المؤلفون: Patterson, Rachel A.¹, Juarez, Michelle T.², Hermann, Anita¹, Sasik, Roman³, Hardiman, Gary³, McGinnis, William¹ wmcginnis@ucsd.edu

المصدر: PLoS ONE. Apr2013, Vol. 8 Issue 4, p1-26. 26p.

مصطلحات موضوعية: *SERINE proteinases, *PROTEOLYTIC enzymes, *ENZYME activation, *EPIDERMIS, *WOUND healing, *DROSOPHILA, *GENE expression, *MOLECULAR genetics, *IMMUNE response, *CELLULAR signal transduction

مستخلص: : After injury to the animal epidermis, a variety of genes are transcriptionally activated in nearby cells to regenerate the missing cells and facilitate barrier repair. The range and types of diffusible wound signals that are produced by damaged epidermis and function to activate repair genes during epidermal regeneration remains a subject of very active study in many animals. In Drosophila embryos, we have discovered that serine protease function is locally activated around wound sites, and is also required for localized activation of epidermal repair genes. The serine protease trypsin is sufficient to induce a striking global epidermal wound response without inflicting cell death or compromising the integrity of the epithelial barrier. We developed a trypsin wounding treatment as an amplification tool to more fully understand the changes in the Drosophila transcriptome that occur after epidermal injury. By comparing our array results with similar results on mammalian skin wounding we can see which evolutionarily conserved pathways are activated after epidermal wounding in very diverse animals. Our innovative serine protease-mediated wounding protocol allowed us to identify 8 additional genes that are activated in epidermal cells in the immediate vicinity of puncture wounds, and the functions of many of these genes suggest novel genetic pathways that may control epidermal wound repair. Additionally, our data augments the evidence that clean puncture wounding can mount a powerful innate immune transcriptional response, with different innate immune genes being activated in an interesting variety of ways. These include puncture-induced activation only in epidermal cells in the immediate vicinity of wounds, or in all epidermal cells, or specifically in the fat body, or in multiple tissues. [ABSTRACT FROM AUTHOR]

المؤلفون: Mir, Riyaz A.¹, Chauhan, Shyam S.¹ s_s_chauhan@hotmail.com

المصدر: PLoS ONE. 2011, Vol. 6 Issue 6, p1-11. 11p.

مصطلحات موضوعية: *EXTRACELLULAR matrix, *CYSTEINE proteinases, *CATHEPSINS, *ACETALDEHYDE, *CYSTIC fibrosis, *ENZYME activation, *GENE expression, *CHROMATIN, *PRECIPITIN reaction

مستخلص: Background: The imbalance between extra cellular matrix (ECM) synthesis and degradation is critical aspect of various hepatic pathologies including alcohol induced liver fibrosis. This study was carried out to investigate the effect of acetaldehyde on expression of an extra cellular matrix degrading protease cathepsin L (CTSL) in HepG2 cells. Methodology and Results: We measured the enzymatic activity, protein and, mRNA levels of CTSL in acetaldehyde treated and untreated cells. The binding of CAAT enhancer binding protein a (C/EBP a) to CTSL promoter and its key role in the transcription from this promoter and conferring responsiveness to acetaldehyde was established by site directed mutagenesis, electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP) assays and siRNA technology. Acetaldehyde treatment significantly decreased CTSL activity and protein levels in HepG2 cells. A similar decrease in the mRNA levels and promoter activity was also observed. This decrease by acetaldehyde was attributed to the fall in the liver enriched transcription factor C/EBP a levels and it's binding to the CTSL promoter. Mutagenesis of C/EBP a binding motifs revealed the key role of this factor in CTSL transcription as well as conferring responsiveness to acetaldehyde. The siRNA mediated silencing of the C/EBP a expression mimicked the effect of acetaldehyde on CTSL levels and its promoter activity. It also abolished the responsiveness of this promoter to acetaldehyde. Conclusion: Acetaldehyde down regulates the C/EBP a mediated CTSL expression in hepatic cell lines. The decreased expression of CTSL may at least in part contribute to ECM deposition in liver which is a hallmark of alcoholic liver fibrosis. [ABSTRACT FROM AUTHOR]

المؤلفون: Skjoedt, Mikkel-Ole^1,2 moskjoedt@health.sdu.dk, Palarasah, Yaseelan², Munthe-Fog, Lea¹, Jie Ma, Ying¹, Weiss, Gudrun¹, Skjodt, Karsten², Koch, Claus², Garred, Peter¹

المصدر: Immunobiology. Nov2010, Vol. 215 Issue 11, p921-931. 11p.

مصطلحات موضوعية: *SERINE proteinases, *SERUM, *ENZYME activation, *GENETIC regulation, *LECTINS, *GENE expression, *ENZYME-linked immunosorbent assay, *MONOCLONAL antibodies

مستخلص: Abstract: Background: The human lectin complement pathway (LCP) involves circulating complexes consisting of mannose-binding lectin (MBL) or ficolins in association with serine proteases named MASP-1, -2 and -3 and a non-enzymatic protein, sMAP. MASP-3 originates from the MASP1 gene through differential splicing and little is known about its biological characteristics. For this reason we expressed recombinant MASP-3 and generated specific monoclonal antibodies to establish biochemical characteristics and to determine the serum levels, the interactions with the LCP recognition molecules and the influence on complement activation of MASP-3. Methods: We expressed rMASP-3 in CHO-DG44 cells and used SDS-PAGE and Western blotting for biochemical characterization. We generated monoclonal antibodies against MASP-3 and developed a quantitative MASP-3 assay to establish the serum levels in 100 Danish blood donors. In addition we assessed the association levels between MASP-3 and Ficolin-2, -3 and MBL using both ELISA and immunoprecipitation techniques. Moreover, we assessed the influence on complement factor C4 deposition. Results: We found the mean serum MASP-3 concentration to be 6.4mg/l (range: 2–12.9mg/l) and that MASP-3 in serum is primarily found in complex with Ficolin-3. In contrast to this the MASP-3 association with Ficolin-2 and especially with MBL seems to be less evident. rMASP-3 significantly inhibited Ficolin-3 mediated C4 deposition, while the opposite was the case for rMASP-1. Conclusion: Our results show that MASP-3 is present in relatively high serum concentrations. Moreover, Ficolin-3 is the primary acceptor molecule of MASP-3 among the LCP activator molecules, but MASP-3 appears to down-regulate Ficolin-3 mediated complement activation through the lectin pathway. [ABSTRACT FROM AUTHOR]

المؤلفون: Sakaguchi, Masayoshi¹, Niimiya, Keisuke¹, Takezawa, Makoto¹, Toki, Tsutomu¹, Sugahara, Yasusato¹, Kawakita, Masao¹ bt13004@ns.kogakuin.ac.jp

المصدر: Bioscience, Biotechnology & Biochemistry. Aug2008, Vol. 72 Issue 8, p2012-2018. 7p. 2 Charts, 4 Graphs.

مصطلحات موضوعية: *ESCHERICHIA coli, *SERINE proteinases, *GENETIC regulation, *ENZYME activation, *GENE expression

مستخلص: The article presents a study on the construction of an expression system for aqualysin I, alkaline serine-protease in Escherichia coli for the efficient production of enzyme protein. It says that the mature protein is being processed from a precursor through cleavage of N- and C-terminal peptide fragments. Moreover, significant protease activities were detected during the production of aqualysin I in E.coli in crude extract.

المؤلفون: Pechan, T.¹, Ma, P.W.K.², Luthe, D.S.¹ dsluthe@ra.msstate.edu

المصدر: Protein Expression & Purification. Mar2004, Vol. 34 Issue 1, p134. 8p.

مصطلحات موضوعية: *CYSTEINE proteinases, *GENE expression, *CORN, *ENZYME activation

مستخلص: Several heterologous expression systems were tested for their ability to express a unique maize cysteine proteinase Mir1. A baculovirus-based expression system using Trichoplusia ni larvae as host resulted in the expression of Mir1 that was correctly processed and exhibited proteinase activity. Expression in Escherichia coli resulted in accumulation of Mir1, but it had limited solubility and enzymatic activity. Large quantities of Mir1 were produced when Pichia pastoris was used as the host, but the enzyme was insoluble and inactive. [Copyright &y& Elsevier]

المؤلفون: Miyake, Yuka¹, Tsuzuki, Satoshi², Fushiki, Tohru², Inouye, Kuniyo¹ inouye@kais.kyoto-u.ac.jp

المصدر: Bioscience, Biotechnology & Biochemistry. Apr2010, Vol. 74 Issue 4, p848-850. 3p. 3 Diagrams.

مصطلحات موضوعية: *HEPATOCYTE growth factor, *SERINE proteinases, *ENZYME activation, *EPITHELIAL cells, *GENE expression, *CELL lines

مستخلص: The article discusses a study which examines the necessity of hepatocyte growth factor activator inhibitor type-1 (HAI-1) in the activation of matriptase, a type II transmembrane serine protease. Findings of the study reveal that matriptase activation does not necessarily require HAI-1 as showed on a transfection experiment using Madin-Darby canine kidney epithelial cells. It also indicates that matriptase undergoes activation in intracellular environments when expressed alone in cell lines.

المؤلفون: Patterson, Rachel A., Juarez, Michelle T., Hermann, Anita, Sasik, Roman, Hardiman, Gary, McGinnis, William

المصدر: PLoS ONE; Apr2013, Vol. 8 Issue 4, p1-26, 26p

مصطلحات موضوعية: SERINE proteinases, PROTEOLYTIC enzymes, ENZYME activation, EPIDERMIS, WOUND healing, DROSOPHILA, GENE expression, MOLECULAR genetics, IMMUNE response, CELLULAR signal transduction

مستخلص: : After injury to the animal epidermis, a variety of genes are transcriptionally activated in nearby cells to regenerate the missing cells and facilitate barrier repair. The range and types of diffusible wound signals that are produced by damaged epidermis and function to activate repair genes during epidermal regeneration remains a subject of very active study in many animals. In Drosophila embryos, we have discovered that serine protease function is locally activated around wound sites, and is also required for localized activation of epidermal repair genes. The serine protease trypsin is sufficient to induce a striking global epidermal wound response without inflicting cell death or compromising the integrity of the epithelial barrier. We developed a trypsin wounding treatment as an amplification tool to more fully understand the changes in the Drosophila transcriptome that occur after epidermal injury. By comparing our array results with similar results on mammalian skin wounding we can see which evolutionarily conserved pathways are activated after epidermal wounding in very diverse animals. Our innovative serine protease-mediated wounding protocol allowed us to identify 8 additional genes that are activated in epidermal cells in the immediate vicinity of puncture wounds, and the functions of many of these genes suggest novel genetic pathways that may control epidermal wound repair. Additionally, our data augments the evidence that clean puncture wounding can mount a powerful innate immune transcriptional response, with different innate immune genes being activated in an interesting variety of ways. These include puncture-induced activation only in epidermal cells in the immediate vicinity of wounds, or in all epidermal cells, or specifically in the fat body, or in multiple tissues. [ABSTRACT FROM AUTHOR]

: Copyright of PLoS ONE is the property of Public Library of Science and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

المؤلفون: Hurley, Amanda, Smith, Mindy, Karpova, Tatiana, Hasley, Rebecca B., Belkina, Natalya, Shaw, Stephen, Balenga, Nariman, Druey, Kirk M., Nickel, Erin, Packard, Beverly, Imamichi, Hiromi, Hu, Zonghui, Follmann, Dean, McNally, James, Higgins, Jeanette, Sneller, Michael, Lane, H. Clifford, Catalfamo, Marta

المصدر: Journal of Infectious Diseases; Feb2013, Vol. 207 Issue 4, p638-650, 13p

مصطلحات موضوعية: THERAPEUTICS, HIV infections, THROMBIN, INFLAMMATION, SERINE proteinases, PROTEASE-activated receptors, ENZYME activation, T cells, GENE expression

مستخلص: Disruption of vascular integrity by trauma and other tissue insults leads to inflammation and activation of the coagulation cascade. The serine protease thrombin links these 2 processes. The proinflammatory function of thrombin is mediated by activation of protease-activated receptor 1 (PAR-1). We found that peripheral blood effector memory CD4+ and CD8+ T lymphocytes expressed PAR-1 and that expression was increased in CD8+ T cells from human immunodeficiency virus (HIV)–infected patients. Thrombin enhanced cytokine secretion in CD8+ T cells from healthy controls and HIV-infected patients. In addition, thrombin induced chemokinesis, but not chemotaxis, of CD8+ T cells, which led to structural changes, including cell polarization and formation of a structure rich in F-actin and phosphorylated ezrin-radexin-moesin proteins. These findings suggest that thrombin mediates cross-talk between the coagulation system and the adaptive immune system at sites of vascular injury through increased T-cell motility and production of proinflammatory cytokines. [ABSTRACT FROM AUTHOR]

: Copyright of Journal of Infectious Diseases is the property of Oxford University Press / USA and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

المؤلفون: Gan, J, Greenwood, SM, Cobb, SR, Bushell, TJ, Greenwood, S M, Cobb, S R, Bushell, T J

المصدر: British Journal of Pharmacology; Jul2011, Vol. 163 Issue 5, p984-994, 11p

مصطلحات موضوعية: NEURAL transmission, HIPPOCAMPUS (Brain), ENZYME activation, NEUTRAL proteinases, GENE expression, CENTRAL nervous system, LABORATORY rats

مستخلص: Background and Purpose: Proteinase-activated receptor-2 (PAR2) is widely expressed in the CNS under normal physiological conditions. However, its potential role in modulating neuronal excitability and synaptic transmission remains to be determined. Here, we have investigated whether PAR2 activation modulates synaptic activity in the hippocampus.Experimental Approach: PAR2 activation and its effect on the hippocampus were examined in rat primary cultures and acute slices using whole cell patch clamp and standard extracellular recordings, respectively.Key Results: PAR2 activation leads to a depolarization of hippocampal neurones and a paradoxical reduction in the occurrence of synaptically driven spontaneous action potentials (APs). PAR2-induced neuronal depolarization was abolished following either the inhibition of astrocytic function or antagonism of ionotropic glutamate receptors whilst the PAR2-induced decrease in AP frequency was also reduced when astrocytic function was inhibited. Furthermore, when examined in acute hippocampal slices, PAR2 activation induced a profound long-term depression of synaptic transmission that was dependent on NMDA receptor activation and was sensitive to disruption of astrocytic function.Conclusions and Implications: These novel findings show that PAR2 activation indirectly inhibits hippocampal synaptic activity and indicate that these receptors may play an active role in modulating normal physiological CNS function, in addition to their role in pathophysiological disorders. [ABSTRACT FROM AUTHOR]

: Copyright of British Journal of Pharmacology is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

المؤلفون: Park, Jinseo¹, Rhee, Sangkee^1,2 srheesnu@snu.ac.kr

المصدر: Journal of Biological Chemistry. 5/31/2013, Vol. 288 Issue 22, p15760-15770. 11p.

مصطلحات موضوعية: *COFACTORS (Biochemistry), *GENE expression, *CYSTEINE proteinases, *SYNECHOCOCCUS, *ENZYME activation, *CYANOBACTERIA

مستخلص: Succinic semialdehyde dehydrogenase (SSADH) from cyanobacterium Synechococcus differs from other SSADHs in the γ-aminobutyrate shunt. Synechococcus SSADH (SySSADH) is a TCA cycle enzyme and completes a 2-oxoglutarate dehydrogenase-deficient cyanobacterial TCA cycle through a detour metabolic pathway. SySSADH produces succinate in anNADP+-dependent manner with a single cysteine acting as the catalytic residue in the catalytic loop. Crystal structures of SySSADH were determined in their apo form, as a binary complex with NADP+ and as a ternary complex with succinic semialdehyde and NADPH, providing details about the catalytic mechanism by revealing a covalent adduct of a cofactor with the catalytic cysteine in the binary complex and a proposed thiohemiacetal intermediate in the ternary complex. Further analyses showed that SySSADH is an oxidation-sensitive enzyme and that the formation of the NADP-cysteine adduct is a kinetically preferred event that protects the catalytic cysteine from H2O2-dependent oxidative stress. These structural and functional features of SySSADH provide a molecular basis for cofactor-dependent oxidation protection in 1-Cys SSADH, which is unique relative to other 2-Cys SSADHs employing a redox-dependent formation of a disulfide bridge. [ABSTRACT FROM AUTHOR]