Journal articles: 'Biotin transporter'

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Finkenwirth, Friedrich, Olivia Neubauer, Julia Gunzenhäuser, Janna Schoknecht, Silvia Scolari, Martin Stöckl, Thomas Korte, Andreas Herrmann, and Thomas Eitinger. "Subunit composition of an energy-coupling-factor-type biotin transporter analysed in living bacteria." Biochemical Journal 431, no. 3 (October 11, 2010): 373–81. http://dx.doi.org/10.1042/bj20100813.

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BioMNY, a bacterial high-affinity biotin transporter, is a member of the recently defined class of ECF (energy-coupling factor) transporters. These systems are composed of ABC (ATP-binding-cassette) ATPases (represented by BioM in the case of the biotin transporter), a universally conserved transmembrane protein (BioN) and a core transporter component (BioY), in unknown stoichiometry. The quaternary structure of BioY, which functions as a low-affinity biotin transporter in the absence of BioMN, and of BioMNY was investigated by a FRET (Förster resonance energy transfer) approach using living recombinant Escherichia coli cells. To this end, the donor–acceptor pair, of Cerulean and yellow fluorescent protein respectively, were fused to BioM, BioN and BioY. The fusion proteins were stable and the protein tags did not interfere with transport and ATPase activities. Specific donor–acceptor interactions were characterized by lifetime-based FRET spectroscopy. The results suggest an oligomeric structure for the solitary BioY core transporter and oligomeric forms of BioM and BioY in BioMNY complexes. We surmise that oligomers of BioY are the functional units of the low- and high-affinity biotin transporter in the living cell. Beyond its relevance for clarifying the supramolecular organization of ECF transporters, the results demonstrate the general applicability of lifetime-based FRET studies in living bacteria.

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Zempleni, Janos, and Donald M. Mock. "Mitogen-induced proliferation increases biotin uptake into human peripheral blood mononuclear cells." American Journal of Physiology-Cell Physiology 276, no. 5 (May 1, 1999): C1079—C1084. http://dx.doi.org/10.1152/ajpcell.1999.276.5.c1079.

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We sought to determine whether the proliferation of immune cells affects the cellular uptake of the vitamin biotin. Peripheral blood mononuclear cells (PBMC) were isolated from healthy adults. The proliferation of PBMC was induced by either pokeweed lectin, concanavalin A, or phytohemagglutinin. When the medium contained a physiological concentration of [3H]biotin, nonproliferating PBMC accumulated 406 ± 201 amol [3H]biotin ⋅ 106cells−1 ⋅ 30 min−1. For proliferating PBMC, [3H]biotin uptake increased to between 330 and 722% of nonproliferating values. Maximal transport rates of [3H]biotin in proliferating PBMC were also about four times greater than those in nonproliferating PBMC, suggesting that proliferation was associated with an increase in the number of biotin transporters on the PBMC membrane. The biotin affinities and specificities of the transporter for proliferating and nonproliferating PBMC were similar, providing evidence that the same transporter mediates biotin uptake in both states. [14C]urea uptake values for proliferating and nonproliferating PBMC were similar, suggesting that the increased [3H]biotin uptake was not caused by a global upregulation of transporters during proliferation. We conclude that PBMC proliferation increases the cellular accumulation of biotin.

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Subramanian, Veedamali S., Jonathan S. Marchant, and Hamid M. Said. "Biotin-responsive basal ganglia disease-linked mutations inhibit thiamine transport via hTHTR2: biotin is not a substrate for hTHTR2." American Journal of Physiology-Cell Physiology 291, no. 5 (November 2006): C851—C859. http://dx.doi.org/10.1152/ajpcell.00105.2006.

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The water-soluble micronutrient thiamine is required for normal tissue growth and development in humans. Thiamine is accumulated into cells through the activity of two cell surface thiamine transporters (hTHTR1 and hTHTR2), which are differentially targeted in polarized tissues. Mutational dysfunction of hTHTR1 is associated with the clinical condition of thiamine-responsive megaloblastic anemia: the symptoms of which are alleviated by thiamine supplementation. Recently, two hTHTR2 mutants (G23V, T422A) have been discovered in clinical kindreds manifesting biotin-responsive basal ganglia disease (BBGD): the symptoms of which are alleviated by biotin administration. Why then does mutation of a specific thiamine transporter isoform precipitate a disorder correctable by exogenous biotin? To investigate the suggestion that hTHTR2 can physiologically function as a biotin transporter, we examined 1) the cell biological basis of hTHTR2 dysfunction associated with the G23V and T422A mutations and 2) the substrate specificity of hTHTR2 and these clinically relevant mutants. We show that the G23V and T422A mutants both abrogate thiamine transport activity rather than targeting of hTHTR2 to the cell surface. Furthermore, biotin accumulation was not detectable in cells overexpressing either the full length hTHTR2 or the clinically relevant hTHTR2 mutants, yet was demonstrable in the same assay using cells overexpressing the human sodium-dependent multivitamin transporter, a known biotin transporter. These results cast doubt on the most parsimonious explanation for the BBGD phenotype, namely that hTHTR2 is a physiological biotin transporter.

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Walker, Jennifer R., and Elliot Altman. "Biotinylation Facilitates the Uptake of Large Peptides by Escherichia coli and Other Gram-Negative Bacteria." Applied and Environmental Microbiology 71, no. 4 (April 2005): 1850–55. http://dx.doi.org/10.1128/aem.71.4.1850-1855.2005.

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ABSTRACT Gram-negative bacteria such as Escherichia coli can normally only take up small peptides less than 650 Da, or five to six amino acids, in size. We have found that biotinylated peptides up to 31 amino acids in length can be taken up by E. coli and that uptake is dependent on the biotin transporter. Uptake could be competitively inhibited by free biotin or avidin and blocked by the protonophore carbonyl m-chlorophenylhydrazone and was abolished in E. coli mutants that lacked the biotin transporter. Biotinylated peptides could be used to supplement the growth of a biotin auxotroph, and the transported peptides were shown to be localized to the cytoplasm in cell fractionation experiments. The uptake of biotinylated peptides was also demonstrated for two other gram-negative bacteria, Salmonella enterica serovar Typhimurium and Pseudomonas aeruginosa. This finding may make it possible to create new peptide antibiotics that can be used against gram-negative pathogens. Researchers have used various moieties to cause the illicit transport of compounds in bacteria, and this study demonstrates the illicit transport of the largest known compound to date.

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Kondo, Hiroki, Yasuaki Kazuta, and Tamami Goto. "Search for a microbial biotin transporter." BioFactors 11, no. 1-2 (2000): 101–2. http://dx.doi.org/10.1002/biof.5520110129.

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Zempleni, Janos, and Donald M. Mock. "Uptake and metabolism of biotin by human peripheral blood mononuclear cells." American Journal of Physiology-Cell Physiology 275, no. 2 (August 1, 1998): C382—C388. http://dx.doi.org/10.1152/ajpcell.1998.275.2.c382.

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We studied the uptake of biotin into human peripheral blood mononuclear cells (PBMC) using [3H]biotin and studied the catabolism of biotin in PBMC using [14C]biotin. Over 30 min, [3H]biotin uptake was greater at 37°C than at 25°C ( K T = 2.6 ± 0.4 nM, maximal velocity = 2.9 ± 0.2 fmol ⋅ 106cells−1 ⋅ 30 min−1). Ouabain reduced [3H]biotin uptake to 65% of control values, suggesting that biotin uptake is Na-K-ATPase dependent. Unlabeled biotin and biotin analogs reduced the uptake of [3H]biotin to 22–70% of control values, suggesting the presence of a competition for a structurally specific biotin transporter. When endocytosis by PBMC was stimulated by various acyl glycerols, [3H]biotin uptake was 40–73% of control values; these data are consistent with the hypothesis that stimulated endocytosis reduces biotin transporter density on the cell surface. During a 168-h incubation, PBMC did not catabolize [14C]biotin.

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Ghosal, Abhisek, Stefan Jellbauer, Rubina Kapadia, Manuela Raffatellu, and Hamid M. Said. "Salmonellainfection inhibits intestinal biotin transport: cellular and molecular mechanisms." American Journal of Physiology-Gastrointestinal and Liver Physiology 309, no. 2 (July 15, 2015): G123—G131. http://dx.doi.org/10.1152/ajpgi.00112.2015.

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Infection with the nontyphoidal Salmonella is a common cause of food-borne disease that leads to acute gastroenteritis/diarrhea. Severe/prolonged cases of Salmonella infection could also impact host nutritional status, but little is known about its effect on intestinal absorption of vitamins, including biotin. We examined the effect of Salmonella enterica serovar Typhimurium ( S. typhimurium) infection on intestinal biotin uptake using in vivo (streptomycin-pretreated mice) and in vitro [mouse (YAMC) and human (NCM460) colonic epithelial cells, and human intestinal epithelial Caco-2 cells] models. The results showed that infecting mice with wild-type S. typhimurium, but not with its nonpathogenic isogenic invA spiB mutant, leads to a significant inhibition in jejunal/colonic biotin uptake and in level of expression of the biotin transporter, sodium-dependent multivitamin transporter. In contrast, infecting YAMC, NCM460, and Caco-2 cells with S. typhimurium did not affect biotin uptake. These findings suggest that the effect of S. typhimurium infection is indirect and is likely mediated by proinflammatory cytokines, the levels of which were markedly induced in the intestine of S. typhimurium-infected mice. Consistent with this hypothesis, exposure of NCM460 cells to the proinflammatory cytokines TNF-α and IFN-γ led to a significant inhibition of biotin uptake, sodium-dependent multivitamin transporter expression, and activity of the SLC5A6 promoter. The latter effects appear to be mediated, at least in part, via the NF-κB signaling pathway. These results demonstrate that S. typhimurium infection inhibits intestinal biotin uptake, and that the inhibition is mediated via the action of proinflammatory cytokines.

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Lakhan, Ram, and Hamid M. Said. "Lipopolysaccharide inhibits colonic biotin uptake via interference with membrane expression of its transporter: a role for a casein kinase 2-mediated pathway." American Journal of Physiology-Cell Physiology 312, no. 4 (April 1, 2017): C376—C384. http://dx.doi.org/10.1152/ajpcell.00300.2016.

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Biotin (vitamin B7), an essential micronutrient for normal cellular functions, is obtained from both dietary sources as well as gut microbiota. Absorption of biotin in both the small and large intestine is via a carrier-mediated process that involves the sodium-dependent multivitamin transporter (SMVT). Although different physiological and molecular aspects of intestinal biotin uptake have been delineated, nothing is known about the effect of LPS on the process. We addressed this issue using in vitro (human colonic epithelial NCM460 cells) and in vivo (mice) models of LPS exposure. Treating NCM460 cells with LPS was found to lead to a significant inhibition in carrier-mediated biotin uptake. Similarly, administration of LPS to mice led to a significant inhibition in biotin uptake by native colonic tissue. Although no changes in total cellular SMVT protein and mRNA levels were observed, LPS caused a decrease in the fraction of SMVT expressed at the cell surface. A role for casein kinase 2 (CK2) (whose activity was also inhibited by LPS) in mediating the endotoxin effects on biotin uptake and on membrane expression of SMVT was suggested by findings that specific inhibitors of CK2, as well as mutating the putative CK2 phosphorylation site (Thr78Ala) in the SMVT protein, led to inhibition in biotin uptake and membrane expression of SMVT. This study shows for the first time that LPS inhibits colonic biotin uptake via decreasing membrane expression of its transporter and that these effects likely involve a CK2-mediated pathway.

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Vlasova, Tatyana I., Shawna L. Stratton, Amanda M. Wells, Nell I. Mock, and Donald M. Mock. "Biotin Deficiency Reduces Expression of SLC19A3, a Potential Biotin Transporter, in Leukocytes from Human Blood." Journal of Nutrition 135, no. 1 (January 1, 2005): 42–47. http://dx.doi.org/10.1093/jn/135.1.42.

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10

Griffin, Jacob, Steven Stanley, and Janos Zempleni. "Synthesis of a Rabbit Polyclonal Antibody to the Human Sodium-Dependent Multivitamin Transporter." International Journal for Vitamin and Nutrition Research 72, no. 4 (July 1, 2002): 195–98. http://dx.doi.org/10.1024/0300-9831.72.4.195.

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In mammalian cells, biotin is covalently attached to carboxylases and histones and is required for cell proliferation and function. Cellular uptake of biotin (as well as pantothenic acid and lipoic acid) is mediated by the sodium-dependent multivitamin transporter, SMVT. Studies of cellular biotin homeostasis have been hampered by the lack of an antibody to SMVT. Here, we describe the synthesis of a rabbit polyclonal antibody to human SMVT. Using this antibody, SMVT has been identified in human peripheral blood mononuclear cells, Caco-2 cells, and HepG2 cells. Moreover, we observed that cells respond to proliferation with increased synthesis of SMVT.

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Sabui, Subrata, Jennifer Ann Bohl, Rubina Kapadia, Kyle Cogburn, Abhisek Ghosal, Nils W. Lambrecht, and Hamid M. Said. "Role of the sodium-dependent multivitamin transporter (SMVT) in the maintenance of intestinal mucosal integrity." American Journal of Physiology-Gastrointestinal and Liver Physiology 311, no. 3 (September 1, 2016): G561—G570. http://dx.doi.org/10.1152/ajpgi.00240.2016.

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Utilizing a conditional (intestinal-specific) knockout (cKO) mouse model, we have recently shown that the sodium-dependent multivitamin transporter (SMVT) ( SLC5A6) is the only biotin uptake system that operates in the gut and that its deletion leads to biotin deficiency. Unexpectedly, we also observed that all SMVT-cKO mice develop chronic active inflammation, especially in the cecum. Our aim here was to examine the role of SMVT in the maintenance of intestinal mucosal integrity [permeability and expression of tight junction (TJ) proteins]. Our results showed that knocking out the mouse intestinal SMVT is associated with a significant increase in gut permeability and with changes in the level of expression of TJ proteins. To determine whether these changes are related to the state of biotin deficiency that develops in SMVT-cKO mice, we induced (by dietary means) biotin deficiency in wild-type mice and examined its effect on the above-mentioned parameters. The results showed that dietary-induced biotin deficiency leads to a similar development of chronic active inflammation in the cecum with an increase in the level of expression of proinflammatory cytokines, as well as an increase in intestinal permeability and changes in the level of expression of TJ proteins. We also examined the effect of chronic biotin deficiency on permeability and expression of TJ proteins in confluent intestinal epithelial Caco-2 monolayers but observed no changes in these parameters. These results show that the intestinal SMVT plays an important role in the maintenance of normal mucosal integrity, most likely via its role in providing biotin to different cells of the gut mucosa.

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Ghosal, Abhisek, Nils Lambrecht, Sandeep B. Subramanya, Rubina Kapadia, and Hamid M. Said. "Conditional knockout of the Slc5a6 gene in mouse intestine impairs biotin absorption." American Journal of Physiology-Gastrointestinal and Liver Physiology 304, no. 1 (January 1, 2013): G64—G71. http://dx.doi.org/10.1152/ajpgi.00379.2012.

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The Slc5a6 gene expresses a plasma membrane protein involved in the transport of the water-soluble vitamin biotin; the transporter is commonly referred to as the sodium-dependent multivitamin transporter (SMVT) because it also transports pantothenic acid and lipoic acid. The relative contribution of the SMVT system toward carrier-mediated biotin uptake in the native intestine in vivo has not been established. We used a Cre/lox technology to generate an intestine-specific (conditional) SMVT knockout (KO) mouse model to address this issue. The KO mice exhibited absence of expression of SMVT in the intestine compared with sex-matched littermates as well as the expected normal SMVT expression in other tissues. About two-thirds of the KO mice died prematurely between the age of 6 and 10 wk. Growth retardation, decreased bone density, decreased bone length, and decreased biotin status were observed in the KO mice. Microscopic analysis showed histological abnormalities in the small bowel (shortened villi, dysplasia) and cecum (chronic active inflammation, dysplasia) of the KO mice. In vivo (and in vitro) transport studies showed complete inhibition in carrier-mediated biotin uptake in the intestine of the KO mice compared with their control littermates. These studies provide the first in vivo confirmation in native intestine that SMVT is solely responsible for intestinal biotin uptake. These studies also provide evidence for a casual association between SMVT function and normal intestinal health.

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Subramanian, Veedamali S., Jonathan S. Marchant, Michael J. Boulware, Thomas Y. Ma, and Hamid M. Said. "Membrane targeting and intracellular trafficking of the human sodium-dependent multivitamin transporter in polarized epithelial cells." American Journal of Physiology-Cell Physiology 296, no. 4 (April 2009): C663—C671. http://dx.doi.org/10.1152/ajpcell.00396.2008.

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The human sodium-dependent multivitamin transporter (hSMVT) mediates sodium-dependent uptake of biotin in renal and intestinal epithelia. To date, however, there is nothing known about the structure-function relationship or targeting sequences in the hSMVT polypeptide that control its polarized expression within epithelia. Here, we focused on the role of the COOH-terminal tail of hSMVT in the targeting and functionality of this transporter. A full-length hSMVT-green fluorescent protein (GFP) fusion protein was functional and expressed at the apical membrane in renal and intestinal cell lines. Microtubule disrupting agents disrupted the mobility of trafficking vesicles and impaired cell surface delivery of hSMVT, which was also prevented in cells treated with dynamitin (p50), brefeldin, or monensin. Progressive truncation of the COOH-terminal tail impaired the functionality and targeting of the transporter. First, biotin transport decreased by approximately 20–30% on deletion of up to 15 COOH-terminal amino acids of hSMVT, a decrease mimicked solely by deletion of the terminal PDZ motif (TSL). Second, deletions into the COOH-terminal tail (between residues 584-612, containing a region of predicted high surface accessibility) resulted in a further drop in hSMVT transport (to ∼40% of wild-type). Third, apical targeting was lost on deletion of a helical-prone region between amino acids 570-584. We conclude that the COOH tail of hSMVT contains several determinants important for polarized targeting and biotin transport.

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Entcheva, Plamena, Donald A. Phillips, and Wolfgang R. Streit. "Functional Analysis of Sinorhizobium meliloti Genes Involved in Biotin Synthesis and Transport." Applied and Environmental Microbiology 68, no. 6 (June 2002): 2843–48. http://dx.doi.org/10.1128/aem.68.6.2843-2848.2002.

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ABSTRACT External biotin greatly stimulates bacterial growth and alfalfa root colonization by Sinorhizobium meliloti strain 1021. Several genes involved in responses to plant-derived biotin have been identified in this bacterium, but no genes required for biotin transport are known, and not all loci required for biotin synthesis have been assigned. Searches of the S. meliloti genome database in combination with complementation tests of Escherichia coli biotin auxotrophs indicate that biotin synthesis probably is limited in S. meliloti 1021 by the poor functioning or complete absence of several key genes. Although several open reading frames with significant similarities to genes required for synthesis of biotin in gram-positive and gram-negative bacteria were found, only bioB, bioF, and bioH were demonstrably functional in complementation tests with known E. coli mutants. No sequence or complementation evidence was found for bioA, bioC, bioD, or bioZ. In contrast to other microorganisms, the S. meliloti bioB and bioF genes are not localized in a biotin synthesis operon, but bioB is cotranscribed with two genes coding for ABC transporter-like proteins, designated here bioM and bioN. Mutations in bioM and bioN eliminated growth on alfalfa roots and reduced bacterial capacity to maintain normal intracellular levels of biotin. Taken together, these data suggest that S. meliloti normally grows on exogenous biotin using bioM and bioN to conserve biotin assimilated from external sources.

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Yang, Julianne C., Jonathan P. Jacobs, Michael Hwang, Subrata Sabui, Fengting Liang, Hamid M. Said, and Jonathan Skupsky. "Biotin Deficiency Induces Intestinal Dysbiosis Associated with an Inflammatory Bowel Disease-like Phenotype." Nutrients 15, no. 2 (January 4, 2023): 264. http://dx.doi.org/10.3390/nu15020264.

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Biotin is an essential vitamin and critical cofactor in several metabolic pathways, and its deficiency has been linked to several disorders including inflammatory bowel disease (IBD). We previously reported that biotin deficiency (BD) in mice, whether modeled through intestine-specific deletion of biotin transporter (SMVT-icKO) or through a biotin-deficient diet, resulted in intestinal inflammation consistent with an IBD-like phenotype. To assess whether the gut microbiome is associated with these BD-induced changes, we collected stool and intestinal samples from both of these mouse models and utilized them for 16S rRNA gene sequencing. We find that both diet-mediated and deletion-mediated BD result in the expansion of opportunistic microbes including Klebsiella, Enterobacter, and Helicobacter, at the expense of mucus-resident microbes including Akkermansia. Additionally, microbiome dysbiosis resulting from diet-mediated BD precedes the onset of the IBD-like phenotypic changes. Lastly, through the use of predictive metagenomics, we report that the resulting BD-linked microbiome perturbations exhibit increased biotin biosynthesis in addition to several other perturbed metabolic pathways. Altogether, these results demonstrate that biotin deficiency results in a specific microbiome composition, which may favor microbes capable of biotin synthesis and which may contribute to intestinal inflammation.

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Stolz, J�rgen. "Isolation and characterization of the plasma membrane biotin transporter fromSchizosaccharomyces pombe." Yeast 20, no. 3 (2003): 221–31. http://dx.doi.org/10.1002/yea.959.

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Ghosal, Abhisek, and Hamid M. Said. "Structure-function activity of the human sodium-dependent multivitamin transporter: role of His115 and His254." American Journal of Physiology-Cell Physiology 300, no. 1 (January 2011): C97—C104. http://dx.doi.org/10.1152/ajpcell.00398.2010.

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Intestinal absorption of biotin occurs via a Na+-dependent carrier-mediated process that involves the sodium-dependent multivitamin transporter (SMVT; product of the Slc5a6 gene). The SMVT system is exclusively expressed at the apical membrane domain of the polarized intestinal epithelial cells. Whereas previous studies from our laboratory and others have characterized different physiological and biological aspects of SMVT, little is currently known about its structure-function activity relationship. Using site-directed mutagenesis approach, we examined the role of the positively charged histidine (His) residues of the human SMVT (hSMVT) in transporting the negatively charged biotin. Of the seven conserved (across species) His residues in the hSMVT polypeptide, only His115 and His254 were found to be important for the function of hSMVT as their mutation led to a significant reduction in carrier-mediated biotin uptake. This inhibition was mediated via a significant reduction in the maximal velocity ( Vmax), but not the apparent Michaelis constant ( Km), of the biotin uptake process and was not related to the charge of the His residue. The inhibition was also not due to changes in transcriptional or translational efficiency of the mutated hSMVT compared with wild-type carrier. However, surface biotinylation assay showed a significant reduction in the level of expression of the mutated hSMVT at the cell surface, a finding that was further confirmed by confocal imaging. Our results show important role for His115 and His254 residues in hSMVT function, which is most probably mediated via an effect on level of hSMVT expression at the cell membrane.

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Erbach, Johanna, Florian Bonn, Max Diesner, Anne Arnold, Jürgen Stein, Oliver Schröder, and Ayşegül Aksan. "Relevance of Biotin Deficiency in Patients with Inflammatory Bowel Disease and Utility of Serum 3 Hydroxyisovaleryl Carnitine as a Practical Everyday Marker." Journal of Clinical Medicine 11, no. 4 (February 20, 2022): 1118. http://dx.doi.org/10.3390/jcm11041118.

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Background: Biotin, a water-soluble B vitamin, has demonstrable anti-inflammatory properties. A biotin-deficient diet induced a colitis-like phenotype in mice, alleviable by biotin substitution. Mice with dextran sulfate sodium (DSS)-induced colitis showed biotin deficiency and diminished levels of sodium-dependent multivitamin transporter, a protein involved in biotin absorption. Biotin substitution induced remission by reducing activation of NF-κB, a transcription factor involved in intestinal permeability and inflammatory bowel disease (IBD). We investigated for the first time a possible clinical role of biotin status in IBD. Methods: In a comparative, retrospective, cross-sectional study, serum samples of 138 patients with IBD (67 female; 72 Crohn’s disease (CD), 66 ulcerative colitis (UC)) aged 18–65 years and with a mean age (±SD) of 42.5 ± 14.3 years as well as 80 healthy blood donors (40 female; 40.0 ± 10.0 years; range 20–60 years) were analyzed. Inflammation was defined as hsCRP ≥5 mg/L, and to determine biotin status, serum 3-hydroxyisovaleryl carnitine (3HIVc) levels were measured by LC-MS/MS. Results: A total of 138 patients with IBD (67f; 72CD/66 UC; 42.5 ± 14.3 years) were enrolled: 83/138 had inflammation. Mean serum 3HIVc levels were significantly higher in IBD patients but unaffected by inflammation. Biotin deficiency (95th percentile of controls: >30 nmol/L 3HIVc) was significantly more common in IBD patients versus controls. Conclusion: High serum 3HIVc levels and biotin deficiency were associated with IBD but not inflammatory activity or disease type. Our findings suggest biotin may play a role as cause or effect in IBD pathogenesis. Routine assessment and supplementation of biotin may ameliorate IBD and support intestinal integrity.

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Reidling, Jack C., Svetlana M. Nabokina, and Hamid M. Said. "Molecular mechanisms involved in the adaptive regulation of human intestinal biotin uptake: a study of the hSMVT system." American Journal of Physiology-Gastrointestinal and Liver Physiology 292, no. 1 (January 2007): G275—G281. http://dx.doi.org/10.1152/ajpgi.00327.2006.

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Biotin, a water-soluble micronutrient, is vital for cellular functions, including growth and development. The human intestine utilizes the human sodium-dependent multivitamin transporter (hSMVT) for biotin uptake. Evidence exists showing that the intestinal biotin uptake process is adaptively regulated during biotin deficiency. Nothing, however, is known about molecular mechanism(s) involved during this adaptive regulation. This study compared two human-derived intestinal epithelial cell lines (HuTu-80 and Caco-2) during biotin-deficient or biotin-sufficient states and with an approach that assessed carrier-mediated biotin uptake, hSMVT protein and RNA levels, RNA stability, and hSMVT promoter activity. The results showed that during biotin deficiency, a significant and specific upregulation in carrier-mediated biotin uptake occurred in both human intestinal epithelial cell lines and that this increase was associated with an induction in protein and mRNA levels of hSMVT. The increase in mRNA levels was not due to an increase in RNA stability but was associated with an increase in activity of the hSMVT promoter in transfected human intestinal cells. Using promoter deletion constructs and mutational analysis in transiently transfected HuTu-80 and Caco-2 cells, a biotin deficiency-responsive region was mapped to a 103-bp area within the hSMVT promoter that contains gut-enriched Kruppel-like factor (GKLF) sites that confer the response to biotin deficiency. These results confirm that human intestinal biotin uptake is adaptively regulated and provide novel evidence demonstrating that the upregulation is not mediated via changes in hSMVT RNA stability but rather is due to transcriptional regulatory mechanism(s) that likely involve GKLF sites in the hSMVT promoter.

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Pacheco-Alvarez, Diana, R. Sergio Solórzano-Vargas, Alfonso González-Noriega, Colette Michalak, Janos Zempleni, and Alfonso León-Del-Río. "Biotin availability regulates expression of the sodium-dependent multivitamin transporter and the rate of biotin uptake in HepG2 cells." Molecular Genetics and Metabolism 85, no. 4 (August 2005): 301–7. http://dx.doi.org/10.1016/j.ymgme.2005.04.001.

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Nabokina, Svetlana M., Veedamali S. Subramanian, and Hamid M. Said. "Comparative analysis of ontogenic changes in renal and intestinal biotin transport in the rat." American Journal of Physiology-Renal Physiology 284, no. 4 (April 1, 2003): F737—F742. http://dx.doi.org/10.1152/ajprenal.00364.2002.

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Biotin, an essential water-soluble micronutrient, cannot be synthesized by mammals; rather, it is obtained from exogenous sources via uptake by intestinal epithelia. Renal epithelia reclaim the vitamin that is filtered in the glomeruli. Both epithelia take up biotin via the sodium-dependent multivitamin transporter (SMVT). Little is known about ontogenic regulation of the renal and intestinal biotin transport processes and about the mechanism(s) involved in any such regulation. In this study, we sought to examine and compare ontogenic aspects of the renal and intestinal biotin uptake processes using purified brush-border membrane vesicles (BBMV) isolated from the kidney cortex and jejunum of suckling and adult rats. Clear ontogenic changes were observed in the intestinal biotin uptake process, which were mediated via changes in V max and apparent K m. Parallel changes were also seen in protein, mRNA, and transcription rate of SMVT as indicated by results of Western blotting, RT-PCR, and nuclear run-on assays, respectively. In contrast, biotin uptake by renal BBMV did not show ontogenic changes; i.e., it was similar in suckling and adult rats. Also, the levels of SMVT protein and mRNA were similar in the kidneys of both age groups. These data show that biotin uptake by renal and intestinal epithelial cells responds differently to ontogenic regulation. In addition, the ontogenic changes observed in the intestinal biotin uptake process involve the entry step of the vitamin at the BBM and appear to be mediated via a transcriptional mechanism(s).

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Balamurugan, Krishnaswamy, Alvaro Ortiz, and Hamid M. Said. "Biotin uptake by human intestinal and liver epithelial cells: role of the SMVT system." American Journal of Physiology-Gastrointestinal and Liver Physiology 285, no. 1 (July 2003): G73—G77. http://dx.doi.org/10.1152/ajpgi.00059.2003.

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It has been well established that human intestinal and liver epithelial cells transport biotin via an Na+-dependent carrier-mediated mechanism. The sodium-dependent multivitamin transport (SMVT), a biotin transporter, is expressed in both cell types. However, the relative contribution of SMVT toward total carrier-mediated uptake of physiological (nanomolar) concentrations of biotin by these cells is not clear. Addressing this issue is important, especially in light of the recent identification of a second human high-affinity biotin uptake mechanism that operates at the nanomolar range. Hence, we employed a physiological approach of characterizing biotin uptake by human-derived intestinal Caco-2 and HepG2 cells at the nanomolar concentration range. We also employed a molecular biology approach of selectively silencing the endogenous SMVT of these cells with specific small interfering RNAs (siRNAs), then examining carrier-mediated biotin uptake. The results showed that in both Caco-2 and HepG2 cells, the initial rate of biotin uptake as a function of concentration over the range of 0.1 to 50 nM to be linear. Furthermore, we found that the addition of 100 nM unlabeled biotin, desthiobiotin, or pantothenic acid to the incubation medium had no effect on the uptake of 2.6 nM [3H]biotin. Pretreatment of Caco-2 and HepG2 cells with SMVT specific siRNAs substantially reduced SMVT mRNA and protein levels. In addition, carrier-mediated [3H]biotin (2.6 nM) uptake by Caco-2 and HepG2 cells was severely ( P 0.01) inhibited by the siRNAs pretreatment. These results demonstrate that the recently described human high-affinity biotin uptake system is not functional in intestinal and liver epithelial cells. In addition, the results provide strong evidence that SMVT is the major (if not the only) biotin uptake system that operates in these cells.

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Pommerrenig, Benjamin, Jennifer Popko, Mareike Heilmann, Sylwia Schulmeister, Katharina Dietel, Bianca Schmitt, Ruth Stadler, Ivo Feussner, and Norbert Sauer. "SUCROSE TRANSPORTER 5 supplies Arabidopsis embryos with biotin and affects triacylglycerol accumulation." Plant Journal 73, no. 3 (December 31, 2012): 392–404. http://dx.doi.org/10.1111/tpj.12037.

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Daberkow, Rachel L., Brett R. White, Rebecca A. Cederberg, Jacob B. Griffin, and Janos Zempleni. "Monocarboxylate Transporter 1 Mediates Biotin Uptake in Human Peripheral Blood Mononuclear Cells." Journal of Nutrition 133, no. 9 (September 1, 2003): 2703–6. http://dx.doi.org/10.1093/jn/133.9.2703.

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Nabokina, Svetlana M., Veedamali S. Subramanian, and Hamid M. Said. "Association of PDZ-containing protein PDZD11 with the human sodium-dependent multivitamin transporter." American Journal of Physiology-Gastrointestinal and Liver Physiology 300, no. 4 (April 2011): G561—G567. http://dx.doi.org/10.1152/ajpgi.00530.2010.

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Intestinal absorption of biotin is mediated via the sodium-dependent multivitamin transporter (SMVT). Studies from our laboratory and others have characterized different aspects of the human SMVT (hSMVT), but nothing is currently known about protein(s) that may interact with hSMVT and affect its physiology/biology. In this study, a PDZ-containing protein PDZD11 was identified as an interacting partner with hSMVT using yeast two-hybrid screen of a human intestinal cDNA library. The interaction between hSMVT and PDZD11 was confirmed by in vitro GST-pull-down assay and in vivo in a mammalian cell environment by a two-hybrid luciferase and coimmunoprecipitation assays. Furthermore, confocal imaging of live human intestinal epithelial HuTu-80 cells expressing hSMVT-GFP and DsRed-PDZD11 demonstrated colocalization of these two proteins. We also examined the functional consequence of the interaction between hSMVT and PDZD11 in HuTu-80 cells and observed significant induction in [3H]biotin uptake upon coexpression of hSMVT and PDZD11. In contrast, knocking down of PDZD11 with gene-specific small interfering RNA led to a significant decrease in biotin uptake; biotinylation assay showed this to be associated with a marked decrease in level of expression of hSMVT at the cell membrane. By truncation approach, we also demonstrated that the PDZ binding domain that is located in the COOH-terminal tail of hSMVT polypeptide is involved in the interaction with PDZD11. These results demonstrate for the first time that PDZD11 is an interacting partner with hSMVT in intestinal epithelial cells and that this interaction affects hSMVT function and cell biology.

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26

Hong, Mei, Kunihiko Tanaka, Zui Pan, Jianjie Ma, and Guofeng You. "Determination of the external loops and the cellular orientation of the N- and the C-termini of the human organic anion transporter hOAT1." Biochemical Journal 401, no. 2 (December 21, 2006): 515–20. http://dx.doi.org/10.1042/bj20061171.

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The OAT (organic anion transporter) family mediates the absorption, distribution and excretion of a diverse array of environmental toxins and clinically important drugs. OAT dysfunction significantly contributes to renal, hepatic, neurological and fetal toxicity and disease. As a first step to establish the topological model of hOAT1 (human OAT1), we investigated the external loops and the cellular orientation of the N- and the C-termini of this transporter. Combined approaches of immunofluorescence studies and site-directed chemical labelling were used for such purpose. Immunofluorescence microscopy of Myc-tagged hOAT1 expressed in cultured cells identified that both the N- and the C-termini of the transporter were located in the cytoplasm. Replacement of Lys59 in the predicted extracellular loop I with arginine resulted in a mutant (K59R), which was largely inaccessible for labelling by membrane-impermeable NHS (N-hydroxysuccinimido)-SS (dithio)-biotin present in the extracellular medium. This result suggests that loop I faces outside of the cell membrane. A single lysine residue introduced into putative extracellular loops III, V and VI of mutant K59R, which is devoid of extracellular lysine, reacted readily with membrane-impermeable NHS-SS-biotin, suggesting that these putative extracellular loops are in the extracellular domains of the protein. These studies provided the first experimental evidence on the extracellular loops and the cellular orientation of the N- and the C-termini of hOAT1.

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Ghosal, Abhisek, Thillai V. Sekar, and Hamid M. Said. "Biotin uptake by mouse and human pancreatic beta cells/islets: a regulated, lipopolysaccharide-sensitive carrier-mediated process." American Journal of Physiology-Gastrointestinal and Liver Physiology 307, no. 3 (August 1, 2014): G365—G373. http://dx.doi.org/10.1152/ajpgi.00157.2014.

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Biotin is essential for the normal function of pancreatic beta cells. These cells obtain biotin from their surroundings via transport across their cell membrane. Little is known about the uptake mechanism involved, how it is regulated, and how it is affected by internal and external factors. We addressed these issues using the mouse-derived pancreatic beta-TC-6 cells and freshly isolated mouse and human primary pancreatic beta cells as models. The results showed biotin uptake by pancreatic beta-TC-6 cells occurs via a Na+-dependent, carrier-mediated process, that is sensitive to desthiobiotin, as well as to pantothenic acid and lipoate; the process is also saturable as a function of concentration (apparent Km = 22.24 ± 5.5 μM). These cells express the sodium-dependent multivitamin transporter (SMVT), whose knockdown (with doxycycline-inducible shRNA) led to a sever inhibition in biotin uptake. Similarly, uptake of biotin by mouse and human primary pancreatic islets is Na+-dependent and carrier-mediated, and both cell types express SMVT. Biotin uptake by pancreatic beta-TC-6 cells is also adaptively regulated (via transcriptional mechanism) by extracellular substrate level. Chronic treatment of pancreatic beta-TC-6 cells with bacterial lipopolysaccharides (LPS) leads to inhibition in biotin uptake. This inhibition is mediated via a Toll-Like receptor 4-mediated process and involves a decrease in membrane expression of SMVT. These findings show, for the first time, that pancreatic beta cells/islets take up biotin via a specific and regulated carrier-mediated process, and that the process is sensitive to the effect of LPS.

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Neubauer, Olivia, Anja Alfandega, Janna Schoknecht, Ulrich Sternberg, Anne Pohlmann, and Thomas Eitinger. "Two Essential Arginine Residues in the T Components of Energy-Coupling Factor Transporters." Journal of Bacteriology 191, no. 21 (August 28, 2009): 6482–88. http://dx.doi.org/10.1128/jb.00965-09.

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ABSTRACT Energy-coupling factor (ECF) transporters, a recently discovered class of importers of micronutrients, are composed of a substrate-specific transmembrane component (S component) and a conserved energy-coupling module consisting of a transmembrane protein (T component) and pairs of ABC ATPases (A proteins). Based on utilization of a dedicated (subclass I) or shared (subclass II) energy-coupling module, ECF systems fall into two subclasses. The T components are the least-characterized proteins of ECF importers, and their function is essentially unknown. Using RcBioN and LmEcfT, the T units of the subclass I biotin transporter (RcBioMNY) of a gram-negative bacterium and of the subclass II folate, pantothenate, and riboflavin transporters of a lactic acid bacterium, respectively, we analyzed the role of two strongly conserved short motifs, each containing an arginine residue. Individual replacement of the two Arg residues in RcBioN reduced ATPase activity, an indicator of the transporter function, by two-thirds without affecting the modular assembly of the RcBioMNY complex. A double Arg-to-Glu replacement destroyed the complex and abolished ATPase activity. The corresponding single mutation in motif II of LmEcfT, as well as a double mutation, led to loss of the T unit from the subclass II ECF transporters and inactivated these systems. A single Arg-to-Glu replacement in motif I, however, abolished vitamin uptake activity without affecting assembly of the modules. Our results indicate that the conserved motif I in T components is essential for intramolecular signaling and, in cooperation with motif II, for subunit assembly of modular ECF transporters.

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Balamurugan, Krishnaswamy, Nosratola D. Vaziri, and Hamid M. Said. "Biotin uptake by human proximal tubular epithelial cells: cellular and molecular aspects." American Journal of Physiology-Renal Physiology 288, no. 4 (April 2005): F823—F831. http://dx.doi.org/10.1152/ajprenal.00375.2004.

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Cellular and molecular regulation of renal biotin uptake in humans is not well defined. The contribution of the human Na+-dependent multivitamin transporter (hSMVT) to carrier-mediated biotin uptake by human proximal tubular epithelial cells is not clear. The aim of this study was to address these issues, with the human-derived proximal tubular epithelial HK-2 cells used as a model. First, we characterized the mechanism of biotin uptake by these cells and obtained evidence for involvement of an Na+-, temperature-, and energy-dependent carrier-mediated uptake system. This system was inhibited by the biotin structural analog desthiobiotin, pantothenic acid, and lipoate. These findings suggest involvement of the hSMVT system in the uptake process. This was confirmed by demonstrating that the hSMVT system is expressed in HK-2 cells at the protein and mRNA levels and by selective silencing of the hSMVT gene with the use of gene-specific small interfering RNAs, which led to specific and significant inhibition of carrier-mediated biotin uptake. Of the two recently cloned promoters of the hSMVT gene, promoter 1 was more active than promoter 2 in these cells. Pretreatment of HK-2 cells with modulators of PKC- and Ca2+/calmodulin-mediated pathways (but not those that modulate PKA-, protein tyrosine kinase-, or nitric oxide-mediated pathways) led to significant alterations in biotin uptake. Maintaining the HK-2 cells in a biotin-deficient growth medium led to a marked upregulation in biotin transport, which was associated with an increase in hSMVT protein and RNA levels and an increase in activity of the hSMVT promoters. These results demonstrate that biotin uptake by human renal epithelial cells occurs via the hSMVT system and that the process is regulated by intracellular PKC- and Ca2+/calmodulin-mediated pathways. The uptake process appears to be adaptively regulated by extracellular biotin level, which involves transcriptional regulatory mechanism(s).

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Subramanya, Sandeep B., Veedamali S. Subramanian, Jeyan S. Kumar, Robert Hoiness, and Hamid M. Said. "Inhibition of intestinal biotin absorption by chronic alcohol feeding: cellular and molecular mechanisms." American Journal of Physiology-Gastrointestinal and Liver Physiology 300, no. 3 (March 2011): G494—G501. http://dx.doi.org/10.1152/ajpgi.00465.2010.

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The water-soluble vitamin biotin is essential for normal cellular functions and its deficiency leads to a variety of clinical abnormalities. Mammals obtain biotin from exogenous sources via intestinal absorption, a process mediated by the sodium-dependent multivitamin transporter (SMVT). Chronic alcohol use in humans is associated with a significant reduction in plasma biotin levels, and animal studies have shown inhibition in intestinal biotin absorption by chronic alcohol feeding. Little, however, is known about the cellular and molecular mechanisms involved in the inhibition in intestinal biotin transport by chronic alcohol use. These mechanisms were investigated in this study by using rats and transgenic mice carrying the human full-length SLC5A6 5′-regulatory region chronically fed alcohol liquid diets; human intestinal epithelial Caco-2 cells chronically exposed to alcohol were also used as models. The results showed chronic alcohol feeding of rats to lead to a significant inhibition in carrier-mediated biotin transport events across jejunal brush border and basolateral membrane domains. This inhibition was associated with a significant reduction in level of expression of the SMVT protein, mRNA, and heterogenous nuclear RNA. Chronic alcohol feeding also inhibited carrier-mediated biotin uptake in rat colon. Studies with transgenic mice confirmed the above findings and further showed chronic alcohol feeding significantly inhibited the activity of SLC5A6 5′-regulatory region. Finally, chronic exposure of Caco-2 cells to alcohol led to a significant decrease in the activity of both promoters P1 and P2 of the human SLC5A6 gene. These studies identify for the first time the cellular and molecular parameters of the intestinal biotin absorptive processes that are affected by chronic alcohol feeding.

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Tomlinson, Ian D., Hideki Iwamoto, Randy D. Blakely, and Sandra J. Rosenthal. "Biotin tethered homotryptamine derivatives: High affinity probes of the human serotonin transporter (hSERT)." Bioorganic & Medicinal Chemistry Letters 21, no. 6 (March 2011): 1678–82. http://dx.doi.org/10.1016/j.bmcl.2011.01.102.

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Fisher, Derek J., Reinaldo E. Fernández, Nancy E. Adams, and Anthony T. Maurelli. "Uptake of Biotin by Chlamydia Spp. through the Use of a Bacterial Transporter (BioY) and a Host-Cell Transporter (SMVT)." PLoS ONE 7, no. 9 (September 27, 2012): e46052. http://dx.doi.org/10.1371/journal.pone.0046052.

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Sabui, Subrata, Rubina Kapadia, Abhisek Ghosal, Michael Schneider, Nils W. G. Lambrecht, and Hamid M. Said. "Biotin and pantothenic acid oversupplementation to conditional SLC5A6 KO mice prevents the development of intestinal mucosal abnormalities and growth defects." American Journal of Physiology-Cell Physiology 315, no. 1 (July 1, 2018): C73—C79. http://dx.doi.org/10.1152/ajpcell.00319.2017.

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Intestinal absorption of the water-soluble vitamins biotin and pantothenic acid is carrier mediated and involves the sodium-dependent multivitamin transporter (SMVT; product of the SLC5A6 gene). We recently observed that intestinal-specific (conditional) knockout of the mouse Slc5a6 gene (SMVT-cKO) is associated with growth retardation, the development of spontaneous and severe inflammation, abnormal histology in the large intestine, altered gut permeability, and early death. Our aim in this study was to examine the possibility that biotin and pantothenic acid oversupplementation (BPS) of the SMVT-cKO mice could reverse the above-described abnormalities. BPS was provided in the drinking water to mice before conception, to dams during pregnancy and lactation, and to the SMVT-cKO mice throughout their life. Our findings showed that such a regimen prevents early death, as well as normalizes the growth rate, intestinal integrity, pathology, and inflammation in SMVT-cKO mice. These findings provide clear evidence for a role for biotin and/or pantothenic acid in the maintenance of normal intestinal integrity and health.

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Chatterjee, Nabendu S., Chandira K. Kumar, Alvaro Ortiz, Stanley A. Rubin, and Hamid M. Said. "Molecular mechanism of the intestinal biotin transport process." American Journal of Physiology-Cell Physiology 277, no. 4 (October 1, 1999): C605—C613. http://dx.doi.org/10.1152/ajpcell.1999.277.4.c605.

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Previous studies have characterized different aspects of the cellular/membrane mechanism and regulation of the intestinal uptake process of the water-soluble vitamin biotin. Little, however, is known about the molecular mechanisms of the uptake process. In this study, we have identified a cDNA from rat small intestine that appears to be involved in biotin transport. The open reading frame of this cloned cDNA consisted of 1,905 bases and was identical to that identified for the vitamin transporter in placental tissue. Significant heterogeneity, however, was found in the 5′ untranslated region of this clone, with three distinct variants (II, III, IV) being identified in the small intestine; the placental variant (variant I), however, was not present in the small gut. Variant II was found to be the predominant form expressed in the rat small and large intestines. Functional identity of the cloned intestinal cDNA was confirmed by stable expression in COS-7 cells, which showed a four- to fivefold increase in biotin uptake in transfected COS-7 cells compared with controls. The induced biotin uptake in transfected COS-7 cells was found to be 1) Na+ dependent, 2) saturable as a function of concentration with an apparent K m of 8.77 μM and a V max of 779.7 pmol ⋅ mg protein−1 ⋅ 3 min−1, and 3) inhibited by unlabeled biotin and pantothenic acid and their structural analogs. The distribution of complementary mRNA transcripts of the cloned cDNA along the vertical and longitudinal axes of the intestinal tract was also determined. Results of this study describe the molecular characteristics of the intestinal biotin absorption process and report the identification of a cDNA that encodes a Na+-dependent biotin uptake carrier that appears to exist in the form of multiple variants.

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Subramanian, Veedamali S., Sandeep B. Subramanya, and Hamid M. Said. "Chronic alcohol exposure negatively impacts the physiological and molecular parameters of the renal biotin reabsorption process." American Journal of Physiology-Renal Physiology 300, no. 3 (March 2011): F611—F617. http://dx.doi.org/10.1152/ajprenal.00707.2010.

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Normal body homeostasis of biotin is critically dependent on its renal recovery by kidney proximal tubular epithelial cells, a process that is mediated by the sodium-dependent multivitamin transporter (SMVT; a product of the SLC5A6 gene). Chronic ethanol consumption interferes with the renal reabsorption process of a variety of nutrients, including water-soluble vitamins. To date, however, there is nothing known about the effect of chronic alcohol feeding on physiological and molecular parameters of the renal biotin reabsorption process. We addressed these issues using rats and transgenic mice carrying the human SLC5A6 (P1P2) 5′-regulatory region as an in vivo model systems of alcohol exposure, and cultured human renal proximal tubular epithelial HK-2 cells chronically exposed to alcohol as an in vitro model of alcohol exposure. The [3H]biotin uptake results showed that chronic ethanol feeding in rats leads to a significant inhibition in carrier-mediated biotin transport across both renal brush border and basolateral membrane domains. This inhibition was associated with a marked reduction in the level of expression of SMVT protein, mRNA, and heterogenous nuclear RNA (hnRNA). Furthermore, studies with transgenic mice carrying the SLC5A6 5′-regulatory region showed that chronic alcohol feeding leads to a significant decrease in promoter activity. Studies with HK-2 cells chronically exposed to alcohol again showed a marked reduction in carrier-mediated biotin uptake, which was associated with a significant reduction in promoter activity of the human SLC5A6 5′-regulatory region. These findings demonstrate for the first time that chronic ethanol feeding inhibits renal biotin transport and that this effect is, at least in part, being exerted at the transcriptional level.

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Janoria, Kumar G., Sudharshan Hariharan, Durga Paturi, Dhananjay Pal, and Ashim K. Mitra. "Biotin Uptake by Rabbit Corneal Epithelial Cells: Role of Sodium-Dependent Multivitamin Transporter (SMVT)." Current Eye Research 31, no. 10 (January 2006): 797–809. http://dx.doi.org/10.1080/02713680600900206.

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Stelzl, Tamara, Kerstin E. Geillinger-Kästle, Jürgen Stolz, and Hannelore Daniel. "Glycans in the intestinal peptide transporter PEPT1 contribute to function and protect from proteolysis." American Journal of Physiology-Gastrointestinal and Liver Physiology 312, no. 6 (June 1, 2017): G580—G591. http://dx.doi.org/10.1152/ajpgi.00343.2016.

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Despite the fact that many membrane proteins carry extracellular glycans, little is known about whether the glycan chains also affect protein function. We recently demonstrated that the proton-coupled oligopeptide transporter 1 (PEPT1) in the intestine is glycosylated at six asparagine residues (N50, N406, N439, N510, N515, and N532). Mutagenesis-induced disruption of the individual N-glycosylation site N50, which is highly conserved among mammals, was detected to significantly enhance the PEPT1-mediated inward transport of peptides. Here, we show that for the murine protein the inhibition of glycosylation at sequon N50 by substituting N50 with glutamine, lysine, or cysteine or by replacing S52 with alanine equally altered PEPT1 transport kinetics in oocytes. Furthermore, we provide evidence that the uptake of [14C]glycyl-sarcosine in immortalized murine small intestinal (MODE-K) or colonic epithelial (PTK-6) cells stably expressing the PEPT1 transporter N50Q is also significantly increased relative to the wild-type protein. By using electrophysiological recordings and tracer flux studies, we further demonstrate that the rise in transport velocity observed for PEPT1 N50Q is bidirectional. In line with these findings, we show that attachment of biotin derivatives, comparable in weight with two to four monosaccharides, to the PEPT1 N50C transporter slows down the transport velocity. In addition, our experiments provide strong evidence that glycosylation of PEPT1 confers resistance against proteolytic cleavage by proteinase K, whereas a remarkable intrinsic stability against trypsin, even in the absence of N-linked glycans, was detected. NEW & NOTEWORTHY This study highlights the role of N50-linked glycans in modulating the bidirectional transport activity of the murine peptide transporter PEPT1. Electrophysiological and tracer flux measurements in Xenopus oocytes have shown that removal of the N50 glycans increases the maximal peptide transport rate in the inward and outward directions. This effect could be largely reversed by replacement of N50 glycans with structurally dissimilar biotin derivatives. In addition, N-glycans were detected to stabilize PEPT1 against proteolytic cleavage.

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Srinivasan, Padmanabhan, Rubina Kapadia, Arundhati Biswas, and Hamid M. Said. "Chronic alcohol exposure inhibits biotin uptake by pancreatic acinar cells: possible involvement of epigenetic mechanisms." American Journal of Physiology-Gastrointestinal and Liver Physiology 307, no. 9 (November 1, 2014): G941—G949. http://dx.doi.org/10.1152/ajpgi.00278.2014.

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Chronic exposure to alcohol affects different physiological aspects of pancreatic acinar cells (PAC), but its effect on the uptake process of biotin is not known. We addressed this issue using mouse-derived pancreatic acinar 266-6 cells chronically exposed to alcohol and wild-type and transgenic mice (carrying the human SLC5A6 5′-promoter) fed alcohol chronically. First we established that biotin uptake by PAC is Na+ dependent and carrier mediated and involves sodium-dependent multivitamin transporter (SMVT). Chronic exposure of 266-6 cells to alcohol led to a significant inhibition in biotin uptake, expression of SMVT protein, and mRNA as well as in the activity of the SLC5A6 promoter. Similarly, chronic alcohol feeding of wild-type and transgenic mice carrying the SLC5A6 promoter led to a significant inhibition in biotin uptake by PAC, as well as in the expression of SMVT protein and mRNA and the activity of the SLC5A6 promoters expressed in the transgenic mice. We also found that chronic alcohol feeding of mice is associated with a significant increase in the methylation status of CpG islands predicted to be in the mouse Slc5a6 promoters and a decrease in the level of expression of transcription factor KLF-4, which plays an important role in regulating SLC5A6 promoter activity. These results demonstrate, for the first time, that chronic alcohol exposure negatively impacts biotin uptake in PAC and that this effect is exerted (at least in part) at the level of transcription of the SLC5A6 gene and may involve epigenetic/molecular mechanisms.

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Bolla, Pradeep Kumar, Vrinda Gote, Mahima Singh, Manan Patel, Bradley A. Clark, and Jwala Renukuntla. "Lutein-Loaded, Biotin-Decorated Polymeric Nanoparticles Enhance Lutein Uptake in Retinal Cells." Pharmaceutics 12, no. 9 (August 24, 2020): 798. http://dx.doi.org/10.3390/pharmaceutics12090798.

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Age related macular degeneration (AMD) is one of the leading causes of visual loss and is responsible for approximately 9% of global blindness. It is a progressive eye disorder seen in elderly people (>65 years) mainly affecting the macula. Lutein, a carotenoid, is an antioxidant, and has shown neuroprotective properties in the retina. However, lutein has poor bioavailability owing to poor aqueous solubility. Drug delivery to the posterior segment of the eye is challenging due to the blood–retina barrier. Retinal pigment epithelium (RPE) expresses the sodium-dependent multivitamin transporter (SMVT) transport system which selectively uptakes biotin by active transport. In this study, we aimed to enhance lutein uptake into retinal cells using PLGA–PEG–biotin nanoparticles. Lutein loaded polymeric nanoparticles were prepared using O/W solvent-evaporation method. Particle size and zeta potential (ZP) were determined using Malvern Zetasizer. Other characterizations included differential scanning calorimetry, FTIR, and in-vitro release studies. In-vitro uptake and cytotoxicity studies were conducted in ARPE-19 cells using flow cytometry and confocal microscopy. Lutein was successfully encapsulated into PLGA and PLGA–PEG–biotin nanoparticles (<250 nm) with uniform size distribution and high ZP. The entrapment efficiency of lutein was ≈56% and ≈75% for lutein-loaded PLGA and PLGA–PEG–biotin nanoparticles, respectively. FTIR and DSC confirmed encapsulation of lutein into nanoparticles. Cellular uptake studies in ARPE-19 cells confirmed a higher uptake of lutein with PLGA–PEG–biotin nanoparticles compared to PLGA nanoparticles and lutein alone. In vitro cytotoxicity results confirmed that the nanoparticles were safe, effective, and non-toxic. Findings from this study suggest that lutein-loaded PLGA–PEG–biotin nanoparticles can be potentially used for treatment of AMD for higher lutein uptake.

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Yang, Julianne C., Subrata Sabui, Jonathan Skupsky Jonathan P. Jacobs, and Hamid M. Said. "Sa597 DIETARY-INDUCED BIOTIN DEFICIENCY AND TAMOXIFEN-INDUCED, INTESTINE-SPECIFIC DELETION OF THE BIOTIN TRANSPORTER IN ADULT MICE LEAD TO GUT MICROBIOME PERTURBATIONS." Gastroenterology 160, no. 6 (May 2021): S—567. http://dx.doi.org/10.1016/s0016-5085(21)02053-9.

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41

Finkenwirth, Friedrich, Michael Sippach, Heidi Landmesser, Franziska Kirsch, Anastasia Ogienko, Miriam Grunzel, Cornelia Kiesler, Heinz-Jürgen Steinhoff, Erwin Schneider, and Thomas Eitinger. "ATP-dependent Conformational Changes Trigger Substrate Capture and Release by an ECF-type Biotin Transporter." Journal of Biological Chemistry 290, no. 27 (May 19, 2015): 16929–42. http://dx.doi.org/10.1074/jbc.m115.654343.

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42

Sabui, Subrata, Jonathan Skupsky, Rubina Kapadia, Kyle Cogburn, Nils W. Lambrecht, Anshu Agrawal, and Hamid M. Said. "Tamoxifen-induced, intestinal-specific deletion of Slc5a6 in adult mice leads to spontaneous inflammation: involvement of NF-κB, NLRP3, and gut microbiota." American Journal of Physiology-Gastrointestinal and Liver Physiology 317, no. 4 (October 1, 2019): G518—G530. http://dx.doi.org/10.1152/ajpgi.00172.2019.

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The sodium-dependent multivitamin transporter (SMVT; SLC5A6) is involved in intestinal absorption of vitamin B7 (biotin). We have previously shown that mice with an embryonic intestinal-specific SMVT knockout (KO) develop biotin deficiency and severe spontaneous intestinal inflammation in addition to growth retardation, developmental delays, and death within the first 6–7 wk of life. The profound morbidity and mortality associated with the SMVT-KO has limited our ability to further characterize the intestinal inflammation and other sequelae of this deletion in adult mice with a mature gut microbiota. To overcome this limitation, we generated an intestine-specific, tamoxifen-inducible, conditional SMVT-KO (SMVT-icKO). Our results showed that adult SMVT-icKO mice have reduced body weight, biotin deficiency, shorter colonic length, and bloody diarrhea compared with age- and sex-matched control littermates. All SMVT-icKO mice also developed spontaneous intestinal inflammation associated with induction of calprotectin (S100a8/S100a9), proinflammatory cytokines (IL-1β, TNF-α, IFN-γ, and IL-6), and an increase in intestinal permeability. Additionally, the intestines of SMVT-icKO showed activation of the NF-κB pathway and the nucleotide-binding domain and leucine-rich repeat pyrin 3 domain (NLRP3) inflammasome. Notably, administration of broad-spectrum antibiotics reduced lethality and led to normalization of intestinal inflammation, proinflammatory cytokines, altered mucosal integrity, and reduced expression of the NLRP3 inflammasome. Overall, these findings support our conclusion that the biotin transport pathway plays an important role in the maintenance of intestinal homeostasis, and that NF-κB and the NLRP3 inflammasome, as well as gut microbiota, drive the development of intestinal inflammation when SMVT is absent. NEW & NOTEWORTHY This study demonstrates that deletion of the intestinal biotin uptake system in adult mice leads to the development of spontaneous gut inflammation and that luminal microbiota plays a role in its development.

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Nara, T., T. Kikukawa, and N. Kamo. "Chemical modification of single-Cys EmrE, a small multidrug transporter in Escherichia coli, by biotin-PE-maleimide." Seibutsu Butsuri 43, supplement (2003): S179. http://dx.doi.org/10.2142/biophys.43.s179_1.

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Alfadhel, Majid. "Early Infantile Leigh-like SLC19A3 Gene Defects Have a Poor Prognosis: Report and Review." Journal of Central Nervous System Disease 9 (January 1, 2017): 117957351773752. http://dx.doi.org/10.1177/1179573517737521.

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Solute carrier family 19 (thiamine transporter), member 3 ( SCL19A3) gene defect produces an autosomal recessive neurodegenerative disorder associated with different phenotypes and acronyms. One of the common presentations is early infantile lethal Leigh-like syndrome. We report a case of early infantile Leigh-like SLC19A3 gene defects of patients who died at 4 months of age with no response to a high dose of biotin and thiamine. In addition, we report a novel mutation that was not reported previously. Finally, we review the literature regarding early infantile Leigh-like SLC19A3 gene defects and compare the literature with our patient.

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Prasad, Puttur D., and Vadivel Ganapathy. "Keratinocytes Join Forces with Immune Cells in the Prosecution of SMVT as a “False” Biotin Transporter." Journal of Investigative Dermatology 120, no. 3 (March 2003): xi—xii. http://dx.doi.org/10.1046/j.1523-1747.2003.12075.x.

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Pittman, Julian T., Celia A. Dodd, and Bradley G. Klein. "Immunohistochemical Changes in the Mouse Striatum Induced by the Pyrethroid Insecticide Permethrin." International Journal of Toxicology 22, no. 5 (September 2003): 359–70. http://dx.doi.org/10.1177/109158180302200504.

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Epidemiological studies have linked insecticide exposure and Parkinson's disease. In addition, some insecticides produce damage or physiological disruption within the dopaminergic nigrostriatal pathway of non-humans. This study employed immunohistochemical analysis in striatum of the C57BL/6 mouse to clarify tissue changes suggested by previous pharmacological studies of the pyrethroid insecticide permethrin. Dopamine transporter, tyrosine hydroxylase, and glial fibrillary acidic protein immunoreactivities were examined in caudate-putamen to distinguish changes in amount of dopamine transporter immunoreactive protein from degeneration or other damage to dopaminergic neuropil. Weight-matched pairs of pesticide-treated and vehicle-control mice were dosed and sacrificed on the same days. Permethrin at 0.8, 1.5 and 3.0 mg/kg were the low doses and at 200 mg/kg the high dose. Brains from matched pairs of mice were processed on the same slides using the avidin-biotin technique. Four fields were morphometrically located in each of the serial sections of caudateputamen, digitally photographed, and immunopositive image pixels were counted and compared between members of matched pairs of permethrin-treated and vehicle-control mice. For low doses, only 3.0 mg/kg produced a significant decrease in dopamine transporter immunostaining. The high dose of permethrin did not produce a significant change in dopamine transporter or tyrosine hydroxylase immunostaining, but resulted in a significant increase in glial fibrillary acidic protein immunostaining. These data suggest that a low dose of permethrin can reduce the amount of dopamine transporter immunoreactive protein in the caudate-putamen. They also suggest that previously reported reductions in dopamine uptake of striatal synaptosomes of high-dose mice may be due to nondegenerative tissue damage within this region as opposed to reductions of dopamine transporter protein or death of nigrostriatal terminals. These data provide further evidence that insecticides can affect the primary neurodegenerative substrate of Parkinson's disease.

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Farrell, C. L., J. Yang, and W. M. Pardridge. "GLUT-1 glucose transporter is present within apical and basolateral membranes of brain epithelial interfaces and in microvascular endothelia with and without tight junctions." Journal of Histochemistry & Cytochemistry 40, no. 2 (February 1992): 193–99. http://dx.doi.org/10.1177/40.2.1552163.

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We investigated the diversity of cellular localization of the GLUT-1 glucose transporter protein at epithelial and endothelial barriers either possessing or lacking occluding junctions. The avidin-biotin immunoperoxidase and the immunogold-silver staining (IGSS) techniques were used. A rabbit polyclonal antiserum prepared against a synthetic peptide encoding the 13 amino acids at the carboxyl terminus of the GLUT-1 glucose transporter protein was used. Both techniques were found to have comparable sensitivity in detecting immunoreactive GLUT-1. The IGSS experiments employed a light-insensitive stabilizer, and no immunoreactive GLUT-1 was found in brain cells (neurons, glial cells), but abundant immunoreactive GLUT-1 was found in brain capillary endothelium, which is composed of cells with occluding junctions. However, immunoreactive GLUT-1 was also found in endothelium known not to contain occluding junctions, such as testicular microvascular endothelium and endothelium on the fetal side of the syncytiotrophoblast of the placenta. In epithelial barriers, GLUT-1 was also found in the basolateral membrane of renal collecting duct epithelium, choroid plexus, and the placental syncytiotrophoblast layer. However, immunoreactive GLUT-1 was found in the apical membrane of ependymal epithelium near the lower portion of the third ventricle. In conclusion, there is diversity underlying the expression of the GLUT-1 glucose transporter protein in different cell types, and the transporter protein can be found in endothelium with and without occluding junctions, and in both apical and basolateral membranes of epithelial barriers.

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Budipitojo, Teguh, Ariana, Tri Wahyu Pangestiningsih, Hery Wijayanto, Dwi Liliek Kusindarta, and Dewi Kania Musana. "Studi Distribusi Glukosa Transporter 4 pada Otot Skelet Ayam Kedu Cemani." Jurnal Sain Veteriner 35, no. 2 (December 1, 2017): 254. http://dx.doi.org/10.22146/jsv.34698.

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Glucose transporter (GLUT 4) is glucose transporter protein regulated by insulin, found in adipose tissue and striated muscle (skeletal and cardiac muscle). Kedu cemani chicken is one of Indonesia endemic animal, found in Kedu, Temanggung regency, Central Java. This study was required to complete microscopic documentation of Indonesia’s native biodiversity. The objective of this study was to clarify GLUT 4 distribution in skeletal muscle fibers of kedu cemani chicken by using avidin-biotin-peroxidase complex (ABC) immunohistochemistry method. This study was conducted by using pectorales major, biceps brachii, and biceps femoris muscle tissue from 5 kedu cemani chicken. The result showed that GLUT 4 immunoreactivity were detected in sarcolemma and myofibrils component of pectorales major, biceps brachii, and biceps femoris muscle tissue. Intensity of GLUT 4 immunorectivites increased from weak intensity in pectorales major muscle tissue, moderate intensity in biceps brachii muscle tissue, then strong intensity in biceps femoris muscle tissue. This result might motivate to further exploration about the other kedu cemani chicken specific features to complete microscopic documentation of Indonesia’s native biodiversity.

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Aluri, Srinivas, Rongbao Zhao, Andras Fiser, and I. David Goldman. "Substituted-cysteine accessibility and cross-linking identify an exofacial cleft in the 7th and 8th helices of the proton-coupled folate transporter (SLC46A1)." American Journal of Physiology-Cell Physiology 314, no. 3 (March 1, 2018): C289—C296. http://dx.doi.org/10.1152/ajpcell.00215.2017.

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The proton-coupled folate transporter (PCFT-SLC46A1) is required for folate transport across the apical membrane of the small intestine and across the choroid plexus. This study focuses on the structure/function of the 7th transmembrane domain (TMD), and its relationship to the 8th TMD as assessed by the substituted cysteine accessibility method (SCAM) and dicysteine cross-linking. Nine exofacial residues (I278C; H281C–L288C) of 23 residues in the 7th TMD were accessible to 2-((biotinoyl)amino)ethyl methanethiosulfonate (MTSEA-biotin). Pemetrexed, a high-affinity substrate for PCFT, decreased or abolished biotinylation of seven of these residues consistent with their location in or near the folate binding pocket. Homology models of PCFT based on Glut5 fructose transporter structures in both inward- and outward- open conformations were constructed and predicted that two pairs of residues (T289-I304C and Q285-Q311C) from the 7th and 8th TMDs should be in sufficiently close proximity to form a disulfide bond when substituted with cysteines. The single Cys-substituted mutants were accessible to MTSEA-biotin and functional with and without pretreatment with dithiotreitol. However, the double mutants were either not accessible at all, or accessibility was markedly reduced and function markedly impaired. This occurred spontaneously without inclusion of an oxidizing agent. Dithiotreitol restored accessibility and function consistent with disulfide bond disruption. The data establish the proximity of exofacial regions of the 7th and 8th TMDs and their role in defining the aqueous translocation pathway and suggest that these helices may be a component of an exofacial cleft through which substrates enter the protein binding pocket in its outward-open conformation.

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Tabarki, Brahim, and Majid Alfadhel. "SLC19A3 Gene Defects Sorting the Phenotype and Acronyms: Review." Neuropediatrics 49, no. 02 (September 29, 2017): 083–92. http://dx.doi.org/10.1055/s-0037-1607191.

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AbstractThiamine metabolism dysfunction syndrome type 2 is also known by other terms including: “SCL19A3 gene defect,” “biotin-responsive basal ganglia disease” (BBGD), and “biotin-thiamine–responsive basal ganglia disease” (BTBGD). The worldwide incidence and prevalence of this disorder are unknown, but the syndrome has primarily been reported in Saudi Arabia (52% of reported cases). It is caused by a defect in thiamine transporter 2 (hTHTR2), which is encoded by the SLC19A3 gene. The clinical presentations of these syndromes are heterogeneous and are likely related to the age of onset. They can be classified into three major categories: classical childhood BBGD; early-infantile Leigh-like syndrome/atypical infantile spasms; and adult Wernicke's-like encephalopathy. These variable phenotypes have common features in that all are triggered by stressors, such as fever, trauma, or vaccinations. Affected brain areas include the basal ganglia, cerebral cortex, thalamus, and periaqueductal regions. Free thiamine is a potential biomarker for diagnosis and monitoring of treatment. Definitive diagnosis is usually made by molecular testing for the SLC19A3 gene defect, and treatment consists of thiamine alone or in combination with biotin for life. In this report, we review all reported cases of the SLC19A3 gene defect, discuss the history, epidemiology, metabolic pathways, clinical phenotypes, biochemical abnormalities, brain pathology, diagnosis, genetic issues, and treatment of this devastating disorder. Finally, we recommend instituting an international registry to further the basic scientific and clinical research to elucidate multiple unanswered questions about SLC19A3 gene syndromes.

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Journal articles on the topic 'Biotin transporter'

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