The Experts below are selected from a list of 127770 Experts worldwide ranked by ideXlab platform
Ulla M Wewer - One of the best experts on this subject based on the ideXlab platform.
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Selective inhibition of ADAM12 catalytic activity through engineering of tissue inhibitor of metalloproteinases (TIMP)-2
Biochemical Journal, 2010Co-Authors: Marie Kveiborg, Ulla M Wewer, Hideaki Nagase, Meng-huee Lee, Jonas Jacobsen, Gillian MurphyAbstract:The disintegrin and metalloprotease ADAM12 has important functions in normal physiology as well as in diseases, such as cancer. Little is known about how ADAM12 confers its protumorigenic effect; however, its proteolytic capacity is likely a key component. Thus, selective inhibition of ADAM12 activity may be of great value therapeutically and as an investigative tool to elucidate its mechanisms of actions. We have previously reported the inhibitory profile of tissue inhibitors of metalloproteinases (TIMPs) against ADAM12, demonstrating in addition to TIMP-3, a unique ADAM inhibitory activity of TIMP-2. These findings strongly suggest that it is feasible to design a TIMP mutant selectively inhibiting ADAM12. With this purpose, we characterized the molecular determinants of the ADAM12–TIMP complex formation as compared with known molecular requirements for TIMP-mediated inhibition of ADAM17/TACE. Kinetic analysis using a fluorescent-peptide substrate demonstrated that the molecular interactions of N-terminal domains of TIMPs (N-TIMPs) with ADAM12 and TACE are for the most part comparable, yet revealed striking unique features of TIMP-mediated ADAM12 inhibition. Intriguingly, we found that removal of the AB-loop in N-TIMP-2, which is known to impair its interaction with TACE, resulted in increased affinity to ADAM12. Importantly, using a cell-based epidermal growth factor-shedding assay, we demonstrated for the first time an inhibitory activity of TIMPs against the transmembrane ADAM12-L, verifying the distinctive inhibitory abilities of N-TIMP-2 and engineered N-TIMP-2 mutants in a cellular environment. Together, our findings support the idea that a distinctive ADAM12 inhibitor with future therapeutic potential can be designed.
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Selective inhibition of ADAM12 catalytic activity through engineering of tissue inhibitor of metalloproteinase 2 (TIMP-2)
The Biochemical journal, 2010Co-Authors: Marie Kveiborg, Ulla M Wewer, Hideaki Nagase, Meng-huee Lee, Jonas Jacobsen, Gillian MurphyAbstract:The disintegrin and metalloprotease ADAM12 has important functions in normal physiology as well as in diseases, such as cancer. Little is known about how ADAM12 confers its pro-tumorigenic effect; however, its proteolytic capacity is probably a key component. Thus selective inhibition of ADAM12 activity may be of great value therapeutically and as an investigative tool to elucidate its mechanisms of action. We have previously reported the inhibitory profile of TIMPs (tissue inhibitor of metalloproteinases) against ADAM12, demonstrating in addition to TIMP-3, a unique ADAM-inhibitory activity of TIMP-2. These findings strongly suggest that it is feasible to design a TIMP mutant selectively inhibiting ADAM12. With this purpose, we characterized the molecular determinants of the ADAM12–TIMP complex formation as compared with known molecular requirements for TIMP-mediated inhibition of ADAM17/TACE (tumour necrosis factor α-converting enzyme). Kinetic analysis using a fluorescent peptide substrate demonstrated that the molecular interactions of N-TIMPs (N-terminal domains of TIMPs) with ADAM12 and TACE are for the most part comparable, yet revealed strikingly unique features of TIMP-mediated ADAM12 inhibition. Intriguingly, we found that removal of the AB-loop in N-TIMP-2, which is known to impair its interaction with TACE, resulted in increased affinity to ADAM12. Importantly, using a cell-based epidermal growth factor-shedding assay, we demonstrated for the first time an inhibitory activity of TIMPs against the transmembrane ADAM12-L (full-length ADAM12), verifying the distinctive inhibitory abilities of N-TIMP-2 and engineered N-TIMP-2 mutants in a cellular environment. Taken together, our findings support the idea that a distinctive ADAM12 inhibitor with future therapeutic potential can be designed.
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ADAM12 syndecan 4 signaling promotes beta 1 integrin dependent cell spreading through protein kinase calpha and rhoa
Journal of Biological Chemistry, 2003Co-Authors: Charles Kumar Thodeti, Reidar Albrechtsen, Morten Grauslund, Meena Asmar, Yoshikazu Takada, Arthur M Mercurio, John R Couchman, Christer Larsson, Ulla M WewerAbstract:Abstract The ADAMs (a disintegrinand metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers β1integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent. In the present study, we investigated ADAM12/syndecan-4 signaling leading to cell spreading and stress fiber formation. We demonstrate that syndecan-4, when present in significant amounts, promotes β1 integrin-dependent cell spreading and stress fiber formation in response to rADAM12-cys. A mutant form of syndecan-4 deficient in protein kinase C (PKC)α activation or a different member of the syndecan family, syndecan-2, was unable to promote cell spreading. GF109203X and Go6976, inhibitors of PKC, completely inhibited ADAM12/syndecan-4-induced cell spreading. Expression of syndecan-4, but not syn4ΔI, resulted in the accumulation of activated β1 integrins at the cell periphery in Chinese hamster ovary β1 cells as revealed by 12G10 staining. Further, expression of myristoylated, constitutively active PKCα resulted in β1 integrin-dependent cell spreading, but additional activation of RhoA was required to induce stress fiber formation. In summary, these data provide novel insights into syndecan-4 signaling. Syndecan-4 can promote cell spreading in a β1 integrin-dependent fashion through PKCα and RhoA, and PKCα and RhoA likely function in separate pathways.
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ADAM12 syndecan 4 signaling promotes beta 1 integrin dependent cell spreading through protein kinase calpha and rhoa
Journal of Biological Chemistry, 2003Co-Authors: Charles Kumar Thodeti, Reidar Albrechtsen, Morten Grauslund, Meena Asmar, Yoshikazu Takada, Arthur M Mercurio, John R Couchman, Christer Larsson, Ulla M WewerAbstract:The ADAMs (a disintegrin and metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers beta(1) integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent. In the present study, we investigated ADAM12/syndecan-4 signaling leading to cell spreading and stress fiber formation. We demonstrate that syndecan-4, when present in significant amounts, promotes beta(1) integrin-dependent cell spreading and stress fiber formation in response to rADAM12-cys. A mutant form of syndecan-4 deficient in protein kinase C (PKC)alpha activation or a different member of the syndecan family, syndecan-2, was unable to promote cell spreading. GF109203X and Go6976, inhibitors of PKC, completely inhibited ADAM12/syndecan-4-induced cell spreading. Expression of syndecan-4, but not syn4DeltaI, resulted in the accumulation of activated beta(1) integrins at the cell periphery in Chinese hamster ovary beta1 cells as revealed by 12G10 staining. Further, expression of myristoylated, constitutively active PKCalpha resulted in beta(1) integrin-dependent cell spreading, but additional activation of RhoA was required to induce stress fiber formation. In summary, these data provide novel insights into syndecan-4 signaling. Syndecan-4 can promote cell spreading in a beta(1) integrin-dependent fashion through PKCalpha and RhoA, and PKCalpha and RhoA likely function in separate pathways.
Gillian Murphy - One of the best experts on this subject based on the ideXlab platform.
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shedding of endogenous interleukin 6 receptor il 6r is governed by a disintegrin and metalloproteinase adam proteases while a full length il 6r isoform localizes to circulating microvesicles
Journal of Biological Chemistry, 2015Co-Authors: Neele Schumacher, Dorte Meyer, Andre Mauermann, Jan Von Der Heyde, Janina Wolf, Jeanette Schwarz, Katharina Knittler, Matthias Michalek, Gillian Murphy, Christoph GarbersAbstract:Generation of the soluble interleukin-6 receptor (sIL-6R) is a prerequisite for pathogenic IL-6 trans-signaling, which constitutes a distinct signaling pathway of the pleiotropic cytokine interleukin-6 (IL-6). Although in vitro experiments using ectopically overexpressed IL-6R and candidate proteases revealed major roles for the metalloproteinases ADAM10 and ADAM17 in IL-6R shedding, the identity of the protease(s) cleaving IL-6R in more physiological settings, or even in vivo, remains unknown. By taking advantage of specific pharmacological inhibitors and primary cells from ADAM-deficient mice we established that endogenous IL-6R of both human and murine origin is shed by ADAM17 in an induced manner, whereas constitutive release of endogenous IL-6R is largely mediated by ADAM10. Although circulating IL-6R levels are altered in various diseases, the origin of blood-borne IL-6R is still poorly understood. It has been shown previously that ADAM17 hypomorphic mice exhibit unaltered levels of serum sIL-6R. Here, by quantification of serum sIL-6R in protease-deficient mice as well as human patients we also excluded ADAM10, ADAM8, neutrophil elastase, cathepsin G, and proteinase 3 from contributing to circulating sIL-6R. Furthermore, we ruled out alternative splicing of the IL-6R mRNA as a potential source of circulating sIL-6R in the mouse. Instead, we found full-length IL-6R on circulating microvesicles, establishing microvesicle release as a novel mechanism for sIL-6R generation.
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Selective inhibition of ADAM12 catalytic activity through engineering of tissue inhibitor of metalloproteinases (TIMP)-2
Biochemical Journal, 2010Co-Authors: Marie Kveiborg, Ulla M Wewer, Hideaki Nagase, Meng-huee Lee, Jonas Jacobsen, Gillian MurphyAbstract:The disintegrin and metalloprotease ADAM12 has important functions in normal physiology as well as in diseases, such as cancer. Little is known about how ADAM12 confers its protumorigenic effect; however, its proteolytic capacity is likely a key component. Thus, selective inhibition of ADAM12 activity may be of great value therapeutically and as an investigative tool to elucidate its mechanisms of actions. We have previously reported the inhibitory profile of tissue inhibitors of metalloproteinases (TIMPs) against ADAM12, demonstrating in addition to TIMP-3, a unique ADAM inhibitory activity of TIMP-2. These findings strongly suggest that it is feasible to design a TIMP mutant selectively inhibiting ADAM12. With this purpose, we characterized the molecular determinants of the ADAM12–TIMP complex formation as compared with known molecular requirements for TIMP-mediated inhibition of ADAM17/TACE. Kinetic analysis using a fluorescent-peptide substrate demonstrated that the molecular interactions of N-terminal domains of TIMPs (N-TIMPs) with ADAM12 and TACE are for the most part comparable, yet revealed striking unique features of TIMP-mediated ADAM12 inhibition. Intriguingly, we found that removal of the AB-loop in N-TIMP-2, which is known to impair its interaction with TACE, resulted in increased affinity to ADAM12. Importantly, using a cell-based epidermal growth factor-shedding assay, we demonstrated for the first time an inhibitory activity of TIMPs against the transmembrane ADAM12-L, verifying the distinctive inhibitory abilities of N-TIMP-2 and engineered N-TIMP-2 mutants in a cellular environment. Together, our findings support the idea that a distinctive ADAM12 inhibitor with future therapeutic potential can be designed.
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Selective inhibition of ADAM12 catalytic activity through engineering of tissue inhibitor of metalloproteinase 2 (TIMP-2)
The Biochemical journal, 2010Co-Authors: Marie Kveiborg, Ulla M Wewer, Hideaki Nagase, Meng-huee Lee, Jonas Jacobsen, Gillian MurphyAbstract:The disintegrin and metalloprotease ADAM12 has important functions in normal physiology as well as in diseases, such as cancer. Little is known about how ADAM12 confers its pro-tumorigenic effect; however, its proteolytic capacity is probably a key component. Thus selective inhibition of ADAM12 activity may be of great value therapeutically and as an investigative tool to elucidate its mechanisms of action. We have previously reported the inhibitory profile of TIMPs (tissue inhibitor of metalloproteinases) against ADAM12, demonstrating in addition to TIMP-3, a unique ADAM-inhibitory activity of TIMP-2. These findings strongly suggest that it is feasible to design a TIMP mutant selectively inhibiting ADAM12. With this purpose, we characterized the molecular determinants of the ADAM12–TIMP complex formation as compared with known molecular requirements for TIMP-mediated inhibition of ADAM17/TACE (tumour necrosis factor α-converting enzyme). Kinetic analysis using a fluorescent peptide substrate demonstrated that the molecular interactions of N-TIMPs (N-terminal domains of TIMPs) with ADAM12 and TACE are for the most part comparable, yet revealed strikingly unique features of TIMP-mediated ADAM12 inhibition. Intriguingly, we found that removal of the AB-loop in N-TIMP-2, which is known to impair its interaction with TACE, resulted in increased affinity to ADAM12. Importantly, using a cell-based epidermal growth factor-shedding assay, we demonstrated for the first time an inhibitory activity of TIMPs against the transmembrane ADAM12-L (full-length ADAM12), verifying the distinctive inhibitory abilities of N-TIMP-2 and engineered N-TIMP-2 mutants in a cellular environment. Taken together, our findings support the idea that a distinctive ADAM12 inhibitor with future therapeutic potential can be designed.
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The isolated N-terminal domains of TIMP-1 and TIMP-3 are insufficient for ADAM10 inhibition
Biochemical Journal, 2008Co-Authors: Magdalini Rapti, Susan J Atkinson, Meng-huee Lee, Andrew Trim, Marcia Moss, Gillian MurphyAbstract:ADAM10 is a key member of the ADAM family of disintegrin and metalloproteinases which process membrane associated proteins to soluble forms in a process known as 'shedding'. Among the major targets of ADAM10 are Notch, EphrinA2 and CD44. In many cell based studies of shedding the activity of ADAM10 appears to overlap with that of ADAM17 which has a similar active site topology relative to the other proteolytically active ADAMs. The tissue inhibitors of metalloproteinases, TIMPs, have proved useful in the study of ADAM function since TIMP-1 inhibits ADAM10 but not ADAM17, but both enzymes are inhibited by TIMP-3. In this study we show that, in comparison to ADAM17 and the MMPs, the N-terminal domains of TIMPs alone are insufficient for the inhibition of ADAM10.This knowledge could form the basis for the design of directed inhibitors against different metalloproteinases.
Yoshikazu Takada - One of the best experts on this subject based on the ideXlab platform.
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ADAM12 syndecan 4 signaling promotes beta 1 integrin dependent cell spreading through protein kinase calpha and rhoa
Journal of Biological Chemistry, 2003Co-Authors: Charles Kumar Thodeti, Reidar Albrechtsen, Morten Grauslund, Meena Asmar, Yoshikazu Takada, Arthur M Mercurio, John R Couchman, Christer Larsson, Ulla M WewerAbstract:Abstract The ADAMs (a disintegrinand metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers β1integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent. In the present study, we investigated ADAM12/syndecan-4 signaling leading to cell spreading and stress fiber formation. We demonstrate that syndecan-4, when present in significant amounts, promotes β1 integrin-dependent cell spreading and stress fiber formation in response to rADAM12-cys. A mutant form of syndecan-4 deficient in protein kinase C (PKC)α activation or a different member of the syndecan family, syndecan-2, was unable to promote cell spreading. GF109203X and Go6976, inhibitors of PKC, completely inhibited ADAM12/syndecan-4-induced cell spreading. Expression of syndecan-4, but not syn4ΔI, resulted in the accumulation of activated β1 integrins at the cell periphery in Chinese hamster ovary β1 cells as revealed by 12G10 staining. Further, expression of myristoylated, constitutively active PKCα resulted in β1 integrin-dependent cell spreading, but additional activation of RhoA was required to induce stress fiber formation. In summary, these data provide novel insights into syndecan-4 signaling. Syndecan-4 can promote cell spreading in a β1 integrin-dependent fashion through PKCα and RhoA, and PKCα and RhoA likely function in separate pathways.
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ADAM12 syndecan 4 signaling promotes beta 1 integrin dependent cell spreading through protein kinase calpha and rhoa
Journal of Biological Chemistry, 2003Co-Authors: Charles Kumar Thodeti, Reidar Albrechtsen, Morten Grauslund, Meena Asmar, Yoshikazu Takada, Arthur M Mercurio, John R Couchman, Christer Larsson, Ulla M WewerAbstract:The ADAMs (a disintegrin and metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers beta(1) integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent. In the present study, we investigated ADAM12/syndecan-4 signaling leading to cell spreading and stress fiber formation. We demonstrate that syndecan-4, when present in significant amounts, promotes beta(1) integrin-dependent cell spreading and stress fiber formation in response to rADAM12-cys. A mutant form of syndecan-4 deficient in protein kinase C (PKC)alpha activation or a different member of the syndecan family, syndecan-2, was unable to promote cell spreading. GF109203X and Go6976, inhibitors of PKC, completely inhibited ADAM12/syndecan-4-induced cell spreading. Expression of syndecan-4, but not syn4DeltaI, resulted in the accumulation of activated beta(1) integrins at the cell periphery in Chinese hamster ovary beta1 cells as revealed by 12G10 staining. Further, expression of myristoylated, constitutively active PKCalpha resulted in beta(1) integrin-dependent cell spreading, but additional activation of RhoA was required to induce stress fiber formation. In summary, these data provide novel insights into syndecan-4 signaling. Syndecan-4 can promote cell spreading in a beta(1) integrin-dependent fashion through PKCalpha and RhoA, and PKCalpha and RhoA likely function in separate pathways.
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Functional Classification of ADAMs Based on a Conserved Motif for Binding to Integrin α9β1 IMPLICATIONS FOR SPERM-EGG BINDING AND OTHER CELL INTERACTIONS
The Journal of biological chemistry, 2002Co-Authors: Koji Eto, Clotilde Huet, Takehiko Tarui, Sergey Kupriyanov, Hai Zhen Liu, Wilma Puzon-mclaughlin, Xi Ping Zhang, Dean Sheppard, Eva Engvall, Yoshikazu TakadaAbstract:Abstract ADAMs (a disintegrin and metalloproteases) are members of the metzincin superfamily of metalloproteases. Among integrins binding to disintegrin domains of ADAMs are α9β1 and αvβ3, and they bind in an RGD-independent and an RGD-dependent manner, respectively. Human ADAM15 is the only ADAM with the RGD motif in the disintegrin domain. Thus, both integrin α9β1 and αvβ3 recognize the ADAM15 disintegrin domain. We determined how these integrins recognize the ADAM15 disintegrin domain by mutational analysis. We found that the Arg481 and the Asp-Leu-Pro-Glu-Phe residues (residues 488–492) were critical for α9β1 binding, but the RGD motif (residues 484–486) was not. In contrast, the RGD motif was critical for αvβ3 binding, but the other residues flanking the RGD motif were not. As the RX 6DLPEF α9β1recognition motif (residues 481–492) is conserved among ADAMs, except for ADAM10 and 17, we hypothesized that α9β1 may recognize disintegrin domains in all ADAMs except ADAM10 and 17. Indeed we found that α9β1 bound avidly to the disintegrin domains of ADAM1, 2, 3, and 9 but not to the disintegrin domains of ADAM10 and 17. As several ADAMs have been implicated in sperm-oocyte interaction, we tested whether the functional classification of ADAMs, based on specificity for integrin α9β1, applies to sperm-egg binding. We found that the ADAM2 and 15 disintegrin domains bound to oocytes, but the ADAM17 disintegrin domain did not. Furthermore, the ADAM2 and 15 disintegrin domains effectively blocked binding of sperm to oocytes, but the ADAM17 disintegrin domain did not. These results suggest that oocytes and α9β1 have similar binding specificities for ADAMs and that α9β1, or a receptor with similar specificity, may be involved in sperm-egg interaction during fertilization. As α9β1 is a receptor for many ADAM disintegrins and α9β1 and ADAMs are widely expressed, α9β1-ADAM interaction may be of a broad biological importance.
Charles Kumar Thodeti - One of the best experts on this subject based on the ideXlab platform.
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ADAM12 syndecan 4 signaling promotes beta 1 integrin dependent cell spreading through protein kinase calpha and rhoa
Journal of Biological Chemistry, 2003Co-Authors: Charles Kumar Thodeti, Reidar Albrechtsen, Morten Grauslund, Meena Asmar, Yoshikazu Takada, Arthur M Mercurio, John R Couchman, Christer Larsson, Ulla M WewerAbstract:Abstract The ADAMs (a disintegrinand metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers β1integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent. In the present study, we investigated ADAM12/syndecan-4 signaling leading to cell spreading and stress fiber formation. We demonstrate that syndecan-4, when present in significant amounts, promotes β1 integrin-dependent cell spreading and stress fiber formation in response to rADAM12-cys. A mutant form of syndecan-4 deficient in protein kinase C (PKC)α activation or a different member of the syndecan family, syndecan-2, was unable to promote cell spreading. GF109203X and Go6976, inhibitors of PKC, completely inhibited ADAM12/syndecan-4-induced cell spreading. Expression of syndecan-4, but not syn4ΔI, resulted in the accumulation of activated β1 integrins at the cell periphery in Chinese hamster ovary β1 cells as revealed by 12G10 staining. Further, expression of myristoylated, constitutively active PKCα resulted in β1 integrin-dependent cell spreading, but additional activation of RhoA was required to induce stress fiber formation. In summary, these data provide novel insights into syndecan-4 signaling. Syndecan-4 can promote cell spreading in a β1 integrin-dependent fashion through PKCα and RhoA, and PKCα and RhoA likely function in separate pathways.
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ADAM12 syndecan 4 signaling promotes beta 1 integrin dependent cell spreading through protein kinase calpha and rhoa
Journal of Biological Chemistry, 2003Co-Authors: Charles Kumar Thodeti, Reidar Albrechtsen, Morten Grauslund, Meena Asmar, Yoshikazu Takada, Arthur M Mercurio, John R Couchman, Christer Larsson, Ulla M WewerAbstract:The ADAMs (a disintegrin and metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers beta(1) integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent. In the present study, we investigated ADAM12/syndecan-4 signaling leading to cell spreading and stress fiber formation. We demonstrate that syndecan-4, when present in significant amounts, promotes beta(1) integrin-dependent cell spreading and stress fiber formation in response to rADAM12-cys. A mutant form of syndecan-4 deficient in protein kinase C (PKC)alpha activation or a different member of the syndecan family, syndecan-2, was unable to promote cell spreading. GF109203X and Go6976, inhibitors of PKC, completely inhibited ADAM12/syndecan-4-induced cell spreading. Expression of syndecan-4, but not syn4DeltaI, resulted in the accumulation of activated beta(1) integrins at the cell periphery in Chinese hamster ovary beta1 cells as revealed by 12G10 staining. Further, expression of myristoylated, constitutively active PKCalpha resulted in beta(1) integrin-dependent cell spreading, but additional activation of RhoA was required to induce stress fiber formation. In summary, these data provide novel insights into syndecan-4 signaling. Syndecan-4 can promote cell spreading in a beta(1) integrin-dependent fashion through PKCalpha and RhoA, and PKCalpha and RhoA likely function in separate pathways.
Alexander G Beristain - One of the best experts on this subject based on the ideXlab platform.
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ADAM12 and papp a candidate regulators of trophoblast invasion and first trimester markers of healthy trophoblasts
Cell Adhesion & Migration, 2016Co-Authors: Julian K Christians, Alexander G BeristainAbstract:Proper placental development and function is crucial for a healthy pregnancy, and there has been substantial research to identify markers of placental dysfunction for the early detection of pregnancy complications. Low first-trimester levels of a disintegrin and metalloproteinase 12 (ADAM12) and pregnancy-associated plasma protein-A (PAPP-A) have been consistently associated with the subsequent development of preeclampsia and fetal growth restriction. These molecules are both metalloproteinases secreted by the placenta that cleave insulin-like growth factor binding proteins (IGFBPs), although ADAM12 also has numerous other substrates. Recent work has identified ADAM12, and particularly its shorter variant, ADAM12S, as a regulator of the migration and invasion of trophoblasts into the lining of the uterus, a critical step in normal placental development. While the mechanisms underlying this regulation are not yet clear, they may involve the liberation of heparin-binding EGF-like growth factor (HB-EGF) and/or IGFs from IGFBPs. In contrast, there has been relatively little functional work examining PAPP-A or the IGFBP substrates of ADAM12 and PAPP-A. Understanding the functions of these markers and the mechanisms underlying their association with disease could improve screening strategies and enable the development of new therapeutic interventions.
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ADAM12-directed ectodomain shedding of E-cadherin potentiates trophoblast fusion
Cell Death & Differentiation, 2015Co-Authors: Mohsen Aghababaei, K Hogg, S Perdu, W P Robinson, Alexander G BeristainAbstract:Trophoblasts, placental cells of epithelial lineage, undergo extensive differentiation to form the cellular components of the placenta. Trophoblast progenitor cell differentiation into the multinucleated syncytiotrophoblast is a key developmental process required for placental function, where defects in syncytiotrophoblast formation and turnover associate with placental pathologies and link to poor pregnancy outcomes. The cellular and molecular processes governing syncytiotrophoblast formation are poorly understood, but require the activation of pathways that direct cell fusion. The protease, A Disintegrin and Metalloproteinase 12 (ADAM12), controls cell fusion in myoblasts and is highly expressed in the placenta localizing to multiple trophoblast populations. However, the importance of ADAM12 in regulating trophoblast fusion is unknown. Here, we describe a function for ADAM12 in regulating trophoblast fusion. Using two distinct trophoblast models of cell fusion, we show that ADAM12 is dynamically upregulated and is under the transcriptional control of protein kinase A. siRNA-directed loss of ADAM12 impedes spontaneous fusion of primary cytotrophoblasts, whereas overexpression of the secreted variant, ADAM12S, potentiates cell fusion in the Bewo trophoblast cell line. Mechanistically, both ectopic and endogenous levels of ADAM12 were shown to control trophoblast fusion through E-cadherin ectodomain shedding and remodeling of intercellular boundaries. This study describes a novel role for ADAM12 in placental development, specifically highlighting its importance in controlling the differentiation of villous cytotrophoblasts into multinucleated cellular structures. Moreover, this work identifies E-cadherin as a novel ADAM12 substrate, and highlights the significance that cell adhesion molecule ectodomain shedding has in normal development.
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a disintegrin and metalloproteinase 12 ADAM12 localizes to invasive trophoblast promotes cell invasion and directs column outgrowth in early placental development
Molecular Human Reproduction, 2014Co-Authors: Mohsen Aghababaei, S Perdu, Alexander G Beristain, K IrvineAbstract:During pregnancy, stromal- and vascular-remodeling trophoblasts serve critical roles in directing placental development acquiring pro-invasive characteristics. The A Disintegrin and Metalloproteinase (ADAM) family of multifunctional proteins direct cellular processes across multiple organ systems via their intrinsic catalytic, cell adhesive and intracellular signaling properties. ADAM12, existing as two distinct splice variants (ADAM12L and ADAM12S), is highly expressed in the human placenta and promotes cell migration and invasion in several tumor cell lines; however, its role in trophoblast biology is unknown. In this study, ADAM12 was localized to anchoring trophoblast columns in first trimester placentas and to highly invasive extracellular matrix-degrading trophoblasts in placental villous explants. The importance of ADAM12 in directing trophoblast invasion was tested using loss-of and gain-of-function strategies, where siRNA-directed knockdown of ADAM12 inhibited trophoblast cell invasion while over-expression promoted migration and invasion in two trophoblastic cell models. In placental villous explant cultures, siRNA-directed loss of ADAM12 significantly dampened trophoblast column outgrowth. Additionally, we provide functional evidence for the ADAM12S variant in promoting trophoblast invasion and column outgrowth through a mechanism requiring its catalytic activity. This is the first study to assign a function for ADAM12 in trophoblast biology, where ADAM12 may play a central role regulating the behavior of invasive trophoblast subsets in early pregnancy. This study also underlines the importance of ADAM12L and ADAM12S in directing cell motility in normal developmental processes outside of cancer, specifically highlighting a potentially important function of ADAM12S in directing early placental development.
