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Raman, Sahadevan, Xiaoling Puyang, Tan-Yun Cheng, David C. Young, D. Branch Moody, and Robert N. Husson. "Mycobacterium tuberculosis SigM Positively Regulates Esx Secreted Protein and Nonribosomal Peptide Synthetase Genes and Down Regulates Virulence-Associated Surface Lipid Synthesis." Journal of Bacteriology 188, no. 24 (October 6, 2006): 8460–68. http://dx.doi.org/10.1128/jb.01212-06.
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ABSTRACT The Mycobacterium tuberculosis genome encodes 12 alternative sigma factors, several of which regulate stress responses and are required for virulence in animal models of acute infection. In this work we investigated M. tuberculosis SigM, a member of the extracytoplasmic function subfamily of alternative sigma factors. This sigma factor is expressed at low levels in vitro and does not appear to function in stress response regulation. Instead, SigM positively regulates genes required for the synthesis of surface or secreted molecules. Among these are genes encoding two pairs of Esx secreted proteins, a multisubunit nonribosomal peptide synthetase operon, and genes encoding two members of the proline-proline-glutamate (PPE) family of proteins. Genes up regulated in a sigM mutant strain include a different PPE gene, as well as several genes involved in surface lipid synthesis. Among these are genes involved in synthesis of phthiocerol dimycocerosate (PDIM), a surface lipid critical for virulence during acute infection, and the kasA-kasB operon, which is required for mycolic acid synthesis. Analysis of surface lipids showed that PDIM synthesis is increased in a sigM-disrupted strain and is undetectable in a sigM overexpression strain. These findings demonstrate that SigM positively and negatively regulates cell surface and secreted molecules that are likely to function in host-pathogen interactions.
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Nakunst, Diana, Christof Larisch, Andrea T. Hüser, Andreas Tauch, Alfred Pühler, and Jörn Kalinowski. "The Extracytoplasmic Function-Type Sigma Factor SigM of Corynebacterium glutamicum ATCC 13032 Is Involved in Transcription of Disulfide Stress-Related Genes." Journal of Bacteriology 189, no. 13 (May 4, 2007): 4696–707. http://dx.doi.org/10.1128/jb.00382-07.
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ABSTRACT The gene for the extracytoplasmic function (ECF) sigma factor SigM was deleted from the chromosome of the gram-positive soil bacterium Corynebacterium glutamicum to elucidate the role of the SigM protein in the regulation of gene expression. Comparative DNA microarray hybridizations of the C. glutamicum wild type and sigM-deficient mutant C. glutamicum DN1 revealed 23 genes with enhanced expression in the sigM-proficient strain, encoding functions in the assembly of iron-sulfur clusters (suf operon), thioredoxin reductase (trxB), thioredoxins (trxC, trxB1), chaperones (groES, groEL, clpB), and proteins involved in the heat shock response (hspR, dnaJ, grpE). Deletion of the sigM gene rendered the C. glutamicum cells more sensitive to heat, cold, and the presence of the thiol oxidant diamide. Transcription of the sigM gene increased under different stress conditions, including heat shock, cold shock, and disulfide stress caused by diamide treatment, suggesting a regulatory role for SigM under thiol-oxidative stress conditions. Stress-responsive promoters were determined upstream of the suf operon and of the trxB, trxC, and trxB1 genes. The deduced SigM consensus promoter is characterized by the −35 hexamer gGGAAT and the −10 hexamer YGTTGR. Transcription of the sigM gene is apparently controlled by the ECF sigma factor SigH, since a sigH mutant was unable to enhance the expression of sigM and the SigM regulon under thiol-oxidative stress conditions. A typical SigH-responsive promoter was mapped upstream of the sigM gene. The ECF sigma factor SigM is apparently part of a regulatory cascade, and its transcription is controlled by SigH under conditions of thiol-oxidative stress.
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Fernandes, Norvin D., Qi-long Wu, Dequan Kong, Xiaoling Puyang, Sumeet Garg, and Robert N. Husson. "A Mycobacterial Extracytoplasmic Sigma Factor Involved in Survival following Heat Shock and Oxidative Stress." Journal of Bacteriology 181, no. 14 (July 15, 1999): 4266–74. http://dx.doi.org/10.1128/jb.181.14.4266-4274.1999.
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ABSTRACT Extracytoplasmic function (ECF) sigma factors are a heterogeneous group of alternative sigma factors that regulate gene expression in response to a variety of conditions, including stress. We previously characterized a mycobacterial ECF sigma factor, SigE, that contributes to survival following several distinct stresses. A gene encoding a closely related sigma factor, sigH, was cloned fromMycobacterium tuberculosis and Mycobacterium smegmatis. A single copy of this gene is present in these and other fast- and slow-growing mycobacteria, including M. fortuitum and M. avium. While the M. tuberculosis and M. smegmatis sigH genes encode highly similar proteins, there are multiple differences in adjacent genes. The single in vivo transcriptional start site identified inM. smegmatis and one of two identified in M. bovis BCG were found to have −35 promoter sequences that match the ECF-dependent −35 promoter consensus. Expression from these promoters was strongly induced by 50°C heat shock. In comparison to the wild type, an M. smegmatis sigH mutant was found to be more susceptible to cumene hydroperoxide stress but to be similar in logarithmic growth, stationary-phase survival, and survival following several other stresses. Survival of an M. smegmatis sigH sigE double mutant was found to be markedly decreased following 53°C heat shock and following exposure to cumene hydroperoxide. Expression of the second gene in the sigH operon is required for complementation of the sigH stress phenotypes. SigH is an alternative sigma factor that plays a role in the mycobacterial stress response.
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Singh, Rakesh Kumar, Lav Kumar Jaiswal, Tanmayee Nayak, Ravindra Singh Rawat, Sanjit Kumar, Sachchida Nand Rai, and Ankush Gupta. "Expression, Purification, and In Silico Characterization of Mycobacterium smegmatis Alternative Sigma Factor SigB." Disease Markers 2022 (May 20, 2022): 1–11. http://dx.doi.org/10.1155/2022/7475704.
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Sigma factor B (SigB), an alternative sigma factor (ASF), is very similar to primary sigma factor SigA (σ70) but dispensable for growth in both Mycobacterium smegmatis (Msmeg) and Mycobacterium tuberculosis (Mtb). It is involved in general stress responses including heat, oxidative, surface, starvation stress, and macrophage infections. Despite having an extremely short half-life, SigB tends to operate downstream of at least three stress-responsive extra cytoplasmic function (ECF) sigma factors (SigH, SigE, SigL) and SigF involved in multiple signaling pathways. There is very little information available regarding the regulation of SigB sigma factor and its interacting protein partners. Hence, we cloned the SigB gene into pET28a vector and optimized its expression in three different strains of E. coli, viz., (BL21 (DE3), C41 (DE3), and CodonPlus (DE3)). We also optimized several other parameters for the expression of recombinant SigB including IPTG concentration, temperature, and time duration. We achieved the maximum expression of SigB at 25°C in the soluble fraction of the cell which was purified by affinity chromatography using Ni-NTA and further confirmed by Western blotting. Further, structural characterization demonstrates the instability of SigB in comparison to SigA that is carried out using homology modeling and structure function relationship. We have done protein-protein docking of RNA polymerase (RNAP) of Msmeg and SigB. This effort provides a platform for pulldown assay, structural, and other studies with the recombinant protein to deduce the SigB interacting proteins, which might pave the way to study its signaling networks along with its regulation.
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White, Mark J., Hongjun He, Renee M. Penoske, Sally S. Twining, and Thomas C. Zahrt. "PepD Participates in the Mycobacterial Stress Response Mediated through MprAB and SigE." Journal of Bacteriology 192, no. 6 (January 8, 2010): 1498–510. http://dx.doi.org/10.1128/jb.01167-09.
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ABSTRACT Currently, one-third of the world's population is believed to be latently infected with Mycobacterium tuberculosis. The mechanisms by which M. tuberculosis establishes latent infection remain largely undefined. mprAB encodes a two-component signal transduction system required by M. tuberculosis for aspects of persistent infection. MprAB regulates a large and diverse group of genetic determinants in response to membrane stress, including the extracytoplasmic function (ECF) sigma factor sigE and the HtrA-like serine protease pepD. Recent studies have demonstrated that PepD functions as both a protease and chaperone in vitro. In addition, inactivation of pepD alters the virulence of M. tuberculosis in a mouse model system of infection. Here, we demonstrate that PepD plays an important role in the stress response network of Mycobacterium mediated through MprAB and SigE. In particular, we demonstrate that the protease activity of PepD requires the PDZ domain, in addition to the catalytic serine at position 317. pepD expression initiates from at least three promoters in M. tuberculosis, including one that is regulated by SigE and is located upstream of the mprA coding sequence. Deletion of pepD or mprAB in Mycobacterium smegmatis and M. tuberculosis alters the stress response phenotypes of these strains, including increasing sensitivity to SDS and cell wall antibiotics and upregulating the expression of stress-responsive determinants, including sigE. Taking these data together, we hypothesize that PepD utilizes its PDZ domain to recognize and process misfolded proteins at the cell membrane, leading to activation of the MprAB and SigE signaling pathways and subsequent establishment of a positive feedback loop that facilitates bacterial adaptation.
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Kim, Eun Sook, Ju Yeon Song, Dae Wi Kim, Keith F. Chater, and Kye Joon Lee. "A Possible Extended Family of Regulators of Sigma Factor Activity in Streptomyces coelicolor." Journal of Bacteriology 190, no. 22 (September 12, 2008): 7559–66. http://dx.doi.org/10.1128/jb.00470-08.
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ABSTRACT SCO4677 is one of a large number of similar genes in Streptomyces coelicolor that encode proteins with an HATPase_c domain resembling that of anti-sigma factors such as SpoIIAB of Bacillus subtilis. However, SCO4677 is not located close to genes likely to encode a cognate sigma or anti-anti-sigma factor. SCO4677 was found to regulate antibiotic production and morphological differentiation, both of which were significantly enhanced by the deletion of SCO4677. Through protein-protein interaction screening of candidate sigma factor partners using the yeast two-hybrid system, SCO4677 protein was found to interact with the developmentally specific σF, suggesting that it is an antagonistic regulator of σF. Two other proteins, encoded by SCO0781 and SCO0869, were found to interact with the SCO4677 anti-σF during a subsequent global yeast two-hybrid screen, and the SCO0869-SCO4677 protein-protein interaction was confirmed by coimmunoprecipitation. The SCO0781 and SCO0869 proteins resemble well-known anti-anti-sigma factors such as SpoIIAA of B. subtilis. It appears that streptomycetes may possess an extraordinary abundance of anti-sigma factors, some of which may influence diverse processes through interactions with multiple partners: a novel feature for such regulatory proteins.
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Kazmierczak, Mark J., Martin Wiedmann, and Kathryn J. Boor. "Alternative Sigma Factors and Their Roles in Bacterial Virulence." Microbiology and Molecular Biology Reviews 69, no. 4 (December 2005): 527–43. http://dx.doi.org/10.1128/mmbr.69.4.527-543.2005.
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SUMMARY Sigma factors provide promoter recognition specificity to RNA polymerase holoenzyme, contribute to DNA strand separation, and then dissociate from the core enzyme following transcription initiation. As the regulon of a single sigma factor can be composed of hundreds of genes, sigma factors can provide effective mechanisms for simultaneously regulating expression of large numbers of prokaryotic genes. One newly emerging field is identification of the specific roles of alternative sigma factors in regulating expression of virulence genes and virulence-associated genes in bacterial pathogens. Virulence genes encode proteins whose functions are essential for the bacterium to effectively establish an infection in a host organism. In contrast, virulence-associated genes can contribute to bacterial survival in the environment and therefore may enhance the capacity of the bacterium to spread to new individuals or to survive passage through a host organism. As alternative sigma factors have been shown to regulate expression of both virulence and virulence-associated genes, these proteins can contribute both directly and indirectly to bacterial virulence. Sigma factors are classified into two structurally unrelated families, the σ70 and the σ54 families. The σ70 family includes primary sigma factors (e.g., Bacillus subtilis σA) as well as related alternative sigma factors; σ54 forms a distinct subfamily of sigma factors referred to as σN in almost all species for which these proteins have been characterized to date. We present several examples of alternative sigma factors that have been shown to contribute to virulence in at least one organism. For each sigma factor, when applicable, examples are drawn from multiple species.
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S Thompson, L., and E. J Harry. "Alternative sigma factors: the master regulators." Microbiology Australia 27, no. 3 (2006): 118. http://dx.doi.org/10.1071/ma06118.
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When a bacterial cell encounters a change in environmental conditions, it responds by producing a different complement of cellular proteins. Which proteins are produced and maintained is regulated in a number of ways, including regulation of gene transcription, stabilising or degrading mRNA transcripts, post translational modifications and targeted degradation of proteins.
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Yang, Fumeng, Wenjun Wang, Qian Liu, Xizhen Wang, Guangrong Bian, Shijie Teng, and Wei Liang. "The application of Six Sigma to perform quality analyses of plasma proteins." Annals of Clinical Biochemistry: International Journal of Laboratory Medicine 57, no. 2 (December 4, 2019): 121–27. http://dx.doi.org/10.1177/0004563219892023.
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Background The Six Sigma theory is an important tool for laboratory quality management. It has been widely used in clinical chemistry, haematology and other disciplines. The aim of our study was to evaluate the analytical performance of plasma proteins by application of Sigma metric and to compare the differences among three different allowable total errors in evaluating the analytical performance of plasma proteins. Methods Three different allowable total error values were used as quality goals. Data from an external quality assessment were used as bias, and the cumulative coefficient of variation in internal quality control data was used to represent the amount of imprecision during the same period. Sigma metric of analytes was calculated using the above data. The quality goal index was calculated to provide corrected measures for continuous improvements in analytical quality. Results The Sigma metric was highest using the external quality assessment standards of China: it was sigma ≥6 or higher in 57.1% of plasma proteins. But Sigma metric was lower by using RiliBÄK or biological variation standards. IgG, C3 and C-reactive protein all required quality improvements in imprecision. A single-rule 13s for internal quality control was recommended for IgA, IgM, C4 and rheumatoid factor, whereas multiple rules (13s/22s/R4s) were recommended for IgG, C3 and C-reactive protein, according to the external quality assessment standards of China. Conclusions Different quality goals can lead to different Sigma metric for the same analyte. As the lowest acceptable standard in clinical practice, the external quality assessment standard of China can guide laboratories to formulate reasonable quality improvement programmes.
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Mortillaro, N. A., and A. E. Taylor. "Microvascular permeability to endogenous plasma proteins in the jejunum." American Journal of Physiology-Heart and Circulatory Physiology 258, no. 6 (June 1, 1990): H1650—H1654. http://dx.doi.org/10.1152/ajpheart.1990.258.6.h1650.
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Steady-state lymph flow and lymph (CL) and plasma (CP) protein concentrations were measured at venous outflow pressures of 0, 10, 20, and 30 mmHg in an autoperfused segment of cat jejunum. In addition to determining total protein concentrations in lymph and plasma, polyacrylamide gradient gel electrophoresis was used to determine lymph and plasma protein concentrations of albumin and nine other plasma proteins. The osmotic reflection coefficient (sigma d) for total proteins, albumin, and each of the nine protein fractions was estimated using CL/CP at a capillary filtration rate independent state, when 1 - CL/CP = sigma d. A sigma d of 0.83 was obtained for total proteins, a value similar to that reported for both dog jejunum and descending colon (0.85) but appreciably different from that reported for both cat stomach (0.78) and ileum (0.92). Additionally, sigma d for albumin and each of the nine plasma protein fractions (molecular radii ranging from 37 to 120 A) increased as molecular radius increased. A two-pore model composed of a ratio of 3,750 48-A radius small pores to one 250-A radius large pore describes the data obtained, with 82% of the total volume flow occurring through the small pores and 16% through the large pores.
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Parker, R. E., R. J. Roselli, and K. L. Brigham. "Effects of prolonged elevated microvascular pressure on lung fluid balance in sheep." Journal of Applied Physiology 58, no. 3 (March 1, 1985): 869–75. http://dx.doi.org/10.1152/jappl.1985.58.3.869.
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Experiments were conducted in seven chronically instrumented unanesthetized sheep to estimate the osmotic reflection coefficient (sigma d) for total proteins and the solvent-drag reflection coefficients (sigma f) for six endogenous protein fractions. We measured the lymph-to-plasma ratio of total proteins (CL/CP) and six protein fractions during base-line conditions and after left atrial pressure elevations of 24–26 h per elevation. We also monitored pulmonary arterial pressure, left atrial pressure, systemic arterial pressure, and lung lymph flow at the various levels of pulmonary microvascular pressure. Our results indicate the CL/CP may require up to 24 h to reach a true steady state. It was found that sigma d is at least 0.89 for total proteins and sigma f is at least 0.84, 0.87, 0.86, 0.92, 0.95, and 0.96 for protein fractions with effective molecular radii of 36, 39.5, 44, 66, 105, and 123 A, respectively. In addition, the sigma f values for various protein fractions obtained from this investigation are compared with the predicted values of various mathematical models of the lung microcirculation.
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Wilson, Megan J., and Iain L. Lamont. "Mutational Analysis of an Extracytoplasmic-Function Sigma Factor To Investigate Its Interactions with RNA Polymerase and DNA." Journal of Bacteriology 188, no. 5 (March 1, 2006): 1935–42. http://dx.doi.org/10.1128/jb.188.5.1935-1942.2006.
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ABSTRACT The extracytoplasmic-function (ECF) family of sigma factors comprises a large group of proteins required for synthesis of a wide variety of extracytoplasmic products by bacteria. Residues important for core RNA polymerase (RNAP) binding, DNA melting, and promoter recognition have been identified in conserved regions 2 and 4.2 of primary sigma factors. Seventeen residues in region 2 and eight residues in region 4.2 of an ECF sigma factor, PvdS from Pseudomonas aeruginosa, were selected for alanine-scanning mutagenesis on the basis of sequence alignments with other sigma factors. Fourteen of the mutations in region 2 had a significant effect on protein function in an in vivo assay. Four proteins with alterations in regions 2.1 and 2.2 were purified as His-tagged fusions, and all showed a reduced affinity for core RNAP in vitro, consistent with a role in core binding. Region 2.3 and 2.4 mutant proteins retained the ability to bind core RNAP, but four mutants had reduced or no ability to cause core RNA polymerase to bind promoter DNA in a band-shift assay, identifying residues important for DNA binding. All mutations in region 4.2 reduced the activity of PvdS in vivo. Two of the region 4.2 mutant proteins were purified, and each showed a reduced ability to cause core RNA polymerase to bind to promoter DNA. The results show that some residues in PvdS have functions equivalent to those of corresponding residues in primary sigma factors; however, they also show that several residues not shared with primary sigma factors contribute to protein function.
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Negrini, D., D. Venturoli, M. I. Townsley, and R. K. Reed. "Permeability of parietal pleura to liquid and proteins." Journal of Applied Physiology 76, no. 2 (February 1, 1994): 627–33. http://dx.doi.org/10.1152/jappl.1994.76.2.627.
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The permselectivity of the parietal pleura was determined in spontaneously breathing anesthetized rabbits and dogs. In rabbits, we injected intrapleurally 5 ml of 1-g/dl albumin solution containing 100 microCi of 131I-labeled albumin plus 100 microCi of either lactate dehydrogenase (LDH) or alpha 2–125I-macroglobulin. Dogs received 100 ml of 1-g/dl albumin solution containing 100 microCi of 131I-albumin plus 100 microCi of alpha 2–125I-macroglobulin. A transpleural pressure gradient was set, lowering the intracapsular pressure to -30 cmH2O. The solvent drag reflection coefficients (sigma f) were calculated as the ratio between tracer concentrations in capsular and pleural liquid collected at 60–180 min. In rabbits sigma f was 0.44 +/- 0.2 (SD) for albumin, 0.84 +/- 0.1 for LDH, and 0.93 +/- 0.05 for alpha 2-macroglobulin. In dogs sigma f was 0.30 +/- 0.19 for albumin and 0.53 +/- 0.15 for alpha 2-macroglobulin. The hydraulic conductivity of the parietal pleura was 2.18 +/- 1.54 microliters.h-1.cmH2O-1.cm-2 in rabbits and 1.22 +/- 1.13 microliters.h-1.cmH2O-1.cm-2 in dogs. The parietal pleura could be modeled by two pore populations with radii of 83–89 and 156-222 A. The permeability coefficient averaged 0.08–0.21 x 10(-6) cm/s for albumin, 0.06–0.09 x 10(-6) cm/s for LDH, and 0.01-0.03 x 10(-6) cm/s for alpha 2-macroglobulin.
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Maron, M. B., and C. F. Pilati. "Calculation of the reflection coefficient from measurements of endogenous vascular indicators." Journal of Applied Physiology 64, no. 4 (April 1, 1988): 1746–48. http://dx.doi.org/10.1152/jappl.1988.64.4.1746.
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The solvent drag reflection coefficient (sigma) for total proteins can be estimated by comparing the relative degrees of concentration of erythrocytes and plasma proteins that occur during fluid filtration in an isolated perfused organ. In this analysis, we evaluated the accuracy of equations proposed by Pilati and Maron [Am. J. Physiol. 247 (Heart Circ. Physiol. 16): H1-H7, 1984] and Wolf et al. [Am. J. Physiol. 253 (Heart Circ. Physiol. 22): H194-H204, 1987] to calculate sigma from these concentration changes. We calculated sigma with each equation using data generated from a mathematical model of fluid and solute flux in membranes with known sigma's. We found that the equation of Wolf et al. provided the closest approximation to the true sigma over the entire range of filtration fractions tested (0.1-0.6), with the differences between the two equations increasing with filtration fraction. At low filtration fractions, the difference in sigma obtained using either approach was found to be inconsequential. At larger filtration fractions, a closer approximation of the true sigma can be obtained using the equation of Wolf et al.
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Liberek, K., and C. Georgopoulos. "Autoregulation of the Escherichia coli heat shock response by the DnaK and DnaJ heat shock proteins." Proceedings of the National Academy of Sciences 90, no. 23 (December 1, 1993): 11019–23. http://dx.doi.org/10.1073/pnas.90.23.11019.
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All organisms respond to various forms of stress, including heat shock. The heat shock response has been universally conserved from bacteria to humans. In Escherichia coli the heat shock response is under the positive transcriptional control of the sigma 32 polypeptide and involves transient acceleration in the rate of synthesis of a few dozen genes. Three of the heat shock genes--dnaK, dnaJ, and grpE--are special because mutations in any one of these lead to constitutive levels of heat shock gene expression, implying that their products negatively autoregulate their own synthesis. The DnaK, DnaJ, and GrpE proteins have been known to function in various biological situations, including bacteriophage lambda replication. Here, we report the formation of an ATP hydrolysis-dependent complex of DnaJ, sigma 32, and DnaK proteins in vitro. This DnaJ-sigma 32-DnaK complex has been seen under different conditions, including glycerol gradient sedimentation and co-immunoprecipitation. The DnaK and DnaJ proteins in the presence of ATP can interfere with the efficient binding of sigma 32 to the RNA polymerase core, and are capable of disrupting a preexisting sigma 32-RNA polymerase complex. Our results suggest a possible mechanism for the autoregulation of the heat shock response.
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Gao, Forson, Amy E. Danson, Fuzhou Ye, Milija Jovanovic, Martin Buck, and Xiaodong Zhang. "Bacterial Enhancer Binding Proteins—AAA+ Proteins in Transcription Activation." Biomolecules 10, no. 3 (February 25, 2020): 351. http://dx.doi.org/10.3390/biom10030351.
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Bacterial enhancer-binding proteins (bEBPs) are specialised transcriptional activators. bEBPs are hexameric AAA+ ATPases and use ATPase activities to remodel RNA polymerase (RNAP) complexes that contain the major variant sigma factor, σ54 to convert the initial closed complex to the transcription competent open complex. Earlier crystal structures of AAA+ domains alone have led to proposals of how nucleotide-bound states are sensed and propagated to substrate interactions. Recently, the structure of the AAA+ domain of a bEBP bound to RNAP-σ54-promoter DNA was revealed. Together with structures of the closed complex, an intermediate state where DNA is partially loaded into the RNAP cleft and the open promoter complex, a mechanistic understanding of how bEBPs use ATP to activate transcription can now be proposed. This review summarises current structural models and the emerging understanding of how this special class of AAA+ proteins utilises ATPase activities to allow σ54-dependent transcription initiation.
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Alhadlaq, Meshari Ahmed, Jeffrey Green, and Bassam K. Kudhair. "Analysis of Kytococcus sedentarius Strain Isolated from a Dehumidifier Operating in a University Lecture Theatre: Systems for Aerobic Respiration, Resisting Osmotic Stress, and Sensing Nitric Oxide." Microbial Physiology 31, no. 2 (2021): 135–45. http://dx.doi.org/10.1159/000512751.
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A strain of <i>Kytococcus sedentarius</i> was isolated from a dehumidifier operating in a university lecture theatre. Genome analysis and phenotypic characterisation showed that this strain, <i>K. sedentarius</i> MBB13, was a moderately halotolerant aerobe with a branched aerobic electron transport chain and genes that could contribute to erythromycin resistance. The major compatible solute was glycine betaine, with ectoine and proline being deployed at higher osmolarities. Actinobacteria possess multiple WhiB-like (Wbl) regulatory proteins, and <i>K. sedentarius</i> MBB13 has four (WhiB1, WhiB2, WhiB3, and WhiB7). Wbls are iron-sulfur proteins that regulate gene expression through interactions with RNA polymerase sigma factors and/or other regulatory proteins. Bacterial two-hybrid analyses suggested that WhiB1 and WhiB2, but not WhiB3 and WhiB7, interact with the C-terminal domain of the major sigma factor, σ<sup>A</sup>; no interaction was detected between any of the Wbl proteins and the only alternative sigma factors, σ<sup>B</sup>, σ<sup>H</sup>, or σ<sup>J</sup>. The interaction between σ<sup>A</sup> and WhiB1 or WhiB2 was disrupted in a heterologous system under growth conditions that produce nitric oxide and the iron-sulfur clusters of the isolated WhiB1 and WhiB2 proteins reacted with nitric oxide. Thus, <i>K. sedentarius</i> strain exhibits the major phenotypic characteristics of the type strain and a comprehensive examination of the interactions between its four Wbl proteins and four sigma factors suggested that the Wbl proteins all operate through interaction with σ<sup>A</sup>.
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Kuwana, Ritsuko, Satoko Yamamura, Hiromi Ikejiri, Kazuo Kobayashi, Naotake Ogasawara, Kei Asai, Yoshito Sadaie, Hiromu Takamatsu, and Kazuhito Watabe. "Bacillus subtilis spoVIF (yjcC) gene, involved in coat assembly and spore resistance." Microbiology 149, no. 10 (October 1, 2003): 3011–21. http://dx.doi.org/10.1099/mic.0.26432-0.
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In systematic screening four sporulation-specific genes, yjcA, yjcB, yjcZ and yjcC, of unknown function were found in Bacillus subtilis. These genes are located just upstream of the cotVWXYZ gene cluster oriented in the opposite direction. Northern blot analysis showed that yjcA was transcribed by the SigE RNA polymerase beginning 2 h (t 2) after the onset of sporulation, and yjcB, yjcZ and yjcC were transcribed by the SigK RNA polymerase beginning at t 4 of sporulation. The transcription of yjcZ was dependent on SigK and GerE. The consensus sequences of the appropriate sigma factors were found upstream of each gene. There were putative GerE-binding sites upstream of yjcZ. Insertional inactivation of the yjcC gene resulted in a reduction in resistance of the mutant spores to lysozyme and heat. Transmission electron microscopic examination of yjcC spores revealed a defect of sporulation at stage VI, resulting in loss of spore coats. These results suggest that YjcC is involved in assembly of spore coat proteins that have roles in lysozyme resistance. It is proposed that yjcC should be renamed as spoVIF.
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Huang, Pi H., Ying J. Li, Yu P. Su, Long H. Lee, and Hung J. Liu. "Epitope mapping and functional analysis of sigma A and sigma NS proteins of avian reovirus." Virology 332, no. 2 (February 2005): 584–95. http://dx.doi.org/10.1016/j.virol.2004.12.005.
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da Silva Neto, José F., Tie Koide, Suely L. Gomes, and Marilis V. Marques. "The Single Extracytoplasmic-Function Sigma Factor of Xylella fastidiosa Is Involved in the Heat Shock Response and Presents an Unusual Regulatory Mechanism." Journal of Bacteriology 189, no. 2 (November 10, 2006): 551–60. http://dx.doi.org/10.1128/jb.00986-06.
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ABSTRACT Genome sequence analysis of the bacterium Xylella fastidiosa revealed the presence of two genes, named rpoE and rseA, predicted to encode an extracytoplasmic function (ECF) sigma factor and an anti-sigma factor, respectively. In this work, an rpoE null mutant was constructed in the citrus strain J1a12 and shown to be sensitive to exposure to heat shock and ethanol. To identify the X. fastidiosa σE regulon, global gene expression profiles were obtained by DNA microarray analysis of bacterial cells under heat shock, identifying 21 σE-dependent genes. These genes encode proteins belonging to different functional categories, such as enzymes involved in protein folding and degradation, signal transduction, and DNA restriction modification and hypothetical proteins. Several putative σE-dependent promoters were mapped by primer extension, and alignment of the mapped promoters revealed a consensus sequence similar to those of ECF sigma factor promoters of other bacteria. Like other ECF sigma factors, rpoE and rseA were shown to comprise an operon in X. fastidiosa, together with a third open reading frame (XF2241). However, upon heat shock, rpoE expression was not induced, while rseA and XF2241 were highly induced at a newly identified σE-dependent promoter internal to the operon. Therefore, unlike many other ECF sigma factors, rpoE is not autoregulated but instead positively regulates the gene encoding its putative anti-sigma factor.
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McQuade, Ryan S., Natalia Comella, and Alan D. Grossman. "Control of a Family of Phosphatase Regulatory Genes (phr) by the Alternate Sigma Factor Sigma-H ofBacillus subtilis." Journal of Bacteriology 183, no. 16 (August 15, 2001): 4905–9. http://dx.doi.org/10.1128/jb.183.16.4905-4909.2001.
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ABSTRACT A family of 11 phosphatases can help to modulate the activity of response regulator proteins in Bacillus subtilis. Downstream of seven of the rap (phosphatase) genes arephr genes, encoding secreted peptides that function as phosphatase regulators. By using fusions to lacZ and primer extension analysis, we found that six of the sevenphr genes are controlled by the alternate sigma factor sigma-H. These results expand the potential of sigma-H to contribute to the output of several response regulators by controlling expression of inhibitors of phosphatases.
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Ostrov, David A., Andrew P. Bluhm, Danmeng Li, Juveriya Qamar Khan, Megha Rohamare, Karthic Rajamanickam, Kalpana K. Bhanumathy, et al. "Highly Specific Sigma Receptor Ligands Exhibit Anti-Viral Properties in SARS-CoV-2 Infected Cells." Pathogens 10, no. 11 (November 20, 2021): 1514. http://dx.doi.org/10.3390/pathogens10111514.
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(1) Background: There is a strong need for prevention and treatment strategies for COVID-19 that are not impacted by SARS-CoV-2 mutations emerging in variants of concern. After virus infection, host ER resident sigma receptors form direct interactions with non-structural SARS-CoV-2 proteins present in the replication complex. (2) Methods: In this work, highly specific sigma receptor ligands were investigated for their ability to inhibit both SARS-CoV-2 genome replication and virus induced cellular toxicity. This study found antiviral activity associated with agonism of the sigma-1 receptor (e.g., SA4503), ligation of the sigma-2 receptor (e.g., CM398), and a combination of the two pathways (e.g., AZ66). (3) Results: Intermolecular contacts between these ligands and sigma receptors were identified by structural modeling. (4) Conclusions: Sigma receptor ligands and drugs with off-target sigma receptor binding characteristics were effective at inhibiting SARS-CoV-2 infection in primate and human cells, representing a potential therapeutic avenue for COVID-19 prevention and treatment.
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Maron, Michael B., and Cahrles F. Pilati. "Effect of papaverine on pulmonary vascular permeability to proteins." Journal of Applied Physiology 66, no. 6 (June 1, 1989): 2919. http://dx.doi.org/10.1152/jappl.1989.66.6.2919-s.
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Page 1367–1371: Michael B. Maron and Charles F. Pilati. “Effect of papaverine on pulmonary vascular permeability to proteins.” Page 1370: right-hand column, sentence beginning on line 23 should read: For example, we previously found that histamine does not decrease sigma in this preparation, although we found it to do so in an isolated perfused canine forelimb preparation (11).
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Maron, M. B., and C. F. Pilati. "Effect of papaverine on pulmonary vascular permeability to proteins." Journal of Applied Physiology 65, no. 3 (September 1, 1988): 1367–71. http://dx.doi.org/10.1152/jappl.1988.65.3.1367.
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Previous studies have suggested that papaverine, a drug commonly used in studies of transvascular fluid and solute exchange to eliminate confounding effects of changes in vascular tone, may itself increase vascular permeability. In this study, we determined the ability of papaverine to alter pulmonary vascular protein permeability by measuring the osmotic reflection coefficient (sigma) for total proteins in a canine isolated perfused left lower lung lobe (LLL) preparation. The reflection coefficient, determined by the hematocrit-protein double-indicator technique, for control LLL's was 0.83 +/- 0.04 (SE) (n = 7). In separate groups of LLL's, blood papaverine HCl concentrations of 10(-5), 10(-4), and 10(-3) M resulted in sigma's of 0.84 +/- 0.02 (n = 6), 0.73 +/- 0.04 (n = 7), and 0.53 +/- 0.04 (n = 6), respectively. When two LLL's from the 10(-4) M group with sigma's of 0.56 and 0.57 were excluded from the analysis, the average sigma for this group was 0.79 +/- 0.02. We conclude that papaverine increases protein permeability at a concentration of 10(-3) M but does so in only some lobes at 10(-4) M. These results suggest that caution be taken when using high concentrations of papaverine in fluid balance studies.
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Shadel, G. S., and D. A. Clayton. "A Saccharomyces cerevisiae mitochondrial transcription factor, sc-mtTFB, shares features with sigma factors but is functionally distinct." Molecular and Cellular Biology 15, no. 4 (April 1995): 2101–8. http://dx.doi.org/10.1128/mcb.15.4.2101.
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In Saccharomyces cerevisiae mitochondria, sc-mtTFB is a 341-amino-acid transcription factor required for initiation of transcription from mitochondrial DNA promoters. Specific transcription in vitro requires only sc-mtTFB and the bacteriophage-related core sc-mtRNA polymerase. Mutational analysis of sc-mtTFB has defined two regions of the protein that are important for normal function both in vivo and in vitro. These regions overlap portions of the protein that exhibit similarity to conserved region 2 of bacterial sigma factors. One mutation in this region of sc-mtTFB (tyrosine 108 to arginine [Y108R]) has a defective phenotype that matches that observed for mutations in the corresponding residue of Bacillus subtilis sigma A and sigma E proteins. However, mutations in the sigma 2.4-like region, including a 5-amino-acid deletion corresponding to crucial promoter-contacting amino acids of sigma factors, did not eliminate the ability of sc-mtTFB to initiate transcription specifically in vitro. This suggests a mechanism of promoter recognition for sc-mtRNA polymerase different from that used by bacterial RNA polymerases. Two mutations in a basic region of sc-mtTFB resulted in defective proteins that were virtually dependent on supercoiled DNA templates in vitro. These mutations may have disrupted a DNA-unwinding function of sc-mtTFB that is only manifested in vitro and is partially rescued by DNA supercoiling.
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Lee, Jong-Hee, Petros C. Karakousis, and William R. Bishai. "Roles of SigB and SigF in the Mycobacterium tuberculosis Sigma Factor Network." Journal of Bacteriology 190, no. 2 (November 9, 2007): 699–707. http://dx.doi.org/10.1128/jb.01273-07.
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ABSTRACTTo characterize the roles of SigB and SigF in sigma factor regulation inMycobacterium tuberculosis, we used chemically inducible recombinant strains to conditionally overexpresssigBandsigF.Using whole genomic microarray analysis and quantitative reverse transcription-PCR, we investigated the resulting global transcriptional changes aftersigBinduction, and we specifically tested the relative expression of other sigma factor genes after knock-in expression ofsigBandsigF. Overexpression ofsigBresulted in significant upregulation of genes encoding several early culture filtrate antigens (ESAT-6-like proteins), ribosomal proteins, PE-PGRS proteins, the keto-acyl synthase, KasA, and the regulatory proteins WhiB2 and IdeR. Of note, the induction ofsigBdid not alter the expression of other sigma factor genes, indicating that SigB is likely to serve as an end regulator for at least one branch of theM. tuberculosissigma factor regulatory cascade. Analysis of the 5′-untranslated region (UTR) of SigB-dependent transcripts revealed a putative consensus sequence of NGTGG-N14-18-NNGNNG. This sequence appeared upstream of bothsigB(Rv2710) and the gene following it,ideR(Rv2711), and in vitro transcription analysis with recombinant SigB-reconstituted RNA polymerase confirmed SigB-dependent transcription from each of these promoters. Knock-in expression ofsigFrevealed that only thesigCgene was significantly upregulated 6 and 12 h aftersigFinduction. The previously identified SigF promoter consensus sequence AGTTTG-N15-GGGTTT was identified in the 5′ UTR of thesigCgene, and SigF-dependent in vitro transcription of the promoter upstream ofsigCwas confirmed by using recombinant SigF-reconstituted RNA polymerase. These two knock-in recombinant strains were tested in a macrophage model of infection which showed that overexpression ofsigBandsigFresulted in reduced rates ofM. tuberculosisintracellular growth. These results define the SigB promoter consensus recognition sequence and members of the SigB regulon. Moreover, the data suggest that, in addition to serving as an end regulator in a sigma factor cascade, SigB may auto-amplify its own expression under certain conditions.
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Babu, Suganthan Rajan, Yumiko Sato, Shoji Yagi, Gyedu Ampaabeng, and Nobuo Shimamoto. "1P208 Possible contradiction of the consensus mechanism of E. coli sigma-70 and the observed activity of its N- terminus half proteins(7. Nucleic acid binding protein,Poster Session,Abstract,Meeting Program of EABS & BSJ 2006)." Seibutsu Butsuri 46, supplement2 (2006): S198. http://dx.doi.org/10.2142/biophys.46.s198_4.
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Sexton, W. L., R. J. Korthuis, and M. H. Laughlin. "Ischemia-reperfusion injury in isolated rat hindquarters." Journal of Applied Physiology 68, no. 1 (January 1, 1990): 387–92. http://dx.doi.org/10.1152/jappl.1990.68.1.387.
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The purpose of this study was to determine the suitability of the maximally vasodilated (papaverine) isolated rat hindquarters preparation to study the effects of ischemia and reperfusion on the microvasculature of skeletal muscle. The osmotic reflection coefficient for plasma proteins (sigma) and total vascular resistance (RT, mmHg.ml-1.min.100 g-1) were determined before ischemic periods of 30, 60, 120, 180, and 240 min in intact (with skin) and 30, 60, and 120 min in skinned hindquarters and again after 60 min of reperfusion. In both intact and skinned hindquarters, reductions in sigma and increases in RT were observed during reperfusion and were correlated with the ischemic period duration. After 120 min of ischemia in intact and skinned hindquarters, sigma was reduced from preischemia values of 0.92 +/- 0.02 and 0.89 +/- 0.02 to 0.61 +/- 0.03 and 0.57 +/- 0.03, respectively, whereas RT was increased from preischemia levels of 8.9 +/- 0.3 and 8.1 +/- 0.1 to 28.4 +/- 2.9 and 74.2 +/- 16.8, respectively. The increases in RT were associated with proportional increases in skeletal muscle vascular resistance. Thus, in isolated rat hindquarters, increasing the duration of ischemia results in progressive increases in the permeability to plasma proteins (decreased sigma) and RT, which are associated primarily with skeletal muscle.
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Bosso, F. J., M. B. Maron, C. F. Pilati, and D. G. Jarjoura. "Pulmonary vascular protein sieving capability after exposure to high vascular pressures." Journal of Applied Physiology 73, no. 1 (July 1, 1992): 50–58. http://dx.doi.org/10.1152/jappl.1992.73.1.50.
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We evaluated the ability of the canine in situ left lower lobe (LLL) vasculature to sieve endogenous plasma proteins of various molecular radii (34–124 A) after LLL arterial pressure had been transiently elevated to 23.8 +/- 0.9 (control group, n = 5) or 92.3 +/- 1.4 (SE) Torr (high-pressure group, n = 9) by restricting LLL venous outflow under conditions of constant flow. After LLL flow was returned to natural perfusion, left atrial pressure was elevated in step increments, and LLL lymph and blood samples were collected until filtration-independent lymph-to-plasma protein concentration ratios (CL/CP) were obtained. The osmotic reflection coefficients (sigma d) for total proteins and seven protein fractions (separated by gradient gel electrophoresis) were calculated. The average total protein sigma d of the high-pressure group [0.51 +/- 0.06 (SE)] was significantly lower than that of the control group (0.68 +/- 0.03). Several LLLs of the high-pressure group, however, exhibited normal sigma d′s. Protein fraction CL/CP′s decreased with increasing molecular radius in both groups, but the CL/CP-molecular radius relationship was displaced upward in the high-pressure group. Pore analysis suggested that the decreases in sigma d could be explained by increases in the fractional flow through a large-pore system.
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Sipahutar, Rizki Ananda, and Ismail. "Analisis Pengendalian Kualitas Produk Kemasan Makanan Ternak Dengan Metode Six Sigma Dan Analisa Kaizen di PT. Central Proteina Prima Tbk." Journal Technology and Industrial Engineering (JTIE) 1, no. 2 (February 2, 2023): 153–61. http://dx.doi.org/10.59840/jtie.v1i2.170.
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Guna menjaga image tersebut, pastinya industri ini senantiasa berinovasi dalam perihal kenaikan mutu produk yang bertujuan buat tingkatkan performansi industri dengan menggapai zero defect. Metode Six Sigma merupakan konsep statistik yang mengukur sebuah proses yang berkaitan dengan cacat ataupun kehancuran. Metode ini dinilai selaku suatu metode yang sanggup melaksanakan pendekatan secara komprehensif, yang secara dramatis hendak tingkatkan mutu dari produk yang dihasilkan. Bersumber pada hasil pengolahan informasi dengan metode Six Sigma, jumlah penciptaan 140, 330 jumlah cacat penciptaan terbanyak ada pada parameter kecacatan kandas timbang ialah sebesar 187 pcs produk dan nilai DPMO proses sebesar 824, 2476 serta nilai sigma proses sebesar 4,65 yang artinya implementasi six sigma berpengaruh positif terhadap pengendalian kualiatas kemasan di PT.Central Proteina Prima Tbk.
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Parashar, Archana, Kimberley R. Colvin, Dawn R. D. Bignell, and Brenda K. Leskiw. "BldG and SCO3548 Interact Antagonistically To Control Key Developmental Processes in Streptomyces coelicolor." Journal of Bacteriology 191, no. 8 (February 6, 2009): 2541–50. http://dx.doi.org/10.1128/jb.01695-08.
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ABSTRACT The similarity of BldG and the downstream coexpressed protein SCO3548 to anti-anti-sigma and anti-sigma factors, respectively, together with the phenotype of a bldG mutant, suggests that BldG and SCO3548 interact as part of a regulatory system to control both antibiotic production and morphological differentiation in Streptomyces coelicolor. A combination of bacterial two-hybrid, affinity purification, and far-Western analyses demonstrated that there was self-interaction of both BldG and SCO3548, as well as a direct interaction between the two proteins. Furthermore, a genetic complementation experiment demonstrated that SCO3548 antagonizes the function of BldG, similar to other anti-anti-sigma/anti-sigma factor pairs. It is therefore proposed that BldG and SCO3548 form a partner-switching pair that regulates the function of one or more sigma factors in S. coelicolor. The conservation of bldG and sco3548 in other streptomycetes demonstrates that this system is likely a key regulatory switch controlling developmental processes throughout the genus Streptomyces.
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Laborenz, Janina, Yury S. Bykov, Katharina Knöringer, Markus Räschle, Sabine Filker, Cristina Prescianotto-Baschong, Anne Spang, et al. "The ER protein Ema19 facilitates the degradation of nonimported mitochondrial precursor proteins." Molecular Biology of the Cell 32, no. 8 (April 15, 2021): 664–74. http://dx.doi.org/10.1091/mbc.e20-11-0748.
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Ema19 is an integral endoplasmic reticulum (ER) protein with relevance for mitochondrial biogenesis. This yeast, representative of the TMEM97/sigma 2 receptor family, serves as quality control factor which supports the degradation of nonimported mitochondrial proteins. This study again demonstrates the important role of the ER in mitochondrial protein targeting.
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Pampura, A. N., E. F. Zhukalina, M. A. Morenko, and O. P. Usenova. "Modern approaches to the diagnosis and management of children with allergy to cow’s milk proteins." Rossiyskiy Vestnik Perinatologii i Pediatrii (Russian Bulletin of Perinatology and Pediatrics) 68, no. 2 (April 27, 2023): 39–46. http://dx.doi.org/10.21508/1027-4065-2023-68-2-39-46.
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Allergy to cow’s milk proteins is the most common cause of allergic reactions in young children, with a significant impact on the quality of life of children and their families. The most significant biomarker of herbivore milk allergy is allergen-specific IgE (sIgE), which can be assessed both for the whole allergen (for example, cow’s milk (CM), mare’s milk, goat’s milk, etc.) and a specific molecule, included in their composition. This article focuses on the use of sIgE in infants with suspected cow’s milk protein allergy.
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Jiang, Yuan-Hong, Jia-Fong Jhang, Lori A. Birder, and Hann-Chorng Kuo. "Sensory Receptor, Inflammatory, and Apoptotic Protein Expression in the Bladder Urothelium of Patients with Different Subtypes of Interstitial Cystitis/Bladder Pain Syndrome." International Journal of Molecular Sciences 24, no. 1 (January 3, 2023): 820. http://dx.doi.org/10.3390/ijms24010820.
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The aim of this study was to investigate the expression levels of sensory receptors, inflammatory proteins, and pro-apoptotic proteins in the urothelium of non-Hunner’s interstitial cystitis (NHIC) bladders of patients with different clinical and cystoscopic phenotypes. The urothelia from the bladders of 52 NHIC patients were harvested. The expression of sensory receptors, including TRPV1, TRPV4, TRPA1, H1-receptors, and sigma-1 receptors; the inflammatory proteins p38 and tryptase; and the pro-apoptotic proteins, such as caspase-3, BAD, and BAX in the urothelium, were investigated using immunohistochemistry and Western blotting. We compared the expression levels of these proteins in NHIC subtypes according to IC symptom scores, visual analog scores of bladder pain, maximal bladder capacity, glomerulation grades, and combined maximal bladder capacity and glomerulations after cystoscopic hydrodistention. The expression levels of TRPV1, TRPV4, sigma-1, P38, tryptase, caspase-3, and BAD were significantly increased in the urothelium of IC/BPS patients compared with the expression levels in the controls. TRPV1 was significantly associated with IC symptom severity. However, no significant differences in sensory receptor expression in the IC/BPS bladders with different bladder conditions were detected. Inflammatory and pro-apoptotic protein expression levels in the urothelium were similar among the IC/BPS subgroups. This study concluded that IC/BPS patients with frequency and bladder pain complaints have higher levels of urothelial sensory receptors, and inflammatory and pro-apoptotic proteins. The expression levels of these sensory receptors, inflammatory proteins, and pro-apoptotic proteins are not significantly different among IC/BPS bladders with different conditions.
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Min, K. T., and M. D. Yudkin. "Activity of mutant sigma F proteins truncated near the C terminus." Journal of Bacteriology 174, no. 22 (1992): 7144–48. http://dx.doi.org/10.1128/jb.174.22.7144-7148.1992.
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Miyazaki, Eishi, Jong-Min Chen, Chiew Ko, and William R. Bishai. "The Staphylococcus aureus rsbW(orf159) Gene Encodes an Anti-Sigma Factor of SigB." Journal of Bacteriology 181, no. 9 (May 1, 1999): 2846–51. http://dx.doi.org/10.1128/jb.181.9.2846-2851.1999.
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ABSTRACT SigB, a newly discovered alternative sigma factor ofStaphylococcus aureus, has been shown to play an important role in stress responses and the regulation of virulence factors. ThersbW (orf159) gene is immediately upstream ofsigB. Its gene product is homologous to Bacillus subtilis RsbW which under appropriate conditions binds toB. subtilis SigB and functions as an anti-sigma factor or negative posttranslational regulator. To define the function ofS. aureus RsbW, both the S. aureus SigB and RsbW proteins were expressed in Escherichia coli and purified. Cross-linking experiments with these purified proteins revealed that RsbW was capable of specific binding to SigB. In an in vitro transcription runoff assay, RsbW prevented SigB-directed transcription from the sar P3 promoter, a known SigB-dependent promoter, and the inhibitory activity of RsbW was found to be concentration dependent. We also identified SigB promoter consensus sequences upstream of the genes encoding alkaline shock protein 23 and coagulase and have demonstrated SigB and RsbW dependence for the promoters in vitro. These results show that RsbW is a protein sequestering anti-sigma factor of S. aureus SigB and suggest that SigB activity in S. aureus is regulated posttranslationally.
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Martínez-Salazar, Jaime M., Emmanuel Salazar, Sergio Encarnación, Miguel A. Ramírez-Romero, and Javier Rivera. "Role of the Extracytoplasmic Function Sigma Factor RpoE4 in Oxidative and Osmotic Stress Responses in Rhizobium etli." Journal of Bacteriology 191, no. 13 (April 17, 2009): 4122–32. http://dx.doi.org/10.1128/jb.01626-08.
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ABSTRACT The aims of this study were to functionally characterize and analyze the transcriptional regulation and transcriptome of the Rhizobium etli rpoE4 gene. An R. etli rpoE4 mutant was sensitive to oxidative, saline, and osmotic stresses. Using transcriptional fusions, we determined that RpoE4 controls its own transcription and that it is negatively regulated by rseF (regulator of sigma rpoE4; CH03274), which is cotranscribed with rpoE4. rpoE4 expression was induced not only after oxidative, saline, and osmotic shocks, but also under microaerobic and stationary-phase growth conditions. The transcriptome analyses of an rpoE4 mutant and an rpoE4-overexpressing strain revealed that the RpoE4 extracytoplasmic function sigma factor regulates about 98 genes; 50 of them have the rpoE4 promoter motifs in the upstream regulatory regions. Interestingly, 16 of 38 genes upregulated in the rpoE4-overexpressing strain encode unknown putative cell envelope proteins. Other genes controlled by RpoE4 include rpoH2, CH00462, CH02434, CH03474, and xthA1, which encode proteins involved in the stress response (a heat shock sigma factor, a putative Mn-catalase, an alkylation DNA repair protein, pyridoxine phosphate oxidase, and exonuclease III, respectively), as well as several genes, such as CH01253, CH03555, and PF00247, encoding putative proteins involved in cell envelope biogenesis (a putative peptidoglycan binding protein, a cell wall degradation protein, and phospholipase D, respectively). These results suggest that rpoE4 has a relevant function in cell envelope biogenesis and that it plays a role as a general regulator in the responses to several kinds of stress.
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Schiff, L. A., M. L. Nibert, M. S. Co, E. G. Brown, and B. N. Fields. "Distinct binding sites for zinc and double-stranded RNA in the reovirus outer capsid protein sigma 3." Molecular and Cellular Biology 8, no. 1 (January 1988): 273–83. http://dx.doi.org/10.1128/mcb.8.1.273-283.1988.
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By atomic absorption analysis, we determined that the reovirus outer capsid protein sigma 3, which binds double-stranded RNA (dsRNA), is a zinc metalloprotein. Using Northwestern blots and a novel zinc blotting technique, we localized the zinc- and dsRNA-binding activities of sigma 3 to distinct V8 protease-generated fragments. Zinc-binding activity was contained within an amino-terminal fragment that contained a transcription factor IIIA-like zinc-binding sequence, and dsRNA-binding activity was associated with a carboxy-terminal fragment. By these techniques, new zinc- and dsRNA-binding activities were also detected in reovirus core proteins. A sequence similarity was observed between the catalytic site of the picornavirus proteases and the transcription factor IIIA-like zinc-binding site within sigma 3. We suggest that the zinc- and dsRNA-binding activities of sigma 3 may be important for its proposed regulatory effects on viral and host cell transcription and translation.
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Schiff, L. A., M. L. Nibert, M. S. Co, E. G. Brown, and B. N. Fields. "Distinct binding sites for zinc and double-stranded RNA in the reovirus outer capsid protein sigma 3." Molecular and Cellular Biology 8, no. 1 (January 1988): 273–83. http://dx.doi.org/10.1128/mcb.8.1.273.
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By atomic absorption analysis, we determined that the reovirus outer capsid protein sigma 3, which binds double-stranded RNA (dsRNA), is a zinc metalloprotein. Using Northwestern blots and a novel zinc blotting technique, we localized the zinc- and dsRNA-binding activities of sigma 3 to distinct V8 protease-generated fragments. Zinc-binding activity was contained within an amino-terminal fragment that contained a transcription factor IIIA-like zinc-binding sequence, and dsRNA-binding activity was associated with a carboxy-terminal fragment. By these techniques, new zinc- and dsRNA-binding activities were also detected in reovirus core proteins. A sequence similarity was observed between the catalytic site of the picornavirus proteases and the transcription factor IIIA-like zinc-binding site within sigma 3. We suggest that the zinc- and dsRNA-binding activities of sigma 3 may be important for its proposed regulatory effects on viral and host cell transcription and translation.
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Rippe, B., M. Townsley, J. C. Parker, and A. E. Taylor. "Osmotic reflection coefficient for total plasma protein in lung microvessels." Journal of Applied Physiology 58, no. 2 (February 1, 1985): 436–42. http://dx.doi.org/10.1152/jappl.1985.58.2.436.
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The osmotic reflection coefficient (sigma) for total plasma proteins was estimated in 11 isolated blood-perfused canine lungs. Sigma's were determined by first measuring the capillary filtration coefficient (Kf,C in ml X min-1 X 100g-1 X cmH2O-1) using increased hydrostatic pressures and time 0 extrapolation of the slope of the weight gain curve. Kf,C averaged 0.19 +/- 0.05 (mean +/- SD) for 14 separate determinations in the 11 lungs. Following a Kf,C determination, the isogravimetric capillary pressure (Pc,i) was determined and averaged 9.9 +/- 0.5 cmH2O for all controls reported in this study. Then the blood colloids in the perfusate were either diluted or concentrated. The lung either gained or lost weight, respectively, and an initial slope of the weight gain curve (delta W/delta t)0 was estimated. The change in plasma protein colloid osmotic pressure (delta IIP) was measured using a membrane osmometer. The measured delta IIP was related to the effective colloid osmotic pressure (delta IIM) by delta IIM = (delta W/delta t)0/Kf,C = sigma delta IIP. Using this relationship, sigma averaged 0.65 +/- 0.06, and the least-squares linear regression equation relating Pc,i and the measured IIP was Pc,i = -3.1 + 0.67 IIP. The mean estimate of sigma (0.65) for total plasma proteins is similar to that reported for dog lung using lymphatic protein flux analyses, although lower than estimates made in skeletal muscle using the present methods (approximately 0.95).
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Miller, Eric S., Elizabeth Kutter, Gisela Mosig, Fumio Arisaka, Takashi Kunisawa, and Wolfgang Rüger. "Bacteriophage T4 Genome." Microbiology and Molecular Biology Reviews 67, no. 1 (March 2003): 86–156. http://dx.doi.org/10.1128/mmbr.67.1.86-156.2003.
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SUMMARY Phage T4 has provided countless contributions to the paradigms of genetics and biochemistry. Its complete genome sequence of 168,903 bp encodes about 300 gene products. T4 biology and its genomic sequence provide the best-understood model for modern functional genomics and proteomics. Variations on gene expression, including overlapping genes, internal translation initiation, spliced genes, translational bypassing, and RNA processing, alert us to the caveats of purely computational methods. The T4 transcriptional pattern reflects its dependence on the host RNA polymerase and the use of phage-encoded proteins that sequentially modify RNA polymerase; transcriptional activator proteins, a phage sigma factor, anti-sigma, and sigma decoy proteins also act to specify early, middle, and late promoter recognition. Posttranscriptional controls by T4 provide excellent systems for the study of RNA-dependent processes, particularly at the structural level. The redundancy of DNA replication and recombination systems of T4 reveals how phage and other genomes are stably replicated and repaired in different environments, providing insight into genome evolution and adaptations to new hosts and growth environments. Moreover, genomic sequence analysis has provided new insights into tail fiber variation, lysis, gene duplications, and membrane localization of proteins, while high-resolution structural determination of the “cell-puncturing device,” combined with the three-dimensional image reconstruction of the baseplate, has revealed the mechanism of penetration during infection. Despite these advances, nearly 130 potential T4 genes remain uncharacterized. Current phage-sequencing initiatives are now revealing the similarities and differences among members of the T4 family, including those that infect bacteria other than Escherichia coli. T4 functional genomics will aid in the interpretation of these newly sequenced T4-related genomes and in broadening our understanding of the complex evolution and ecology of phages—the most abundant and among the most ancient biological entities on Earth.
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Reed, R. K., M. I. Townsley, and A. E. Taylor. "Estimation of capillary reflection coefficients and unique PS products in dog paw." American Journal of Physiology-Heart and Circulatory Physiology 257, no. 3 (September 1, 1989): H1037—H1041. http://dx.doi.org/10.1152/ajpheart.1989.257.3.h1037.
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Lymphatic protein flux (Js) obtained in canine hindpaws at low lymph flows were used to determine the capillary osmotic reflection coefficients (sigma d) and unique permeability surface area (PS) products for total proteins, albumin, immunoglobulin (Ig)G, and IgM. This new analysis is based on the phenomenon that when maximal diffusion occurs across the capillary membrane, the Peclet number [x = Jv(1 - sigma d)/PS] attains a unique value defined only by sigma d. The diffusive flux is maximal when the relationship between protein flux and transcapillary fluid flux (Jv) changes from a curvilinear to a linear relationship. The slope of the linear portion of this protein flux relationship was used to determine sigma d as (1 - sigma d) = delta Js/(delta JvCp), where Cp is the plasma protein concentration. With the use of sigma d, the Jv at which the maximal diffusion occurred, and the corresponding Peclet number, a unique value is obtained for the PS product. Experiments performed using lymph from canine hindpaws (n = 6) yielded sigma d's (mean +/- SD) of 0.91 +/- 0.03, 0.83 +/- 0.11, 0.96 +/- 0.03, and virtually 1 for total protein, albumin, IgG, and IgM, respectively. The corresponding PS products for total protein, albumin, and IgG were 25.0 +/- 13.2, 28.4 +/- 6.6, and 14.0 +/- 7.9 microliters.min-1.100 g-1, respectively; PS for IgM was almost zero.(ABSTRACT TRUNCATED AT 250 WORDS)
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Paul, Deborupa, and Sanmitra Ghosh. "An overview of heat-stress response regulation in Gram-negative bacteria considering Escherichia coli as a model organism." Journal of Experimental Biology and Agricultural Sciences 10, no. 1 (February 28, 2022): 190–200. http://dx.doi.org/10.18006/2022.10(1).190.200.
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Response to heat stress (HSR) is a key stress response for endurance in Escherichia coli mediated by transcriptional factor σ-32. Even though there has been extensive investigation on the contribution of proteins and chaperones in retaining protein stability in cells under stress conditions, limited information is available regarding the dynamic nature of mechanisms regulating the activity of the highly conserved heat shock proteins (Hsps). Several gene expression-based studies suggest the pivotal role of Hsp70 (DnaK) in the regulation of the expression of heat shock genes (Hsg). Direct interaction of Hsp70 with σ-32 may regulate this function in E. coli. Recent studies revealed that localization of σ-32 to the membrane interior by SRP-dependent pathway enables them to function appropriately in their role as regulators. The contributions of different cellular components including cell membrane remain unknown. Other cellular components or σ-32 interfere with polypeptides which could play a crucial role in cell survival. Sigma factor monitors and preserves outer membrane integrity of E. coli by stimulating the genes regulating outer membrane proteins (OMPs) and lipopolysaccharide (LPS) assemblage as well as through expression of small RNAs to down-regulate surplus unassembled OMPs. σ-E activity is regulated by the rate at which its membrane-encompassing anti-sigma factor, RseA is degraded. Mutations in rseA are reported to constitutively increase the sigma (E) activity that is validated at both genetic and biochemical levels. In this review, the basic mechanism of heat stress regulation in gram-negative bacteria has been elaborated using E. coli as a model organism.
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Ehrhart, I. C., and W. F. Hofman. "Pressure-dependent increase in lung vascular permeability to water but not protein." Journal of Applied Physiology 72, no. 1 (January 1, 1992): 211–18. http://dx.doi.org/10.1152/jappl.1992.72.1.211.
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Simultaneous measures of vascular permeability to fluid (capillary filtration coefficient, Kf) and to plasma proteins (solvent drag reflection coefficient, sigma) were obtained over venous pressures (Pv) from 14 to 105 Torr in the isolated ventilated canine lung lobe (n = 70) pump perfused with autologous blood. The sigma was obtained from the relative increase in the concentration of plasma proteins vs. erythrocytes during fluid filtration. Kf's were obtained from two gravimetric methods as well as from change in hematocrit. All Kf's increased (P less than 0.05) as Pv was increased. However, sigma averaged 0.59 +/- 0.01 (range 0.54–0.67) and was unchanged (P greater than 0.05) by elevation of Pv over 20–105 Torr. In 44 lobes where all three Kf measures were obtained, gravimetric measures of Kf did not differ (P greater than 0.05) and were highly correlated with Kf obtained from hematocrit change, Vf Kf (P less than 0.001). However, both weight-based Kf's exceeded Vf Kf (P less than 0.05), suggesting that fluid filtration was overestimated by rate of lung weight gain or underestimated by hematocrit change. Increased permeability to water but not to protein over Pv from 20 to 105 Torr indicates that permeability to both can change independently and is counter to the theory that elevated vascular pressure “stretches” vascular pores.
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Gordon, Nadria D., Geri L. Ottaviano, Sarah E. Connell, Gregory V. Tobkin, Crystal H. Son, Sebastian Shterental, and Amy M. Gehring. "Secreted-Protein Response to σU Activity in Streptomyces coelicolor." Journal of Bacteriology 190, no. 3 (December 7, 2007): 894–904. http://dx.doi.org/10.1128/jb.01759-07.
Full textAbstract:
ABSTRACT The filamentous bacterium Streptomyces coelicolor forms an aerial mycelium as a prerequisite to sporulation, which occurs in the aerial hyphae. Uncontrolled activity of the extracytoplasmic function sigma factor σU blocks the process of aerial mycelium formation in this organism. Using a green fluorescent protein transcriptional reporter, we have demonstrated that sigU transcription is autoregulated. We have defined a σU-dependent promoter sequence and used this to identify 22 likely σU regulon members in the S. coelicolor genome. Since many of these genes encode probable secreted proteins, we characterized the extracellular proteome of a mutant with high σU activity caused by disruption of rsuA, the presumed cognate anti-sigma factor of σU. This mutant secreted a much greater quantity and diversity of proteins than the wild-type strain. Peptide mass fingerprinting was used to identify 79 proteins from the rsuA mutant culture supernatant. The most abundant species, SCO2217, SCO0930, and SCO2207, corresponded to secreted proteins or lipoproteins of unknown functions whose genes are in the proposed σU regulon. Several unique proteases were also detected in the extracellular proteome of the mutant, and the levels of the protease inhibitor SCO0762 were much reduced compared to those of the wild type. Consequently, extracellular protease activity was elevated about fourfold in the rsuA mutant. The functions of the proteins secreted as a result of σU activity may be important for combating cell envelope stress and modulating morphological differentiation in S. coelicolor.
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Wolf, M. B., P. D. Watson, and D. R. Scott. "Integral-mass balance method for determination of solvent drag reflection coefficient." American Journal of Physiology-Heart and Circulatory Physiology 253, no. 1 (July 1, 1987): H194—H204. http://dx.doi.org/10.1152/ajpheart.1987.253.1.h194.
Full textAbstract:
We have developed the integral-mass balance (IMB) method to measure the solvent drag reflection coefficient (sigma f) for transcapillary macromolecular transport in skeletal muscle and other organs. Of course, sigma f is calculated from the cumulative amounts of water and macromolecule that move convectively across the microvascular membrane as determined from changes in hematocrit and plasma macromolecule concentration over a period of fluid filtration. We have investigated the effects of both theoretical and experimental factors that affect the validity and accuracy of the method. The effect of the following factors on sigma f determination by the IMB method were explored: low Peclet number; random-measurement errors; and systematic errors due to vascular leakage, hemolysis of red blood cells, evaporation, and osmolality changes. We found that all of these factors produced overestimations of sigma f, but their effects could be corrected. Also, appropriate experimental design could minimize these effects. Experiments using the IMB method in the isolated, perfused cat hindlimb preparation to determine sigma f for albumin and plasma proteins resulted in mean values of 0.82 +/- 0.08 (SD) (n = 7) and 0.83 +/- 0.02 (n = 4), respectively.
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Hahn, Mi-Young, Sahadevan Raman, Mauricio Anaya, and Robert N. Husson. "The Mycobacterium tuberculosis Extracytoplasmic-Function Sigma Factor SigL Regulates Polyketide Synthases and Secreted or Membrane Proteins and Is Required for Virulence." Journal of Bacteriology 187, no. 20 (October 15, 2005): 7062–71. http://dx.doi.org/10.1128/jb.187.20.7062-7071.2005.
Full textAbstract:
ABSTRACT Mycobacterium tuberculosis sigL encodes an extracytoplasmic function (ECF) sigma factor and is adjacent to a gene for a membrane protein (Rv0736) that contains a conserved HXXXCXXC sequence. This motif is found in anti-sigma factors that regulate several ECF sigma factors, including those that control oxidative stress responses. In this work, SigL and Rv0736 were found to be cotranscribed, and the intracellular domain of Rv0736 was shown to interact specifically with SigL, suggesting that Rv0736 may encode an anti-sigma factor of SigL. An M. tuberculosis sigL mutant was not more susceptible than the parental strain to several oxidative and nitrosative stresses, and sigL expression was not increased in response to these stresses. In vivo, sigL is expressed from a weak SigL-independent promoter and also from a second SigL-dependent promoter. To identify SigL-regulated genes, sigL was overexpressed and microarray analysis of global transcription was performed. Four small operons, sigL (Rv0735)-Rv0736, mpt53 (Rv2878c)-Rv2877c, pks10 (Rv1660)-pks7 (Rv1661), and Rv1139c-Rv1138c, were among the most highly upregulated genes in the sigL-overexpressing strain. SigL-dependent transcription start sites of these operons were mapped, and the consensus promoter sequences TGAACC in the −35 region and CGTgtc in the −10 region were identified. In vitro, purified SigL specifically initiated transcription from the promoters of sigL, mpt53, and pks10. Additional genes, including four PE_PGRS genes, appear to be regulated indirectly by SigL. In an in vivo murine infection model, the sigL mutant strain showed marked attenuation, indicating that the sigL regulon is important in M. tuberculosis pathogenesis.
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Mathews, Sarah A., and Peter Timms. "Identification and Mapping of Sigma-54 Promoters inChlamydia trachomatis." Journal of Bacteriology 182, no. 21 (November 1, 2000): 6239–42. http://dx.doi.org/10.1128/jb.182.21.6239-6242.2000.
Full textAbstract:
ABSTRACT The first ς54 promoters in Chlamydia trachomatis L2 were mapped upstream of hypothetical proteins CT652.1 and CT683. Comparative genomics indicated that these ς54 promoters and potential upstream activation binding sites are conserved in orthologous C. trachomatis D,C. trachomatis mouse pneumonitis strain, andChlamydia pneumoniae (CWL029 and AR39) genes.
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Reed, R. K., M. I. Townsley, R. J. Korthuis, and A. E. Taylor. "Analysis of lymphatic protein flux data. V. Unique PS products and sigma dS at low lymph flows." American Journal of Physiology-Heart and Circulatory Physiology 261, no. 3 (September 1, 1991): H728—H740. http://dx.doi.org/10.1152/ajpheart.1991.261.3.h728.
Full textAbstract:
The selectivity of the capillary membrane to protein (osmotic reflection coefficient, sigma d) can be measured at high transcapillary volume flow when the capillary membrane can be considered as a true sieve. However, the diffusive capacity of the membrane (permeability-surface area product, PS) for macromolecules has not been directly measured, only estimated by assuming that transcapillary volume flow was zero. Based on unique properties of the Peclet number, a parameter that describes the ratio of solute convective flux relative to diffusive capacity, we have developed three new techniques using lymph protein fluxes to estimate a unique PS product that is independent of transcapillary fluid flux. Two of these techniques require a measure of sigma d when the ratio of protein concentration in lymph relative to plasma is equal to (1- sigma d), which occurs at high capillary filtration rates. However, the third method allows both sigma d and the PS product to be determined at relatively low lymph flow rates, eliminating the need for high capillary pressures to determine sigma d. For each protein, these techniques yield an estimate of PS and sigma d for the total membrane. However, by analysis of several different sized proteins and estimation of small- and large-pore volume flows, sigma d and PS can be determined separately for the small- and large-pore pathways. These techniques for estimating sigma d and PS were evaluated by modeling the total solute flux of albumin and immunoglobulins G and M in a heteroporous membrane.
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Malhotra, Sonal, Laura A. Silo-Suh, Kalai Mathee, and Dennis E. Ohman. "Proteome Analysis of the Effect of Mucoid Conversion on Global Protein Expression in Pseudomonas aeruginosa Strain PAO1 Shows Induction of the Disulfide Bond Isomerase, DsbA." Journal of Bacteriology 182, no. 24 (December 15, 2000): 6999–7006. http://dx.doi.org/10.1128/jb.182.24.6999-7006.2000.
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ABSTRACT Pseudomonas aeruginosa strains that cause chronic pulmonary infections in cystic fibrosis patients typically undergo mucoid conversion. The mucoid phenotype indicates alginate overproduction and is often due to defects in MucA, an antisigma factor that controls the activity of sigma-22 (AlgT [also called AlgU]), which is required for the activation of genes for alginate biosynthesis. In this study we hypothesized that mucoid conversion may be part of a larger response that activates genes other than those for alginate synthesis. To address this, a two-dimensional (2-D) gel analysis was employed to compare total proteins in strain PAO1 to those of its mucA22 derivative, PDO300, in order to identify protein levels enhanced by mucoid conversion. Six proteins that were clearly more abundant in the mucoid strain were observed. The amino termini of such proteins were determined and used to identify the gene products in the genomic database. Proteins involved in alginate biosynthesis were expected among these, and two (AlgA and AlgD) were identified. This result verified that the 2-D gel approach could identify gene products under sigma-22 control and upregulated bymucA mutation. Two other protein spots were also clearly upregulated in the mucA22 background, and these were identified as porin F (an outer membrane protein) and a homologue of DsbA (a disulfide bond isomerase). Single-copy gene fusions were constructed to test whether these proteins were enhanced in the mucoid strain due to increased transcription. The oprF-lacZ fusion showed little difference in levels of expression in the two strains. However, the dsbA-lacZ fusion showed two- to threefold higher expression in PDO300 than in PAO1, suggesting that its promoter was upregulated by the deregulation of sigma-22 activity. AdsbA-null mutant was constructed in PAO1 and shown to have defects predicted for a cell with reduced disulfide bond isomerase activity, namely, reduction in periplasmic alkaline phosphatase activity, increased sensitivity to dithiothreitol, reduced type IV pilin-mediated twitching motility, and reduced accumulation of extracellular proteases, including elastase. Although efficient secretion of elastase in the dsbA mutant was still demonstrable, the elastase produced appeared to be unstable, possibly as a result of mispaired disulfide bonds. Disruption ofdsbA in the mucoid PDO300 background did not affect alginate production. Thus, even though dsbA is coregulated with mucoid conversion, it was not required for alginate production. This suggests that mucA mutation, which deregulates sigma-22, results in a global response that includes other factors in addition to increasing the production of alginate.