes with fluphenazine, a phospholipase inhibitor, decreased calpain activation and improved cell survival. Melittin, a phospholipase A2 activator, increased calpain activity and potentiated cell killing. Finally, phospholipid degradation preceded the increase in calpain activity. Thus the enhanced calpain activity occurring in hepatocytes during anoxia 1) is regulated at the posttranslational level and 2) appears to be dependent on phospholipase activity. These data suggest a novel cascade for degradative hydrolase activity during hepatocyte necrosis by anoxia with phospholipase-mediated activation of calpains.
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Sultan, Karim R., Bernd T. Dittrich, and Dirk Pette. "Calpain activity in fast, slow, transforming, and regenerating skeletal muscles of rat." American Journal of Physiology-Cell Physiology 279, no. 3 (September 1, 2000): C639—C647. http://dx.doi.org/10.1152/ajpcell.2000.279.3.c639.
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Fiber-type transitions in adult skeletal muscle induced by chronic low-frequency stimulation (CLFS) encompass coordinated exchanges of myofibrillar protein isoforms. CLFS-induced elevations in cytosolic Ca2+ could activate proteases, especially calpains, the major Ca2+-regulated cytosolic proteases. Calpain activity determined by a fluorogenic substrate in the presence of unaltered endogenous calpastatin activities increased twofold in low-frequency-stimulated extensor digitorum longus (EDL) muscle, reaching a level intermediate between normal fast- and slow-twitch muscles. μ- and m-calpains were delineated by a calpain-specific zymographical assay that assessed total activities independent of calpastatin and distinguished between native and processed calpains. Contrary to normal EDL, structure-bound, namely myofibrillar and microsomal calpains, were abundant in soleus muscle. However, the fast-to-slow conversion of EDL was accompanied by an early translocation of cytosolic μ-calpain, suggesting that myofibrillar and microsomal μ-calpain was responsible for the twofold increase in activity and thus involved in controlled proteolysis during fiber transformation. This is in contrast to muscle regeneration where m-calpain translocation predominated. Taken together, we suggest that translocation is an important step in the control of calpain activity in skeletal muscle in vivo.
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Miyazaki, Takuro. "Calpain and Cardiometabolic Diseases." International Journal of Molecular Sciences 24, no. 23 (November 26, 2023): 16782. http://dx.doi.org/10.3390/ijms242316782.
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Calpain is defined as a member of the superfamily of cysteine proteases possessing the CysPC motif within the gene. Calpain-1 and -2, which are categorized as conventional isozymes, execute limited proteolysis in a calcium-dependent fashion. Accordingly, the calpain system participates in physiological and pathological phenomena, including cell migration, apoptosis, and synaptic plasticity. Recent investigations have unveiled the contributions of both conventional and unconventional calpains to the pathogenesis of cardiometabolic disorders. In the context of atherosclerosis, overactivation of conventional calpain attenuates the barrier function of vascular endothelial cells and decreases the immunosuppressive effects attributed to lymphatic endothelial cells. In addition, calpain-6 induces aberrant mRNA splicing in macrophages, conferring atheroprone properties. In terms of diabetes, polymorphisms of the calpain-10 gene can modify insulin secretion and glucose disposal. Moreover, conventional calpain reportedly participates in amino acid production from vascular endothelial cells to induce alteration of amino acid composition in the liver microenvironment, thereby facilitating steatohepatitis. Such multifaceted functionality of calpain underscores its potential as a promising candidate for pharmaceutical targets for the treatment of cardiometabolic diseases. Consequently, the present review highlights the pivotal role of calpains in the complications of cardiometabolic diseases and embarks upon a characterization of calpains as molecular targets.
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Pánico, Pablo, Marcia Hiriart, Patricia Ostrosky-Wegman, and Ana María Salazar. "TUG is a calpain-10 substrate involved in the translocation of GLUT4 in adipocytes." Journal of Molecular Endocrinology 65, no. 3 (October 2020): 45–57. http://dx.doi.org/10.1530/jme-19-0253.
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The calpain-10 (CAPN10) protease is implicated in the translocation of the glucose transporter 4 (GLUT4), which is retained in the Golgi matrix via the Tether containing a UBX domain for GLUT4 (TUG) protein. Insulin stimulation induces the proteolytic processing of TUG, which leads to the translocation of GLUT4 to the cell membrane. We tested whether TUG is a CAPN10 substrate. Proteolysis of TUG by calpains was assessed using a cell-free system containing calpain-1 and TUG. In situ proteolysis of TUG by calpains was demonstrated in 3T3-L1 adipocytes in the presence of insulin or calpain inhibitors to modulate calpain activity. Proteolysis of TUG by CAPN10 was confirmed using transient or stable silencing of CAPN10 in 3T3-L1 adipocytes. Calpains proteolyzed the C-terminus of TUG in vitro. In adipocytes, insulin-induced cleavage of TUG was correlated with the activation of calpains. Treatment with calpain inhibitors reduced TUG cleavage, resulting in impaired GLUT4 translocation without altering Akt phosphorylation. Furthermore, CAPN10 but not calpain-1 or calpain-2 colocalized with GLUT4 in the absence of insulin, and their colocalization was reduced after stimulation with insulin. Finally, we demonstrated that CAPN10 knockdown reduced the proteolysis of TUG without altering the phosphorylation of Akt or the expression of the Usp25m protease. Thus, our results provide evidence that the TUG protein is cleaved by CAPN10 to regulate GLUT4 translocation.
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Mellgren, Ronald L., and Xinhua Huang. "Fetuin A Stabilizes m-Calpain and Facilitates Plasma Membrane Repair." Journal of Biological Chemistry 282, no. 49 (October 17, 2007): 35868–77. http://dx.doi.org/10.1074/jbc.m706929200.
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Yeast two-hybrid experiments identified α2-Heremans-Schmid glycoprotein (human fetuin A) as a binding partner for calpain domain III (DIII). The tandem DIIIs of calpain-10 interacted under the most selective culture conditions, but DIIIs of m-calpain, calpain-3, and calpain-5 also interacted under less stringent selection. DIIIs of μ-calpain, calpain-6, and the tandem DIII-like domains of the Dictyostelium Cpl protein did not interact with α2-Heremans-Schmid glycoprotein in the yeast two-hybrid system. Bovine fetuin A stabilized proteolytic activity of purified m-calpain incubated in the presence of mm calcium chloride and prevented calcium-dependent m-calpain aggregation. Consistent with the yeast two-hybrid studies, fetuin A neither stabilized μ-calpain nor prevented its aggregation. Confocal immunofluorescence microscopy of scratch-damaged L6 myotubes demonstrated accumulation of m-calpain at the wound site in association with the membrane repair protein, dysferlin. m-Calpain also co-localized with fluorescein-labeled fetuin A at the wound site. The effect of fetuin A on calpain-mediated plasma membrane resealing was investigated using fibroblasts from Capns1-/- and Capns1+/+ mouse embryos. Capns1 encodes the small noncatalytic subunit that is required for the proteolytic function of m- and μ-calpains. Thus, Capns1-/- fibroblasts do not express these calpains in active form. Fetuin A increased resealing of scrape-damaged wild-type fibroblasts but not Capns1-/- fibroblasts. These studies identify fetuin A as a potential extracellular regulator of m-calpain at nascent sites of plasma membrane wounding.
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Weber, Jonasz J., Eva Haas, Yacine Maringer, Stefan Hauser, Nicolas L. P. Casadei, Athar H. Chishti, Olaf Riess, and Jeannette Hübener-Schmid. "Calpain-1 ablation partially rescues disease-associated hallmarks in models of Machado-Joseph disease." Human Molecular Genetics 29, no. 6 (January 21, 2020): 892–906. http://dx.doi.org/10.1093/hmg/ddaa010.
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Abstract Proteolytic fragmentation of polyglutamine-expanded ataxin-3 is a concomitant and modifier of the molecular pathogenesis of Machado–Joseph disease (MJD), the most common autosomal dominant cerebellar ataxia. Calpains, a group of calcium-dependent cysteine proteases, are important mediators of ataxin-3 cleavage and implicated in multiple neurodegenerative conditions. Pharmacologic and genetic approaches lowering calpain activity showed beneficial effects on molecular and behavioural disease characteristics in MJD model organisms. However, specifically targeting one of the calpain isoforms by genetic means has not yet been evaluated as a potential therapeutic strategy. In our study, we tested whether calpains are overactivated in the MJD context and if reduction or ablation of calpain-1 expression ameliorates the disease-associated phenotype in MJD cells and mice. In all analysed MJD models, we detected an elevated calpain activity at baseline. Lowering or removal of calpain-1 in cells or mice counteracted calpain system overactivation and led to reduced cleavage of ataxin-3 without affecting its aggregation. Moreover, calpain-1 knockout in YAC84Q mice alleviated excessive fragmentation of important synaptic proteins. Despite worsening some motor characteristics, YAC84Q mice showed a rescue of body weight loss and extended survival upon calpain-1 knockout. Together, our findings emphasize the general potential of calpains as a therapeutic target in MJD and other neurodegenerative diseases.
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Azam, Mohammad, Shaida S. Andrabi, Kenneth E. Sahr, Lakshmi Kamath, Athan Kuliopulos, and Athar H. Chishti. "Disruption of the Mouse μ-Calpain Gene Reveals an Essential Role in Platelet Function." Molecular and Cellular Biology 21, no. 6 (March 15, 2001): 2213–20. http://dx.doi.org/10.1128/mcb.21.6.2213-2220.2001.
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ABSTRACT Conventional calpains are ubiquitous calcium-regulated cysteine proteases that have been implicated in cytoskeletal organization, cell proliferation, apoptosis, cell motility, and hemostasis. There are two forms of conventional calpains: the μ-calpain, or calpain I, which requires micromolar calcium for half-maximal activation, and the m-calpain, or calpain II, which functions at millimolar calcium concentrations. We evaluated the functional role of the 80-kDa catalytic subunit of μ-calpain by genetic inactivation using homologous recombination in embryonic stem cells. The μ-calpain-deficient mice are viable and fertile. The complete deficiency of μ-calpain causes significant reduction in platelet aggregation and clot retraction but surprisingly the mutant mice display normal bleeding times. No detectable differences were observed in the cleavage pattern and kinetics of calpain substrates such as the β3 subunit of αIIbβ3 integrin, talin, and ABP-280 (filamin). However, μ-calpain null platelets exhibit impaired tyrosine phosphorylation of several proteins including the β3 subunit of αIIbβ3 integrin, correlating with the agonist-induced reduction in platelet aggregation. These results provide the first direct evidence that μ-calpain is essential for normal platelet function, not by affecting the cleavage of cytoskeletal proteins but by potentially regulating the state of tyrosine phosphorylation of the platelet proteins.
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Schultz, Bruna, Jéssica Taday, Leonardo Menezes, Anderson Cigerce, Marina C. Leite, and Carlos-Alberto Gonçalves. "Calpain-Mediated Alterations in Astrocytes Before and During Amyloid Chaos in Alzheimer’s Disease." Journal of Alzheimer's Disease 84, no. 4 (December 7, 2021): 1415–30. http://dx.doi.org/10.3233/jad-215182.
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One of the changes found in the brain in Alzheimer’s disease (AD) is increased calpain, derived from calcium dysregulation, oxidative stress, and/or neuroinflammation, which are all assumed to be basic pillars in neurodegenerative diseases. The role of calpain in synaptic plasticity, neuronal death, and AD has been discussed in some reviews. However, astrocytic calpain changes sometimes appear to be secondary and consequent to neuronal damage in AD. Herein, we explore the possibility of calpain-mediated astroglial reactivity in AD, both preceding and during the amyloid phase. We discuss the types of brain calpains but focus the review on calpains 1 and 2 and some important targets in astrocytes. We address the signaling involved in controlling calpain expression, mainly involving p38/mitogen-activated protein kinase and calcineurin, as well as how calpain regulates the expression of proteins involved in astroglial reactivity through calcineurin and cyclin-dependent kinase 5. Throughout the text, we have tried to provide evidence of the connection between the alterations caused by calpain and the metabolic changes associated with AD. In addition, we discuss the possibility that calpain mediates amyloid-β clearance in astrocytes, as opposed to amyloid-β accumulation in neurons.
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Liu, Xiuli, Juanita J. Rainey, Jay F. Harriman, and Rick G. Schnellmann. "Calpains mediate acute renal cell death: role of autolysis and translocation." American Journal of Physiology-Renal Physiology 281, no. 4 (October 1, 2001): F728—F738. http://dx.doi.org/10.1152/ajprenal.2001.281.4.f728.
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The goals of this study were to determine 1) the expression of calpain isoforms in rabbit renal proximal tubules (RPT); 2) calpain autolysis and translocation, and calpastatin levels during RPT injury; and 3) the effect of a calpain inhibitor (PD-150606) on calpain levels, mitochondrial function, and ion transport during RPT injury. RT-PCR, immunoblot analysis, and FITC-casein zymography demonstrated the presence of only μ- and m-calpains in rabbit RPT. The mitochondrial inhibitor antimycin A decreased RPT μ- and m-calpain and calpastatin levels in conjunction with cell death and increased plasma membrane permeability. No increases in either μ- or m-calpain were observed in the membrane nor were increases observed in autolytic forms of either μ- or m-calpain in antimycin A-exposed RPT. PD-150606 blocked antimycin A-induced cell death, preserved calpain levels in antimycin A-exposed RPT, and promoted the recovery of mitochondrial function and active Na+ transport in RPT after hypoxia and reoxygenation. The present study suggests that calpains mediate RPT injury without undergoing autolysis or translocation, and ultimately they leak from cells subsequent to RPT injury/death. Furthermore, PD-150606 allows functional recovery after injury.
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Khoutorsky, Arkady, and Micha E. Spira. "Calpain Inhibitors Alter the Excitable Membrane Properties of Cultured Aplysia Neurons." Journal of Neurophysiology 100, no. 5 (November 2008): 2784–93. http://dx.doi.org/10.1152/jn.90487.2008.
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The calpain superfamily of calcium-dependent papain-like cysteine proteases constitutes highly conserved proteases that function to posttranslationally modify substrates by partial proteolysis. Calpains are known to proteolyze >100 substrates that lack strong sequence homology. Consequently, the calpain superfamily has been implicated in playing a central role in diverse physiological and pathological processes. Investigation of the physiological functions of calpains, on the one hand, and the need to develop pharmacological reagents to inhibit calpain-mediated pathological processes, on the other hand, led to the development of numerous calpain inhibitors. Using cultured Aplysia neurons and voltage-clamp analysis, we report here that the calpain inhibitors calpeptin, MG132, and the calpain inhibitor XII inhibit voltage-gated potassium conductance and moderately reduce the sodium conductance. These consequently lead to spike broadening and increased calcium influx. Such alterations of the excitable membrane properties may alter the normal patterns of neuronal and muscle electrical activities and thus should be taken into account when evaluating the effects of calpain inhibitors as protective/therapeutic drugs and as research tools.
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Ilian, M. A., and N. E. Forsberg. "Gene expression of calpains and their specific endogenous inhibitor, calpastatin, in skeletal muscle of fed and fasted rabbits." Biochemical Journal 287, no. 1 (October 1, 1992): 163–71. http://dx.doi.org/10.1042/bj2870163.
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To investigate the role of calpains in myofibrillar protein degradation in skeletal muscle and the regulation of their activity in vivo, we studied the effects of fasting on gene expression of calpains and calpastatin in the skeletal muscle of rabbits. In response to fasting, myofibrillar protein degradation increased 2-fold and mRNA levels of calpain I, calpain II and calpastatin were also increased. However, calpain and calpastatin activities remained unchanged. To investigate this discrepancy, we analysed polysomal calpain mRNA. Results indicated that fasting caused a 2-fold increase in the loading of calpain I and II mRNAs on ribosomes. Thus transcription of genes encoding calpain may be increased during fasting to ensure adequate synthesis of the proteinases needed to mobilize muscle protein reserves. The effect of fasting on calpain and calpastatin mRNA expression is shared by cathepsin D and proteasome C2 but not by beta-actin, implying that fasting invokes control of several proteolytic systems in skeletal muscle and underscores the possibility that each proteolytic system plays a role in the adaptation of skeletal muscle to the fasted state.
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Ben-Aharon, Irit, Paula R. Brown, Nir Etkovitz, Edward M. Eddy, and Ruth Shalgi. "The expression of calpain 1 and calpain 2 in spermatogenic cells and spermatozoa of the mouse." Reproduction 129, no. 4 (April 2005): 435–42. http://dx.doi.org/10.1530/rep.1.00255.
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There is some evidence suggesting that Ca2+is involved in processes that occur during the development and function of spermatozoa. Calcium-dependent proteins, such as calmodulin, are expressed during mammalian spermatogenesis further suggesting that Ca2+takes part in its regulation. However, the precise roles of Ca2+in spermatogenesis remain to be elucidated. Calpains are a family of Ca2+-dependent cysteine proteases whose members are expressed ubiquitously or in a tissue-specific manner. Calpain has been demonstrated to mediate specific Ca2+-dependent processes including cell fusion, mitosis and meiosis. We herein followed the expression pattern of calpain’s ubiquitous isoforms, 1 and 2, throughout spermatogenesis at the RNA and protein levels by RT-PCR and Western blotting analysis. Both RNA and protein studies revealed that these isoforms are expressed in all spermatogenic cells. The expression of calpain 1 levels is slightly higher in spermatocytes entering the meiotic phase. Both calpain isoforms are also expressed in mouse spermatozoa and are localized to the acrosomal cap. Inducing capacitated spermatozoa to undergo the acrosome reaction in the presence of a selective calpain inhibitor significantly reduced the acrosome reaction rate in a dose-dependent manner. Thus, calpain, a pluripotential protease with numerous substrates, may serve as an effector in more than one pathway in the complex process of spermatogenesis and in the events preceding fertilization, such as the acrosome reaction.
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Nina S. Pestereva, Irina S. Ivleva, Irina M. Kotova, Dmitriy S. Traktirov, and Marina N. Karpenko. "Region-specific changes in expression and activity of calpains in the CNS of native rats." Biomedicine 42, no. 4 (September 12, 2022): 771–74. http://dx.doi.org/10.51248/.v42i4.1653.
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Introduction and Aim: It has been proposed that µ-calpain is responsible for neuronal survival, while m-calpain – for the degeneration. It can be assumed that the "susceptibility" to the damage factor for neurons in different CNS regions depends on the content/activity of calpain isoforms. We analyzed the mRNA levels and the activity of µ-and m-calpain in the different CNS structures of rats. Materials and Methods: After decapitation intact male Wistar rats the prefrontal cortex, striatum, hippocampus, midbrain, brainstem, cerebellum, and spinal cord were removed. Each structure was divided into two parts: casein zymography was performed to determine the activity and real-time RT–PCR - to determine the level of expression mRNA of µ-and m-calpains. Results: We have shown that m-calpain mRNA predominates in the striatum, midbrain and brainstem, while µ-calpain mRNA enrichment was noticed for the hippocampus and cerebellum. The highest µ-calpain activity was in the cervical spinal cord, the lowest - in the striatum. The m-calpain activity was relatively high in the midbrain, striatum, hippocampus and brainstem, while in the cervical spinal cord and cerebellum it was moderate. Conclusion: The selective neuronal death observed during neurodegeneration can be partially determined by the initial level of calpains expression and/or activity.
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Seremwe, Mutsa, Rick G. Schnellmann, and Wendy B. Bollag. "Calpain-10 Activity Underlies Angiotensin II-Induced Aldosterone Production in an Adrenal Glomerulosa Cell Model." Endocrinology 156, no. 6 (June 1, 2015): 2138–49. http://dx.doi.org/10.1210/en.2014-1866.
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Abstract Aldosterone is a steroid hormone important in the regulation of blood pressure. Aberrant production of aldosterone results in the development and progression of diseases including hypertension and congestive heart failure; therefore, a complete understanding of aldosterone production is important for developing more effective treatments. Angiotensin II (AngII) regulates steroidogenesis, in part through its ability to increase intracellular calcium levels. Calcium can activate calpains, proteases classified as typical or atypical based on the presence or absence of penta-EF-hands, which are involved in various cellular responses. We hypothesized that calpain, in particular calpain-10, is activated by AngII in adrenal glomerulosa cells and underlies aldosterone production. Our studies showed that pan-calpain inhibitors reduced AngII-induced aldosterone production in 2 adrenal glomerulosa cell models, primary bovine zona glomerulosa and human adrenocortical carcinoma (HAC15) cells, as well as CYP11B2 expression in the HAC15 cells. Although AngII induced calpain activation in these cells, typical calpain inhibitors had no effect on AngII-elicited aldosterone production, suggesting a lack of involvement of classical calpains in this process. However, an inhibitor of the atypical calpain, calpain-10, decreased AngII-induced aldosterone production. Consistent with this result, small interfering RNA (siRNA)-mediated knockdown of calpain-10 inhibited aldosterone production and CYP11B2 expression, whereas adenovirus-mediated overexpression of calpain-10 resulted in increased AngII-induced aldosterone production. Our results indicate that AngII-induced activation of calpain-10 in glomerulosa cells underlies aldosterone production and identify calpain-10 or its downstream pathways as potential targets for the development of drug therapies for the treatment of hypertension.
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Zhang, Mengxiao, Grace Wang, and Tianqing Peng. "Calpain-Mediated Mitochondrial Damage: An Emerging Mechanism Contributing to Cardiac Disease." Cells 10, no. 8 (August 8, 2021): 2024. http://dx.doi.org/10.3390/cells10082024.
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Calpains belong to the family of calcium-dependent cysteine proteases expressed ubiquitously in mammals and many other organisms. Activation of calpain is observed in diseased hearts and is implicated in cardiac cell death, hypertrophy, fibrosis, and inflammation. However, the underlying mechanisms remain incompletely understood. Recent studies have revealed that calpains target and impair mitochondria in cardiac disease. The objective of this review is to discuss the role of calpains in mediating mitochondrial damage and the underlying mechanisms, and to evaluate whether targeted inhibition of mitochondrial calpain is a potential strategy in treating cardiac disease. We expect to describe the wealth of new evidence surrounding calpain-mediated mitochondrial damage to facilitate future mechanistic studies and therapy development for cardiac disease.
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ELCE, John S., Peter L. DAVIES, Carol HEGADORN, Donald H. MAURICE, and J. Simon C. ARTHUR. "The effects of truncations of the small subunit on m-calpain activity and heterodimer formation." Biochemical Journal 326, no. 1 (August 15, 1997): 31–38. http://dx.doi.org/10.1042/bj3260031.
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In order to study subunit interactions in calpain, the effects of small subunit truncations on m-calpain activity and heterodimer formation have been measured. It has been shown previously that active calpain is formed by co-expression of the large subunit (80 kDa) of rat m-calpain with a δ86 form (21 kDa) of the small subunit. cDNA for the full-length 270 amino acid (28.5 kDa) rat calpain small subunit has now been cloned, both with and without an N-terminal histidine tag (NHis10). The full-length small subunit constructs yielded active calpains on co-expression with the large subunit, and the small subunit was autolysed to 20 kDa on exposure of these calpains to Ca2+. A series of deletion mutants of the small subunit, NHis10-δ86,-δ99, -δ107, and -δ116, gave active heterodimeric calpains with unchanged specific activities, although in decreasing yield, and with a progressive decrease in stability. NHis10-δ125 formed a heterodimer which was inactive and unstable. Removal of 25 C-terminal residues from δ86, leaving residues 87–245, abolished both activity and heterodimer formation. The results show that: (a) generation of active m-calpain in Escherichia coli requires heterodimer formation; (b) small subunit residues between 94 and 116 contribute to the stability of the active heterodimer but do not directly affect the catalytic mechanism; (c) residues in the region 245–270 are essential for subunit binding. Finally, it was shown that an inactive mutant Cys105 → Ser-80k/δ86 calpain, used in order to preclude autolysis, did not dissociate in the presence of Ca2+, a result which does not support the proposal that Ca2+-induced dissociation is involved in calpain activation.
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Piper, Ann-Katrin, Reece A. Sophocleous, Samuel E. Ross, Frances J. Evesson, Omar Saleh, Adam Bournazos, Joe Yasa, et al. "Loss of calpains-1 and -2 prevents repair of plasma membrane scrape injuries, but not small pores, and induces a severe muscular dystrophy." American Journal of Physiology-Cell Physiology 318, no. 6 (June 1, 2020): C1226—C1237. http://dx.doi.org/10.1152/ajpcell.00408.2019.
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The ubiquitous calpains, calpain-1 and -2, play important roles in Ca2+-dependent membrane repair. Mechanically active tissues like skeletal muscle are particularly reliant on mechanisms to repair and remodel membrane injury, such as those caused by eccentric damage. We demonstrate that calpain-1 and -2 are master effectors of Ca2+-dependent repair of mechanical plasma membrane scrape injuries, although they are dispensable for repair/removal of small wounds caused by pore-forming agents. Using CRISPR gene-edited human embryonic kidney 293 (HEK293) cell lines, we established that loss of both calpains-1 and -2 ( CAPNS1−/−) virtually ablates Ca2+-dependent repair of mechanical scrape injuries but does not affect injury or recovery from perforation by streptolysin-O or saponin. In contrast, cells with targeted knockout of either calpain-1 ( CAPN1−/−) or -2 ( CAPN2−/−) show near-normal repair of mechanical injuries, inferring that both calpain-1 and calpain-2 are equally capable of conducting the cascade of proteolytic cleavage events to reseal a membrane injury, including that of the known membrane repair agent dysferlin. A severe muscular dystrophy in a murine model with skeletal muscle knockout of Capns1 highlights vital roles for calpain-1 and/or -2 for health and viability of skeletal muscles not compensated for by calpain-3 ( CAPN3). We propose that the dystrophic phenotype relates to loss of maintenance of plasma membrane/cytoskeletal networks by calpains-1 and -2 in response to directed and dysfunctional Ca2+-signaling, pathways hyperstimulated in the context of membrane injury. With CAPN1 variants associated with spastic paraplegia, a severe dystrophy observed with muscle-specific loss of calpain-1 and -2 activity identifies CAPN2 and CAPNS1 as plausible candidate neuromuscular disease genes.
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Blanc, Fany, Laetitia Furio, Dorothée Moisy, Hui-Ling Yen, Michel Chignard, Emmanuel Letavernier, Nadia Naffakh, Chris Ka Pun Mok, and Mustapha Si-Tahar. "Targeting host calpain proteases decreases influenza A virus infection." American Journal of Physiology-Lung Cellular and Molecular Physiology 310, no. 7 (April 1, 2016): L689—L699. http://dx.doi.org/10.1152/ajplung.00314.2015.
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Influenza A viruses (IAV) trigger contagious acute respiratory diseases. A better understanding of the molecular mechanisms of IAV pathogenesis and host immune responses is required for the development of more efficient treatments of severe influenza. Calpains are intracellular proteases that participate in diverse cellular responses, including inflammation. Here, we used in vitro and in vivo approaches to investigate the role of calpain signaling in IAV pathogenesis. Calpain expression and activity were found altered in IAV-infected bronchial epithelial cells. With the use of small-interfering RNA (siRNA) gene silencing, specific synthetic inhibitors of calpains, and mice overexpressing calpastatin, we found that calpain inhibition dampens IAV replication and IAV-triggered secretion of proinflammatory mediators and leukocyte infiltration. Remarkably, calpain inhibition has a protective impact in IAV infection, since it significantly reduced mortality of mice challenged not only by seasonal H3N2- but also by hypervirulent H5N1 IAV strains. Hence, our study suggests that calpains are promising therapeutic targets for treating IAV acute pneumonia.
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Kuchay, Shafi, Rafael Nunez, Amelia M. Bartholomew, and Athar H. Chishti. "Calpain I Null Mice Display Lymphoid Hyperplasia." Blood 104, no. 11 (November 16, 2004): 1268. http://dx.doi.org/10.1182/blood.v104.11.1268.1268.
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Abstract Calpains are ubiquitous calcium-regulated cysteine proteases that have been implicated in cytoskeletal organization, homeostasis, cell proliferation, cell motility, and apoptosis. The conventional calpains include mu-calpain (calpain I) that requires micromolar calcium for half maximal activation and m-calpain (calpain II) that functions at millimolar calcium concentration in vitro. To evaluate the physiological function(s) of mu-calpain, we utilized genetically altered mu-calpain null mice as the model system. Spleen biopsy of mu-calpain null adult mice showed consistent expansion of white pulp (lymphoid hyperplasia) due to an increase in number and size of follicles and PALS (periarteriolar lymphatic sheets), relative to matched wild type mice. Also, cervical lymph nodes of mu-calpain null mice showed lymphoid hyperplasia and marked plasmacytosis. The majority of mu-calpain null mice displayed multi-organ lymphoid infiltration. Using immunofluorescence labeling and multiparameter flow cytometry, the cellular composition of the hematopoietic compartment was quantified in both bone marrow and spleen of mu-calpain null mice. The mu-calpain null mice exhibited an overall increase in the number of B-cell lineage; showing ~40% increase of B220+ cells in the bone marrow and ~19% increase of B220+ cells in the spleen. Interestingly, T-cell compartment (CD3+, CD4, CD8+) and NK lineage (NK1.1+) cells were significantly reduced. Myeloid specific GR1+, CD11b+ granulocytic cell lineage, and megakaryocytic CD41+ cells were also noticeably increased in the bone marrow of mu-calpain null mice. Together, these results reveal a central role of mu-calpain in the maintenance of lymphoid homeostasis, contributing to the regulation of B-cell lineage. Ongoing experiments involve investigating quantitative effects of mu-calpain deletion on the survival and apoptotic cell death of B220+ B-cell lineage in response to various insults such as Fas ligand, cycloheximide, irradiation, 9-AC, and staurosporine. The outcome of these studies will elucidate the precise biochemical function(s) of mu-calpain by identifying key substrates that regulate apoptosis in a physiologically relevant environment.
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Sorimachi, Hiroyuki, Hiroshi Mamitsuka, and Yasuko Ono. "Understanding the substrate specificity of conventional calpains." Biological Chemistry 393, no. 9 (September 1, 2012): 853–71. http://dx.doi.org/10.1515/hsz-2012-0143.
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Abstract: Calpains are intracellular Ca2+-dependent Cys proteases that play important roles in a wide range of biological phenomena via the limited proteolysis of their substrates. Genetic defects in calpain genes cause lethality and/or functional deficits in many organisms, including humans. Despite their biological importance, the mechanisms underlying the action of calpains, particularly of their substrate specificities, remain largely unknown. Studies show that certain sequence preferences influence calpain substrate recognition, and some properties of amino acids have been related successfully to substrate specificity and to the calpains’ 3D structure. The full spectrum of this substrate specificity, however, has not been clarified using standard sequence analysis algorithms, e.g., the position-specific scoring-matrix method. More advanced bioinformatics techniques were used recently to identify the substrate specificities of calpains and to develop a predictor for calpain cleavage sites, demonstrating the potential of combining empirical data acquisition and machine learning. This review discusses the calpains’ substrate specificities, introducing the benefits of bioinformatics applications. In conclusion, machine learning has led to the development of useful predictors for calpain cleavage sites, although the accuracy of the predictions still needs improvement. Machine learning has also elucidated information about the properties of calpains’ substrate specificities, including a preference for sequences over secondary structures and the existence of a substrate specificity difference between two similar conventional calpains, which has never been indicated biochemically.
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Ou, B. R., and N. E. Forsberg. "Determination of skeletal muscle calpain and calpastatin activities during maturation." American Journal of Physiology-Endocrinology and Metabolism 261, no. 6 (December 1, 1991): E677—E683. http://dx.doi.org/10.1152/ajpendo.1991.261.6.e677.
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Our objectives were to characterize events underlying changes in skeletal muscle calpain and calpastatin activities, using maturation as a model. Muscle samples were taken from rabbits of four ages (newborn and 1, 2, and 5 mo old). Concentrations of RNA and protein and activities of calpains I and II and calpastatin were determined. Steady-state concentrations of mRNAs encoding calpain I, calpain II, calpastatin, alpha- and beta-tubulin, and beta-actin were determined using Northern blot analysis. Calpain and calpastatin activities declined markedly between birth and 1 mo of age and remained unchanged thereafter. Several factors accounted for the neonatal losses of calpains and calpastatin. First, muscle protein concentration increased between birth and 1 mo of age and diluted calpain and calpastatin specific activities. Second, there was a marked reduction of muscle RNA concentration between birth and 1 mo of age, which indicates that protein synthetic capacity declined with age. Finally, calpastatin mRNA concentration declined between birth and 1 mo of age and further contributed to developmental losses of calpastatin activity. Calpain I mRNA concentration was unaffected by age, and although calpain II mRNA concentration declined with age, losses were not detected between birth and 1 mo; hence age-related changes in calpain I and II activities are not mediated at the mRNA level. The age-related reductions in calpain II and calpastatin mRNA concentrations resembled age-related changes in alpha- and beta-tubulin and beta-actin mRNA concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
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Bevers, Matthew B., and Robert W. Neumar. "Mechanistic Role of Calpains in Postischemic Neurodegeneration." Journal of Cerebral Blood Flow & Metabolism 28, no. 4 (December 12, 2007): 655–73. http://dx.doi.org/10.1038/sj.jcbfm.9600595.
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The calpain family of proteases is causally linked to postischemic neurodegeneration. However, the precise mechanisms by which calpains contribute to postischemic neuronal death have not been fully elucidated. This review outlines the key features of the calpain system, and the evidence for its causal role in postischemic neuronal pathology. Furthermore, the consequences of specific calpain substrate cleavage at various subcellular locations are explored. Calpain substrates within synapses, plasma membrane, endoplasmic reticulum, lysosomes, mitochondria, and the nucleus, as well as the overall effect of postischemic calpain activity on calcium regulation and cell death signaling are considered. Finally, potential pathways for calpain-mediated neurodegeneration are outlined in an effort to guide future studies aimed at understanding the downstream pathology of postischemic calpain activity and identifying optimal therapeutic strategies.
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McCartney, Christian-Scott E., Qilu Ye, Robert L. Campbell, and Peter L. Davies. "Insertion sequence 1 from calpain-3 is functional in calpain-2 as an internal propeptide." Journal of Biological Chemistry 293, no. 46 (September 25, 2018): 17716–30. http://dx.doi.org/10.1074/jbc.ra118.004803.
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Calpains are intracellular, calcium-activated cysteine proteases. Calpain-3 is abundant in skeletal muscle, where its mutation-induced loss of function causes limb-girdle muscular dystrophy type 2A. Unlike the small subunit–containing calpain-1 and -2, the calpain-3 isoform homodimerizes through pairing of its C-terminal penta-EF-hand domain. It also has two unique insertion sequences (ISs) not found in the other calpains: IS1 within calpain-3's protease core and IS2 just prior to the penta-EF-hand domain. Production of either native or recombinant full-length calpain-3 to characterize the function of these ISs is challenging. Therefore, here we used recombinant rat calpain-2 as a stable surrogate and inserted IS1 into its equivalent position in the protease core. As it does in calpain-3, IS1 occupied the catalytic cleft and restricted the enzyme's access to substrate and inhibitors. Following activation by Ca2+, IS1 was rapidly cleaved by intramolecular autolysis, permitting the enzyme to freely accept substrate and inhibitors. The surrogate remained functional until extensive intermolecular autoproteolysis inactivated the enzyme, as is typical of calpain-2. Although the small-molecule inhibitors E-64 and leupeptin limited intermolecular autolysis of the surrogate, they did not block the initial intramolecular cleavage of IS1, establishing its role as a propeptide. Surprisingly, the large-molecule calpain inhibitor, calpastatin, completely blocked enzyme activity, even with IS1 intact. We suggest that calpastatin is large enough to oust IS1 from the catalytic cleft and take its place. We propose an explanation for why calpastatin can inhibit calpain-2 bearing the IS1 insertion but cannot inhibit WT calpain-3.
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Baudry, Michel. "Calpain-1 and Calpain-2 in the Brain: Dr. Jekill and Mr Hyde?" Current Neuropharmacology 17, no. 9 (August 22, 2019): 823–29. http://dx.doi.org/10.2174/1570159x17666190228112451.
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While the calpain system has now been discovered for over 50 years, there is still a paucity of information regarding the organization and functions of the signaling pathways regulated by these proteases, although calpains play critical roles in many cell functions. Moreover, calpain overactivation has been shown to be involved in numerous diseases. Among the 15 calpain isoforms identified, calpain-1 (aka µ-calpain) and calpain-2 (aka m-calpain) are ubiquitously distributed in most tissues and organs, including the brain. We have recently proposed that calpain-1 and calpain- 2 play opposite functions in the brain, with calpain-1 activation being required for triggering synaptic plasticity and neuroprotection (Dr. Jekill), and calpain-2 limiting the extent of plasticity and being neurodegenerative (Mr. Hyde). Calpain-mediated cleavage has been observed in cytoskeleton proteins, membrane-associated proteins, receptors/channels, scaffolding/anchoring proteins, and protein kinases and phosphatases. This review will focus on the signaling pathways related to local protein synthesis, cytoskeleton regulation and neuronal survival/death regulated by calpain-1 and calpain-2, in an attempt to explain the origin of the opposite functions of these 2 calpain isoforms. This will be followed by a discussion of the potential therapeutic applications of selective regulators of these 2 calpain isoforms.
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Moraczewski, J., E. Piekarska, M. Zimowska, and M. Sobolewska. "Activity of mu- and m-calpain in regenerating fast and slow twitch skeletal muscles." Acta Biochimica Polonica 43, no. 4 (December 31, 1996): 693–700. http://dx.doi.org/10.18388/abp.1996_4466.
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Calpains--non-lysosomal intracellular calcium-activated neutral proteinases, form a family consisting of several distinct members. Two of the isoenzymes: mu (calpain I) and m (calpain II) responded differently to the injury during complete regeneration of Extensor digitorum longus (EDL) muscle and partial regeneration of Soleus muscle. In the crushed EDL the level of m-calpain on the 3rd and 7th day of regeneration was higher than in non-operated muscles, whereas the activity of this calpain in injured Soleus decreased. The level of mu-calpain in EDL oscillated irregularly during regeneration whereas in Soleus of both injured and contralateral muscles its level rapidly rose. Our results support the hypothesis that m-calpain is involved in the process of fusion of myogenic cells whereas mu-calpain plays a significant but indirect role in muscle regeneration.
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Kashiwagi, Aki, Ernestina Schipani, Mikaela J. Fein, Peter A. Greer, and Masako Shimada. "Targeted Deletion of Capn4 in Cells of the Chondrocyte Lineage Impairs Chondrocyte Proliferation and Differentiation." Molecular and Cellular Biology 30, no. 11 (April 5, 2010): 2799–810. http://dx.doi.org/10.1128/mcb.00157-10.
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ABSTRACT Calpains are calcium-dependent intracellular cysteine proteases, which include ubiquitously expressed μ- and m-calpains. Both calpains are heterodimers consisting of a large catalytic subunit and a small regulatory subunit. The calpain small subunit encoded by the gene Capn4 directly binds to the intracellular C-terminal tail of the receptor for the parathyroid hormone (PTH) and PTH-related peptide and modulates cellular functions in cells of the osteoblast lineage in vitro and in vivo. To investigate a physiological role of the calpain small subunit in cells of the chondrocyte lineage, we generated chondrocyte-specific Capn4 knockout mice. Mutant embryos had reduced chondrocyte proliferation and differentiation in embryonic growth plates compared with control littermates. In vitro analysis further revealed that deletion of Capn4 in cells of the chondrocyte lineage correlated with impaired cell cycle progression at the G1/S transition, reduced cyclin D gene transcription, and accumulated cell cycle proteins known as calpain substrates. Moreover, silencing of p27Kip1 rescued an impaired cell growth phenotype in Capn4 knockdown cells, and reintroducing the calpain small subunit partially normalized cell growth and accumulated cyclin D protein levels in a dose-dependent manner. Collectively, our findings suggest that the calpain small subunit is essential for proper chondrocyte functions in embryonic growth plates.
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Moshal, Karni S., Mahavir Singh, Utpal Sen, Dorothea Susanne E. Rosenberger, Brooke Henderson, Neetu Tyagi, Hong Zhang, and Suresh C. Tyagi. "Homocysteine-mediated activation and mitochondrial translocation of calpain regulates MMP-9 in MVEC." American Journal of Physiology-Heart and Circulatory Physiology 291, no. 6 (December 2006): H2825—H2835. http://dx.doi.org/10.1152/ajpheart.00377.2006.
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Hyperhomocysteinemia (HHcy) is associated with atherosclerosis, stroke, and dementia. Hcy causes extracellular matrix remodeling by the activation of matrix metalloproteinase-9 (MMP-9), in part, by inducing redox signaling and modulating the intracellular calcium dynamics. Calpains are the calcium-dependent cysteine proteases that are implicated in mitochondrial damage via oxidative burst. Mitochondrial abnormalities have been identified in HHcy. The mechanism of Hcy-induced extracellular matrix remodeling by MMP-9 activation via mitochondrial pathway is largely unknown. We report a novel role of calpains in mitochondrial-mediated MMP-9 activation by Hcy in cultured rat heart microvascular endothelial cells. Our observations suggested that calpain regulates Hcy-induced MMP-9 expression and activity. We showed that Hcy activates calpain-1, but not calpain-2, in a calcium-dependent manner. Interestingly, the enhanced calpain activity was not mirrored by the decreased levels of its endogenous inhibitor calpastatin. We presented evidence that Hcy induces the translocation of active calpain from cytosol to mitochondria, leading to MMP-9 activation, in part, by causing intramitochondrial oxidative burst. Furthermore, studies with pharmacological inhibitors of calpain (calpeptin and calpain-1 inhibitor), ERK (PD-98059) and the mitochondrial uncoupler FCCP suggested that calpain and ERK-1/2 are the major events within the Hcy/MMP-9 signal axis and that intramitochondrial oxidative stress regulates MMP-9 via ERK-1/2 signal cascade. Taken together, these findings determine the novel role of mitochondrial translocation of calpain-1 in MMP-9 activation during HHcy, in part, by increasing mitochondrial oxidative tress.
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Ben-Aharon, Irit, Karin Haim, Ruth Shalgi, and Dalit Ben-Yosef. "Expression and possible involvement of calpain isoforms in mammalian egg activation." Reproduction 130, no. 2 (August 2005): 165–75. http://dx.doi.org/10.1530/rep.1.00602.
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At fertilization in mammals, the spermatozoon triggers a unique signal transduction mechanism within the egg, leading to its activation. It is well accepted that the earliest event observed in all activated eggs is an abrupt rise in intracellular calcium concentrations. However, little is known regarding the downstream proteins that are activated by this rise in calcium. Calpains constitute a family of intracellular calcium-dependent cysteine proteases whose members are expressed widely in a variety of cells. We investigated the expression and possible role of the calpain isoforms μ and m throughout egg activation. Both calpains were expressed in the rat egg and localized at the egg cortex as well as in the meiotic spindle. m Calpain translocated to the membrane and to the spindle area during parthenogenetic egg activation and duringin vivofertilization, upon sperm binding to the egg. The cytoskeletal protein α-spectrin (fodrin) was proteolysed by calpain during the egg-activation process, as demonstrated by specific calpain-breakdown products. Following parthenogenetic activation by ionomycin or puromycin, the calpain-selective permeable inhibitor, calpeptin, inhibited the resumption of meiosis and cortical reaction in a dosedependent manner. Calpeptin was also effective in inhibitingin vitrofertilization. These results may imply a correlation between calpain activation and mammalian egg activation at fertilization and a possible role for calpain in the cascade of cellular events leading to resumption of meiosis.
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Gafni, Juliette, Evan Hermel, Jessica E. Young, Cheryl L. Wellington, Michael R. Hayden, and Lisa M. Ellerby. "Inhibition of Calpain Cleavage of Huntingtin Reduces Toxicity." Journal of Biological Chemistry 279, no. 19 (February 23, 2004): 20211–20. http://dx.doi.org/10.1074/jbc.m401267200.
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Huntington's disease (HD) is a neurodegenerative disorder caused by a polyglutamine (polyQ) tract expansion near the N terminus of huntingtin (Htt). Proteolytic processing of mutant Htt and abnormal calcium signaling may play a critical role in disease progression and pathogenesis. Recent work indicates that calpains may participate in the increased and/or altered patterns of Htt proteolysis leading to the selective toxicity observed in HD striatum. Here, we identify two calpain cleavage sites in Htt and show that mutation of these sites renders the polyQ expanded Htt less susceptible to proteolysis and aggregation, resulting in decreased toxicity in anin vitrocell culture model. In addition, we found that calpain- and caspase-derived Htt fragments preferentially accumulate in the nucleus without the requirement of further cleavage into smaller fragments. Calpain family members, calpain-1, -5, -7, and -10, have increased levels or are activated in HD tissue culture and transgenic mouse models, suggesting they may play a key role in Htt proteolysis and disease pathology. Interestingly, calpain-1, -5, -7, and -10 localize to the cytoplasm and the nucleus, whereas the activated forms of calpain-7 and -10 are found only in the nucleus. These results support the role of calpain-derived Htt fragmentation in HD and suggest that aberrant activation of calpains may play a role in HD pathogenesis.
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Wang, Yubin, Yan Liu, Xiaoning Bi, and Michel Baudry. "Calpain-1 and Calpain-2 in the Brain: New Evidence for a Critical Role of Calpain-2 in Neuronal Death." Cells 9, no. 12 (December 16, 2020): 2698. http://dx.doi.org/10.3390/cells9122698.
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Calpains are a family of soluble calcium-dependent proteases that are involved in multiple regulatory pathways. Our laboratory has focused on the understanding of the functions of two ubiquitous calpain isoforms, calpain-1 and calpain-2, in the brain. Results obtained over the last 30 years led to the remarkable conclusion that these two calpain isoforms exhibit opposite functions in the brain. Calpain-1 activation is required for certain forms of synaptic plasticity and corresponding types of learning and memory, while calpain-2 activation limits the extent of plasticity and learning. Calpain-1 is neuroprotective both during postnatal development and in adulthood, while calpain-2 is neurodegenerative. Several key protein targets participating in these opposite functions have been identified and linked to known pathways involved in synaptic plasticity and neuroprotection/neurodegeneration. We have proposed the hypothesis that the existence of different PDZ (PSD-95, DLG and ZO-1) binding domains in the C-terminal of calpain-1 and calpain-2 is responsible for their association with different signaling pathways and thereby their different functions. Results with calpain-2 knock-out mice or with mice treated with a selective calpain-2 inhibitor indicate that calpain-2 is a potential therapeutic target in various forms of neurodegeneration, including traumatic brain injury and repeated concussions.
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GOLL, DARREL E., VALERY F. THOMPSON, HONGQI LI, WEI WEI, and JINYANG CONG. "The Calpain System." Physiological Reviews 83, no. 3 (July 2003): 731–801. http://dx.doi.org/10.1152/physrev.00029.2002.
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Goll, Darrel E., Valery F. Thompson, Hongqi Li, Wei Wei, and Jinyang Cong. The Calpain System. Physiol Rev 83: 731–801, 2003; 10.1152/physrev.00029.2002.—The calpain system originally comprised three molecules: two Ca2+-dependent proteases, μ-calpain and m-calpain, and a third polypeptide, calpastatin, whose only known function is to inhibit the two calpains. Both μ- and m-calpain are heterodimers containing an identical 28-kDa subunit and an 80-kDa subunit that shares 55–65% sequence homology between the two proteases. The crystallographic structure of m-calpain reveals six “domains” in the 80-kDa subunit: 1) a 19-amino acid NH2-terminal sequence; 2) and 3) two domains that constitute the active site, IIa and IIb; 4) domain III; 5) an 18-amino acid extended sequence linking domain III to domain IV; and 6) domain IV, which resembles the penta EF-hand family of polypeptides. The single calpastatin gene can produce eight or more calpastatin polypeptides ranging from 17 to 85 kDa by use of different promoters and alternative splicing events. The physiological significance of these different calpastatins is unclear, although all bind to three different places on the calpain molecule; binding to at least two of the sites is Ca2+dependent. Since 1989, cDNA cloning has identified 12 additional mRNAs in mammals that encode polypeptides homologous to domains IIa and IIb of the 80-kDa subunit of μ- and m-calpain, and calpain-like mRNAs have been identified in other organisms. The molecules encoded by these mRNAs have not been isolated, so little is known about their properties. How calpain activity is regulated in cells is still unclear, but the calpains ostensibly participate in a variety of cellular processes including remodeling of cytoskeletal/membrane attachments, different signal transduction pathways, and apoptosis. Deregulated calpain activity following loss of Ca2+homeostasis results in tissue damage in response to events such as myocardial infarcts, stroke, and brain trauma.
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Ennes-Vidal, Vítor, Marta Helena Branquinha, André Luis Souza dos Santos, and Claudia Masini d’Avila-Levy. "The Diverse Calpain Family in Trypanosomatidae: Functional Proteins Devoid of Proteolytic Activity?" Cells 10, no. 2 (February 1, 2021): 299. http://dx.doi.org/10.3390/cells10020299.
Full textAbstract:
Calpains are calcium-dependent cysteine peptidases that were originally described in mammals and, thereafter, their homologues were identified in almost all known living organisms. The deregulated activity of these peptidases is associated with several pathologies and, consequently, huge efforts have been made to identify selective inhibitors. Trypanosomatids, responsible for life-threatening human diseases, possess a large and diverse family of calpain sequences in their genomes. Considering that the current therapy to treat trypanosomatid diseases is limited to a handful of drugs that suffer from unacceptable toxicity, tough administration routes, like parenteral, and increasing treatment failures, a repurposed approach with calpain inhibitors could be a shortcut to successful chemotherapy. However, there is a general lack of knowledge about calpain functions in these parasites and, currently, the proteolytic activity of these proteins is still an open question. Here, we highlight the current research and perspectives on trypanosomatid calpains, overview calpain description in these organisms, and explore the potential of targeting the calpain system as a therapeutic strategy. This review gathers the current knowledge about this fascinating family of peptidases as well as insights into the puzzle: are we unable to measure calpain activity in trypanosomatids, or are the functions of these proteins devoid of proteolytic activity in these parasites?
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Rose, Aaron, Huang Zhi, Fukun Hoffmann, Robert Norton, and Peter Hoffamann. "Selenoprotein K is a novel target of m-calpain, and cleavage is regulated by toll-like receptor-induced calpastatin in macrophages (54.22)." Journal of Immunology 188, no. 1_Supplement (May 1, 2012): 54.22. http://dx.doi.org/10.4049/jimmunol.188.supp.54.22.
Full textAbstract:
Abstract Calpains are calcium (Ca2+)-activated cysteine proteases that modulate cellular function through cleavage of a variety of intra-cellular targets. Two major isoforms of calpain, u- and m-calpain, are expressed in different tissues and cell-types. Selenoprotein K (SelK) is a selenocysteine containing transmembrane ER protein involved in Ca2+ flux, and western blot analyses revealed one SelK band in mouse lymphocytes (both B and T cells) and two bands for SelK in mouse myeloid cells (macrophages, dendritic cells, and neutrophils). This led us to investigate cleavage of SelK and we found that m-calpain cleaved SelK in myeloid cells. The calpain-mediated cleavage of SelK occured between Arg81 and Gly82 resulting in the removal of the selenocysteine-containing portion of the cytosolic region of the protein. We have further linked this event to the activation state of macrophages. Resting macrophages were found to predominantly contain cleaved SelK while macrophages activated through toll-like receptors contained full length SelK. This was due to increased levels of calpastatin, which is the endogenous inhibitor of calpains. The identification of SelK as a novel target for calpain cleavage activity reveals a novel mechanism by which macrophage activation is regulated and studies are on-going to further characterize the role of calpain/calpastatin in macrophages.
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Arrington, David D., Terry R. Van Vleet, and Rick G. Schnellmann. "Calpain 10: a mitochondrial calpain and its role in calcium-induced mitochondrial dysfunction." American Journal of Physiology-Cell Physiology 291, no. 6 (December 2006): C1159—C1171. http://dx.doi.org/10.1152/ajpcell.00207.2006.
Full textAbstract:
Calpains, Ca2+-activated cysteine proteases, are cytosolic enzymes implicated in numerous cellular functions and pathologies. We identified a mitochondrial Ca2+-inducible protease that hydrolyzed a calpain substrate (SLLVY-AMC) and was inhibited by active site-directed calpain inhibitors as calpain 10, an atypical calpain lacking domain IV. Immunoblot analysis and activity assays revealed calpain 10 in the mitochondrial outer membrane, intermembrane space, inner membrane, and matrix fractions. Mitochondrial staining was observed when COOH-terminal green fluorescent protein-tagged calpain 10 was overexpressed in NIH-3T3 cells and the mitochondrial targeting sequence was localized to the NH2-terminal 15 amino acids. Overexpression of mitochondrial calpain 10 resulted in mitochondrial swelling and autophagy that was blocked by the mitochondrial permeability transition (MPT) inhibitor cyclosporine A. With the use of isolated mitochondria, Ca2+-induced MPT was partially decreased by calpain inhibitors. More importantly, Ca2+-induced inhibition of Complex I of the electron transport chain was blocked by calpain inhibitors and two Complex I proteins were identified as targets of mitochondrial calpain 10, NDUFV2, and ND6. In conclusion, calpain 10 is the first reported mitochondrially targeted calpain and is a mediator of mitochondrial dysfunction through the cleavage of Complex I subunits and activation of MPT.
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Potter, David A., Jennifer S. Tirnauer, Richard Janssen, Dorothy E. Croall, Christina N. Hughes, Kerry A. Fiacco, James W. Mier, Masatoshi Maki, and Ira M. Herman. "Calpain Regulates Actin Remodeling during Cell Spreading." Journal of Cell Biology 141, no. 3 (May 4, 1998): 647–62. http://dx.doi.org/10.1083/jcb.141.3.647.
Full textAbstract:
Previous studies suggest that the Ca2+-dependent proteases, calpains, participate in remodeling of the actin cytoskeleton during wound healing and are active during cell migration. To directly test the role that calpains play in cell spreading, several NIH-3T3– derived clonal cell lines were isolated that overexpress the biological inhibitor of calpains, calpastatin. These cells stably overexpress calpastatin two- to eightfold relative to controls and differ from both parental and control cell lines in morphology, spreading, cytoskeletal structure, and biochemical characteristics. Morphologic characteristics of the mutant cells include failure to extend lamellipodia, as well as abnormal filopodia, extensions, and retractions. Whereas wild-type cells extend lamellae within 30 min after plating, all of the calpastatin-overexpressing cell lines fail to spread and assemble actin-rich processes. The cells genetically altered to overexpress calpastatin display decreased calpain activity as measured in situ or in vitro. The ERM protein ezrin, but not radixin or moesin, is markedly increased due to calpain inhibition. To confirm that inhibition of calpain activity is related to the defect in spreading, pharmacological inhibitors of calpain were also analyzed. The cell permeant inhibitors calpeptin and MDL 28, 170 cause immediate inhibition of spreading. Failure of the intimately related processes of filopodia formation and lamellar extension indicate that calpain is intimately involved in actin remodeling and cell spreading.
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Kalbe, L., A. Leunda, T. Sparre, C. Meulemans, M. T. Ahn, T. Orntoft, M. Kruhoffer, B. Reusens, J. Nerup, and C. Remacle. "Nutritional regulation of proteases involved in fetal rat insulin secretion and islet cell proliferation." British Journal of Nutrition 93, no. 3 (March 2005): 309–16. http://dx.doi.org/10.1079/bjn20041313.
Full textAbstract:
Epidemiological studies have indicated that malnutrition during early life may programme chronic degenerative disease in adulthood. In an animal model of fetal malnutrition, rats received an isoenergetic, low-protein (LP) diet during gestation. This reduced fetal β-cell proliferation and insulin secretion. Supplementation during gestation with taurine prevented these alterations. Since proteases are involved in secretion and proliferation, we investigated which proteases were associated with these alterations and their restoration in fetal LP islets. Insulin secretion and proliferation of fetal control and LP islets exposed to different protease modulators were measured. Lactacystin and calpain inhibitor I, but not isovaleryl-l-carnitine, raised insulin secretion in control islets, indicating that proteasome and cysteinyl cathepsin(s), but not μ-calpain, are involved in fetal insulin secretion. Insulin secretion from LP islets responded normally to lactacystin but was insensitive to calpain inhibitor I, indicating a loss of cysteinyl cathepsin activity. Taurine supplementation prevented this by restoring the response to calpain inhibitor I. Control islet cell proliferation was reduced by calpain inhibitor I and raised by isovaleryl-l-carnitine, indicating an involvement of calpain. Calpain activity appeared to be lost in LP islets and not restored by taurine. Most modifications in the mRNA expression of cysteinyl cathepsins, calpains and calpastatin due to maternal protein restriction were consistent with reduced protease activity and were restored by taurine. Thus, maternal protein restriction affected cysteinyl cathepsins and the calpain–calpastatin system. Taurine normalised fetal LP insulin secretion by protecting cysteinyl cathepsin(s), but the restoration of LP islet cell proliferation by taurine did not implicate calpains.
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Nakagawa, Toshiyuki, and Junying Yuan. "Cross-Talk between Two Cysteine Protease Families." Journal of Cell Biology 150, no. 4 (August 21, 2000): 887–94. http://dx.doi.org/10.1083/jcb.150.4.887.
Full textAbstract:
Calpains and caspases are two cysteine protease families that play important roles in regulating pathological cell death. Here, we report that m-calpain may be responsible for cleaving procaspase-12, a caspase localized in the ER, to generate active caspase-12. In addition, calpain may be responsible for cleaving the loop region in Bcl-xL and, therefore, turning an antiapoptotic molecule into a proapoptotic molecule. We propose that disturbance to intracellular calcium storage as a result of ischemic injury or amyloid β peptide cytotoxicity may induce apoptosis through calpain- mediated caspase-12 activation and Bcl-xL inactivation. These data suggest a novel apoptotic pathway involving calcium-mediated calpain activation and cross-talks between calpain and caspase families.
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Belhadj, Soumaya, Nina Sofia Hermann, Yu Zhu, Gustav Christensen, Torsten Strasser, and François Paquet-Durand. "Visualizing Cell Death in Live Retina: Using Calpain Activity Detection as a Biomarker for Retinal Degeneration." International Journal of Molecular Sciences 23, no. 7 (March 31, 2022): 3892. http://dx.doi.org/10.3390/ijms23073892.
Full textAbstract:
Calpains are a family of calcium-activated proteases involved in numerous disorders. Notably, previous studies have shown that calpain activity was substantially increased in various models for inherited retinal degeneration (RD). In the present study, we tested the capacity of the calpain-specific substrate t-BOC-Leu-Met-CMAC to detect calpain activity in living retina, in organotypic retinal explant cultures derived from wild-type mice, as well as from rd1 and RhoP23H/+ RD-mutant mice. Test conditions were refined until the calpain substrate readily detected large numbers of cells in the photoreceptor layer of RD retina but not in wild-type retina. At the same time, the calpain substrate was not obviously toxic to photoreceptor cells. Comparison of calpain activity with immunostaining for activated calpain-2 furthermore suggested that individual calpain isoforms may be active in distinct temporal stages of photoreceptor cell death. Notably, calpain-2 activity may be a relatively short-lived event, occurring only towards the end of the cell-death process. Finally, our results support the development of calpain activity detection as a novel in vivo biomarker for RD suitable for combination with non-invasive imaging techniques.
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Wang, Lijing, Ligong Duan, Xukun Li, and Guoping Li. "Acute-Exercise-Induced Alterations in Calpain and Calpastatin Expression in Rat Muscle." Journal of Sport Rehabilitation 18, no. 2 (May 2009): 213–28. http://dx.doi.org/10.1123/jsr.18.2.213.
Full textAbstract:
Context:Calpains and calpastatin can degrade muscle proteins, but no research has investigated the expression pattern of calpains and calpastatin after exhaustive exercise.Objective:To investigate the alterations in expression of μ-, m-, and n-calpain and calpastatin after exhaustive exercise and its association with muscle injury.Method:64 rats divided into 2 groups, a nonexercise control group and an acute-exhaustive-exercise (AEE) group. Biopsies in the AEE group were taken at different times after exercise.Results:Calpastatin protein expression and m-calpain activity increased early after exercise, but both n-calpain protein expression and μ-calpain activity generally decreased with time. n-Calpain mRNA expression was down- regulated from late after exercise.Conclusions:The increased m-calpain activity might promote muscle-protein degradation and muscle injury. On the contrary, calpastatin might execute a protective function against muscle injury. The change in p-calpain activity was found earlier than muscle injury and therefore might serve as a useful predictor of muscle injury.
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Lee, Wing-Kee, Blazej Torchalski, and Frank Thévenod. "Cadmium-induced ceramide formation triggers calpain-dependent apoptosis in cultured kidney proximal tubule cells." American Journal of Physiology-Cell Physiology 293, no. 3 (September 2007): C839—C847. http://dx.doi.org/10.1152/ajpcell.00197.2007.
Full textAbstract:
A major target of cadmium (Cd2+) toxicity is the kidney proximal tubule (PT) cell. Cd2+-induced apoptosis of PT cells is mediated by sequential activation of calpains at 3–6 h and caspases-9 and -3 after 24-h exposure. Calpains also partly contribute to caspase activation, which emphasizes the importance of calpains for PT apoptosis by Cd2+. Upstream processes underlying Cd2+-induced calpain activation remain unclear. We describe for the first time that 10–50 μM Cd2+ causes a significant increase in ceramide formation by ∼22% (3 h) and ∼72% (24 h), as measured by diacylglycerol kinase assay. Inhibition of ceramide synthase with fumonisin B1 (3 μM) prevents ceramide formation at 3 h and abolishes calpain activation at 6 h, which is associated with significant attenuation of apoptosis at 3–6 h with Hoechst 33342 nuclear staining and/or 3(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2 H-tetrazolium bromide (MTT) death assays. This indicates that Cd2+ enhances de novo ceramide synthesis and that calpains are a downstream target of ceramides in apoptosis execution. Moreover, addition of C6-ceramide to PT cells increases cytosolic Ca2+ and activates calpains. Apoptosis mediated by C6-ceramide at 24 h is significantly reduced by caspase-3 inhibition, which supports cross talk between calpain- and caspase-dependent apoptotic pathways. We conclude that Cd2+-induced apoptosis of PT cells entails endogenous ceramide elevation and subsequent Ca2+-dependent calpain activation, which propagates kidney damage by Cd2+.
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Macqueen, Daniel J., and Alexander H. Wilcox. "Characterization of the definitive classical calpain family of vertebrates using phylogenetic, evolutionary and expression analyses." Open Biology 4, no. 4 (April 2014): 130219. http://dx.doi.org/10.1098/rsob.130219.
Full textAbstract:
The calpains are a superfamily of proteases with extensive relevance to human health and welfare. Vast research attention is given to the vertebrate ‘classical’ subfamily, making it surprising that the evolutionary origins, distribution and relationships of these genes is poorly characterized. Consequently, there exists uncertainty about the conservation of gene family structure, function and expression that has been principally defined from work with mammals. Here, more than 200 vertebrate classical calpains were incorporated in phylogenetic analyses spanning an unprecedented range of taxa, including jawless and cartilaginous fish. We demonstrate that the common vertebrate ancestor had at least six classical calpains, including a single gene that gave rise to CAPN11 , 1 , 2 and 8 in the early jawed fish lineage, plus CAPN3 , 9 , 12 , 13 and a novel calpain gene, hereafter named CAPN17 . We reveal that while all vertebrate classical calpains have been subject to persistent purifying selection during evolution, the degree and nature of selective pressure has often been lineage-dependent. The tissue expression of the complete classic calpain family was assessed in representative teleost fish, amphibians, reptiles and mammals. This highlighted systematic divergence in expression across vertebrate taxa, with most classic calpain genes from fish and amphibians having more extensive tissue distribution than in amniotes. Our data suggest that classical calpain functions have frequently diverged during vertebrate evolution and challenge the ongoing value of the established system of classifying calpains by expression.
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Muniappan, Latha, Michihiro Okuyama, Aida Javidan, Devi Thiagarajan, Weihua Jiang, Jessica J. Moorleghen, Lihua Yang, et al. "Inducible Depletion of Calpain-2 Mitigates Abdominal Aortic Aneurysm in Mice." Arteriosclerosis, Thrombosis, and Vascular Biology 41, no. 5 (May 5, 2021): 1694–709. http://dx.doi.org/10.1161/atvbaha.120.315546.
Full textAbstract:
Objective: Cytoskeletal structural proteins maintain cell structural integrity by bridging extracellular matrix with contractile filaments. During abdominal aortic aneurysm (AAA) development, (1) aortic medial degeneration is associated with loss of smooth muscle cell integrity and (2) fibrogenic mesenchymal cells mediate extracellular matrix remodeling. Calpains cleave cytoskeletal proteins that maintain cell structural integrity. Pharmacological inhibition of calpains exert beneficial effects on Ang II (angiotensin II)–induced AAAs in LDLR −/− (low-density receptor deficient) mice. Here, we evaluated the functional contribution of fibrogenic mesenchymal cells-derived calpain-2 on (1) cytoskeletal structural protein and extracellular matrix alterations and (2) AAA progression. Approach and Results: Calpain-2 protein and cytoskeletal protein (filamin and talin) fragmentation are significantly elevated in human and Ang II–induced AAAs in mice. To examine the relative contribution of calpain-2 in AAA development, calpain-2 floxed mice in an LDLR −/− background were bred to mice with a tamoxifen-inducible form of Cre under control of either the ubiquitous promoter, chicken β-actin, or fibrogenic mesenchymal cell-specific promoter, Col1α2 (collagen type 1 alpha 2). Ubiquitous or fibrogenic mesenchymal cell-specific depletion of calpain-2 in mice suppressed Ang II–induced AAAs, filamin/talin fragmentation, while promoting extracellular matrix protein, collagen in the aortas. Calpain-2 silencing in aortic smooth muscle cells or fibroblasts reduced Ang II–induced filamin fragmentation. In addition, silencing of filamin in aortic SMCs significantly reduced collagen protein. Furthermore, calpain-2 deficiency suppressed rupture of established Ang II–induced AAAs in mice. Conclusions: Our studies implicate that calpain-2 deficiency prevents (1) Ang II–induced cytoskeletal structural protein fragmentation and AAA development and (2) stabilize and suppress rupture of established AAAs in mice.
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Kuchay, Shafi M., William P. Fay, and Athar H. Chishti. "Double Knockouts Reveal That Protein Tyrosine Phosphatase 1B Is a Physiological Substrate of Calpain-1 in Platelets." Blood 108, no. 11 (November 16, 2006): 396. http://dx.doi.org/10.1182/blood.v108.11.396.396.
Full textAbstract:
Abstract Calpains are ubiquitous calcium-regulated cysteine proteases that have been implicated in cytoskeletal organization, cell proliferation, apoptosis, cell motility, and hemostasis. Previously we used gene-targeting to evaluate the physiological function of mouse calpain-1, and established that its inactivation results in reduced platelet aggregation and clot retraction, potentially by causing dephosphorylation of platelet proteins. Here, we present data showing that calpain-1 null platelets accumulate protein tyrosine phosphatase 1B (PTP1B) that correlates with enhanced tyrosine phosphatase activity and dephosphorylation of multiple substrates in platelets. Using antibodies specific for phosphotyrosines 747 and 759 of the b3 subunit of αIibβ3 integrin, we show that the tyrosine phosphorylation of both tyrosine residues at positions 747 and 759 in the cytoplasmic domain of b3 subunit is reduced by approximately 60–70% in the calpain-1 null platelets. Treatment of calpain-1 null platelets with DMHV, an inhibitor of tyrosine phosphatases, corrected the aggregation defect and recovered impaired clot retraction. Importantly, platelet aggregation, clot retraction, and tyrosine dephosphorylation defects were rescued in the double knockout mice lacking both calpain-1 and PTP1B. Consistent with this paradigm, treatment of wild type mouse platelets as well as human platelets with the tyrosine phosphatase inhibitor DMHV enhanced their aggregation at low doses of thrombin. Conversely, MDL, a cell permeable inhibitor of calpains, potently inhibited aggregation of wild type mouse platelets in a dose-dependent manner upon thrombin activation. Further evaluation of mutant mice by ferric chloride induced arterial injury model suggests that the calpain-1 null mice are relatively resistant to thrombosis in vivo. Finally, the calpain-1 mediated regulation of PTP1B appears to be a systemic event as evident by the enhanced tyrosine dephosphorylation of B lymphocytes and their resistance to apoptosis in calpain-1 null mice. Together, our results demonstrate that PTP1B is a physiological substrate of calpain-1 and suggest that a similar mechanism may regulate calpain-1 mediated tyrosine dephosphorylation in other cells.