Mesenchymal Transglutaminase 2 Activates Epithelial ADAM17: Link to G-Protein-Coupled Receptor 56 (ADGRG1) Signalling (original) (raw)
PLoS ONE
The increased expression of the Disintegrin and Metalloprotease ADAM12 has been associated with human cancers, however its role remain unclear. We have previously reported that ADAM12 expression is induced by the transforming growth factor, TGF-β and promotes TGF-β-dependent signaling through interaction with the type II receptor of TGF-β. Here we explore the implication of ADAM12 in TGF-β-mediated epithelial to mesenchymal transition (EMT), a key process in cancer progression. We show that ADAM12 expression is correlated with EMT markers in human breast cancer cell lines and biopsies. Using a non-malignant breast epithelial cell line (MCF10A), we demonstrate that TGF-β-induced EMT increases expression of the membrane-anchored ADAM12L long form. Importantly, ADAM12L overexpression in MCF10A is sufficient to induce loss of cell-cell contact, reorganization of actin cytoskeleton, up-regulation of EMT markers and chemoresistance. These effects are independent of the proteolytic activity but require the cytoplasmic tail and are specific of ADAM12L since overexpression of ADAM12S failed to induce similar changes. We further demonstrate that ADAM12L-dependent EMT is associated with increased phosphorylation of Smad3, Akt and ERK proteins. Conversely, inhibition of TGF-β receptors or ERK activities reverses ADAM12L-induced mesenchymal phenotype. Together our data demonstrate that ADAM12L is associated with EMT and contributes to TGF-β-dependent EMT by favoring both Smad-dependent and Smad-independent pathways.
Cellular roles of ADAM12 in health and disease
The International Journal of Biochemistry & Cell Biology, 2008
ADAM12 belongs to the large family of ADAMs (a disintegrin and metalloproteases) and possesses extracellular metalloprotease and cell-binding functions, as well as intracellular signaling capacities. Interest in ADAM12 has increased recently because its expression is related to tumor progression and it is a potential biomarker for breast cancer. It is therefore important to understand ADAM12's functions. Many cellular roles for ADAM12 have been suggested. It is an active metalloprotease, and has been implicated in insulin-like growth factor (IGF) receptor signaling, through cleavage of IGF-binding proteins, and in epidermal growth factor receptor (EGFR) pathways, via ectodomain shedding of membrane-tethered EGFR ligands. These proteolytic events may regulate diverse cellular responses, such as altered cell differentiation, proliferation, migration, and invasion. ADAM12 may also regulate cell-cell and cell-extracellular matrix contacts through interactions with cell surface receptors-integrins and syndecans-potentially influencing the actin cytoskeleton. Moreover, ADAM12 interacts with several cytoplasmic signaling and adaptor molecules through its intracellular domain, thereby directly transmitting signals to or from the cell interior. These ADAM12-mediated cellular effects appear to be critical events in both biological and pathological processes. This review presents current knowledge on ADAM12 functions gained from in vitro and in vivo observations, describes ADAM12's role in both normal physiology and pathology, particularly in cancer, and discusses important areas for future investigation.
ADAM17, shedding, TACE as therapeutic targets
a b s t r a c t ADAM17 has been molecularly cloned as the enzyme responsible for cleavage of the transmembrane protein TNF␣ (TNF␣ converting enzyme, TACE). Later it was realized that ADAM17 was also responsible for the processing of cell adhesion proteins, cytokine and growth factor receptors and many ligands of the EGF receptor. Since TNF␣ is a target of anti-inflammatory therapies, it was speculated that inhibition of ADAM17 might be a therapeutic strategy in the treatment of inflammation or inflammation associated cancer. Meanwhile it has been recognized that ADAM17 governs many vital functions in the body and loss of ADAM17 leads to severe defects in the skin and to high susceptibility of the intestine to inflammation. Here I summarize data on the physiologic role of ADAM17 and the feasibility of specific blockade of this enzyme.
Regulation of ADAM12 Cell-surface Expression by Protein Kinase C ϵ
Journal of Biological Chemistry, 2004
The ADAM (a disintegrin and metalloprotease) family consists of multidomain cell-surface proteins that have a major impact on cell behavior. These transmembraneanchored proteins are synthesized as proforms that have (from the N terminus): a prodomain; a metalloprotease-, disintegrin-like-, cysteine-rich, epidermal growth factor-like, and transmembrane domain; and a cytoplasmic tail. The 90-kDa mature form of human ADAM12 is generated in the trans-Golgi through cleavage of the prodomain by a furin-peptidase and is stored intracellularly until translocation to the cell surface as a constitutively active protein. However, little is known about the regulation of ADAM12 cell-surface translocation. Here, we used human RD rhabdomyosarcoma cells, which express ADAM12 at the cell surface, in a temporal pattern. We report that protein kinase C (PKC) ⑀ induces ADAM12 translocation to the cell surface and that catalytic activity of PKC⑀ is required for this translocation. The following results support this conclusion: 1) treatment of cells with 0.1 M phorbol 12-myristate 13-acetate (PMA) enhanced ADAM12 cell-surface immunostaining, 2) ADAM12 and PKC⑀ could be co-immunoprecipitated from membrane-enriched fractions of PMA-treated cells, 3) RD cells transfected with EGFP-tagged, myristoylated PKC⑀ expressed more ADAM12 at the cell surface than did non-transfected cells, and 4) RD cells transfected with a kinase-inactive PKC⑀ mutant did not exhibit ADAM12 cell-surface translocation upon PMA treatment. Finally, we demonstrate that the C1 and C2 domains of PKC⑀ both contain a binding site for ADAM12. These studies show that PKC⑀ plays a critical role in the regulation of ADAM12 cell-surface expression. Cells possess a diverse array of surface proteins, lipids, and carbohydrates that provide active gateways for the selective intake and release of molecular information, which is important in regulating cell behavior. In fact, many disease processes relate to disorganized cell-surface communication systems. ADAMs 1 belong to a large family of cell-surface proteins with over 30 members. The prototypical ADAM molecule is a transmembrane glycoprotein composed of several distinct domains, including a prodomain and a metalloprotease, disintegrin-like, cysteine-rich, epidermal growth factor-like, transmembrane, and cytoplasmic domain. ADAMs play important roles in cell adhesion, interacting with integrins and syndecans, and in the proteolysis of the ectodomains of cell-surface proteins, such as growth factors, growth factor receptors, and cytokines (1-5). For example, ADAM17 (TACE) mediates release of tumor necrosis factor-␣, transforming growth factor-␣, -amyloid, L-selectin, TRANCE, and amphiregulin precursor proteins (6-11). ADAMs 9, 10, and 12 have been shown to cleave membraneanchored, heparin-binding epidermal growth factor (12-15). Important in vivo functions have been reported for several ADAMs (3). For example, the finding that ADAM9,-10, and-17 have, or mediate, ␣-secretase activity could be used to design new treatment strategies for Alzheimer's disease (7, 14, 16, 17). Overexpression of ADAMs has been observed in many human cancers (18, 19), suggesting that ADAMs could promote tumor growth and metastasis by modulating growth factor shedding and cell adhesion. A recent genome-wide scan and polymorphism analysis of a large group of patients identified ADAM33 on chromosome 20 as a putative asthma susceptibility gene (20). We have demonstrated that ADAM12-S, which is present in the serum of pregnant women but not in that of women who are not pregnant (21), can be used as a first-trimester maternal serum marker for Down syndrome (22). Gene-ablation experiments in mice revealed that ADAM17 (TACE)-deficient mice have severe perinatal and postnatal defects primarily related to eye, hair, and skin anomalies, including failure of eyelid fusion (10). In contrast, ADAM9-deficient mice have an apparently normal phenotype (10, 14). ADAM12 deficiency confers increased perinatal mortality, although the reason for this is not yet well understood (23). Surviving ADAM12-null mice have defects in adipose tissue (23), and mice overexpressing ADAM12 under the muscle creatine kinase promoter exhibit increased adipogenesis (24), supporting the idea that ADAM12 is involved in mesenchymal cell differentiation. ADAM12, originally named meltrin-␣ (25), has been implicated in muscle cell function in vivo and in vitro (25-30). In the original study, expression of a truncated version of
iRHOM2-dependent regulation of ADAM17 in cutaneous disease and epidermal barrier function.
iRHOM2 is a highly conserved, catalytically inactive member of the Rhomboid family, which has recently been shown to regulate the maturation of the multi-substrate ectodomain sheddase enzyme ADAM17 (TACE) in macrophages. Dominant iRHOM2 mutations are the cause of the inherited cutaneous and oesophageal cancer-susceptibility syndrome tylosis with oesophageal cancer (TOC), suggesting a role for this protein in epithelial cells. Here, using tissues derived from TOC patients, we demonstrate that TOC-associated mutations in iRHOM2 cause an increase in the maturation and activity of ADAM17 in epidermal keratinocytes, resulting in significantly upregulated shedding of ADAM17 substrates, including EGF-family growth factors and pro-inflammatory cytokines. This activity is accompanied by increased EGFR activity, increased desmosome processing and the presence of immature epidermal desmosomes, upregulated epidermal transglutaminase activity and heightened resistance to Staphylococcal infection in TOC keratinocytes. Many of these features are consistent with the presence of a constitutive wound-healing-like phenotype in TOC epidermis, which may shed light on a novel pathway in skin repair, regeneration and inflammation.
Scientific Reports, 2016
The vitally important skin barrier is formed by extensive cross-linking activity of transglutaminases (TGs) during terminal epidermal differentiation. We have previously shown that epidermal deficiency of a disintegrin and metalloproteinase 17 (ADAM17), the principal EGFR ligand sheddase, results in postnatal skin barrier defects in mice due to impeded TG activity. However, the mechanism by which ADAM17/EGFR signalling maintains TG activity during epidermal differentiation remains elusive. Here we demonstrate that ADAM17-dependent EGFR signalling promotes TG activity in keratinocytes committed to terminal differentiation by direct induction of TG1 expression. Restored TG1 expression of EGF-stimulated differentiated Adam17 −/− keratinocytes was strongly repressed by inhibitors for PLCγ1 or protein kinase C (PKC) pathways, while treatment with the PKC stimulator 12-O-tetradecanoylphorbol-13-acetate restored TG activity in the epidermis of keratinocyte-specific Adam17 −/− (AD17 ΔKC ) m...
Characterization and regulation of ADAMTS-16
Matrix Biology, 2009
The ADAMTS (a disintegrin and metalloproteinase domain with thrombospondin motifs) family includes 19 secreted proteinases in man. ADAMTS16 is a recently cloned gene expressed at high levels in fetal lung and kidney and adult brain and ovary. The ADAMTS-16 protein currently has no known function. ADAMTS16 is also expressed in human cartilage and synovium where its expression is increased in tissues from osteoarthritis patients compared to normal tissues. In this study, we ascertained that the full length ADAMTS16 mRNA was expressed in chondrocytes and cloned the appropriate cDNA. Stable over-expression of ADAMTS16 in chondrosarcoma cells led to a decrease in cell proliferation and migration, though not adhesion, as well as a decrease in the expression of matrix metalloproteinase-13 (MMP13). The transcription start point of the human ADAMTS16 gene was experimentally identified as 138 bp upstream of the translation start ATG and the basal promoter was mapped out to − 1802 bp. Overexpression of Egr1 induced ADAMTS16 promoter constructs of − 157/+138 or longer whilst Sp1 induced all ADAMTS16 promoter constructs. Transforming growth factor beta (TGFβ) stimulated expression of endogenous ADAMTS16 gene expression in chondrocyte cell lines.
Scientific Reports, 2016
The vitally important skin barrier is formed by extensive cross-linking activity of transglutaminases (TGs) during terminal epidermal differentiation. We have previously shown that epidermal deficiency of a disintegrin and metalloproteinase 17 (ADAM17), the principal EGFR ligand sheddase, results in postnatal skin barrier defects in mice due to impeded TG activity. However, the mechanism by which ADAM17/EGFR signalling maintains TG activity during epidermal differentiation remains elusive. Here we demonstrate that ADAM17-dependent EGFR signalling promotes TG activity in keratinocytes committed to terminal differentiation by direct induction of TG1 expression. Restored TG1 expression of EGF-stimulated differentiated Adam17−/− keratinocytes was strongly repressed by inhibitors for PLCγ1 or protein kinase C (PKC) pathways, while treatment with the PKC stimulator 12-O-tetradecanoylphorbol-13-acetate restored TG activity in the epidermis of keratinocyte-specific Adam17−/− (AD17ΔKC) mice....
ADAM-15 inhibits wound healing in human intestinal epithelial cell monolayers
AJP: Gastrointestinal and Liver Physiology, 2005
The disintegrin metalloproteases (or ADAMs) are membrane-anchored glycoproteins that have been implicated in cell-cell or cell-matrix interactions and in proteolysis of molecules on the cell surface. The expression and/or the pathophysiological implications of ADAMs are not known in intestinal epithelial cells. Therefore our aim was to investigate the expression and the role of ADAMs in intestinal epithelial cells.
Deciphering the Role of the ADAM17-Dependent Secretome in Cell Signaling
Journal of Proteome Research, 2014
ADAM17 has been initially identified as the main sheddase responsible for releasing the soluble form of a variety of cell-surface proteins, including growth factors, cytokines, cell adhesion molecules, and receptors, most of which are associated with pathological processes, including cancer and inflammation. However, the function and composition of the ADAM17dependent secretome on a proteome-wide scale is poorly understood. In this study, we observed that the ADAM17-dependent secretome plays an important role in promoting cell proliferation and migration. To further demonstrate the repertoire of proteins involved in this cross-talk, we employed mass-spectrometrybased proteomics using nonmetabolic and metabolic labeling approaches to explore the secretome composition of wild-type and ADAM17 −/− knockout mouse embryonic fibroblast (mEF) cells. Bioinformatic analyses indicated the differential regulation of 277 soluble proteins in the ADAM17-dependent secretome as well as novel direct ADAM17 cleavage substrates, such as mimecan and perlecan. Furthermore, we found that the ADAM17-dependent secretome promoted an opposite regulation of ERK and FAK pathways as well as PPARγ downstream activation. These findings demonstrated fine-tuning of cell signaling rendered by the soluble molecules mediated by ADAM17.
FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 2016
A disintegrin and metalloproteinase with thrombospondin type I motif (ADAMTS)2, 3, and 14 are collectively named procollagen N-proteinases (pNPs) because of their specific ability to cleave the aminopropeptide of fibrillar procollagens. Several reports also indicate that they could be involved in other biological processes, such as blood coagulation, development, and male fertility, but the potential substrates associated with these activities remain unknown. Using the recently described N-terminal amine isotopic labeling of substrate approach, we analyzed the secretomes of human fibroblasts and identified 8, 17, and 22 candidate substrates for ADAMTS2, 3, and 14, respectively. Among these newly identified substrates, many are components of the extracellular matrix and/or proteins related to cell signaling such as latent TGF-β binding protein 1, TGF-β RIII, and dickkopf-related protein 3. Candidate substrates for the 3 ADAMTS have been biochemically validated in different contexts, ...
Respiratory …, 2006
The molecular mechanisms of Idiopathic Pulmonary Fibrosis (IPF) remain elusive. Transforming Growth Factor beta 1(TGF-β1) is a key effector cytokine in the development of lung fibrosis. We used microarray and computational biology strategies to identify genes whose expression is significantly altered in alveolar epithelial cells (A549) in response to TGF-β1, IL-4 and IL-13 and Epstein Barr virus. A549 cells were exposed to 10 ng/ml TGF-β1, IL-4 and IL-13 at serial time points. Total RNA was used for hybridisation to Affymetrix Human Genome U133A microarrays. Each in vitro time-point was studied in duplicate and an average RMA value computed. Expression data for each time point was compared to control and a signal log ratio of 0.6 or greater taken to identify significant differential regulation. Using normalised RMA values and unsupervised Average Linkage Hierarchical Cluster Analysis, a list of 312 extracellular matrix (ECM) proteins or modulators of matrix turnover was curated via Onto-Compare and Gene-Ontology (GO) databases for baited cluster analysis of ECM associated genes. Interrogation of the dataset using ontological classification focused cluster analysis revealed coordinate differential expression of a large cohort of extracellular matrix associated genes. Of this grouping members of the ADAM (A disintegrin and Metalloproteinase domain containing) family of genes were differentially expressed. ADAM gene expression was also identified in EBV infected A549 cells as well as IL-13 and IL-4 stimulated cells. We probed pathologenomic activities (activation and functional activity) of ADAM19 and ADAMTS9 using siRNA and collagen assays. Knockdown of these genes resulted in diminished production of collagen in A549 cells exposed to TGF-β1, suggesting a potential role for these molecules in ECM accumulation in IPF.
The metalloprotease ADAM17 catalyzes the shedding of key signalling molecules from the cell surface, including the inflammatory cytokine TNF (tumour necrosis factor) and activating ligands of the EGFR (epidermal growth factor receptor). ADAM17 exists within an assemblage called the “sheddase complex” containing a rhomboid pseudoprotease (iRhom1 or iRhom2). iRhoms control multiple aspects of ADAM17 biology, including its vesicular trafficking, maturation from its precursor pro-form, activation on the cell surface and specificity for subsets of proteolytic targets. Previous studies from our laboratory and others identified the FERM domain-containing protein Frmd8/iTAP as an iRhom-binding protein. iTAP is required to maintain the cell surface stability of the sheddase complex, thereby preventing the precocious shunting of ADAM17 and iRhom2 to lysosomes and their consequent degradation. As pathophysiological role(s) of iTAP have not been addressed, here we sought to characterize the imp...
Journal of Biological Chemistry, 2004
Osteoarthritis is characterized by the loss of aggrecan and collagen from the cartilage extracellular matrix. The proteinases responsible for the breakdown of cartilage aggrecan include ADAMTS-4 (aggrecanase 1) and ADAMTS-5 (aggrecanase 2). Post-translational inhibition of ADAMTS-4/-5 activity may be important for maintaining normal homeostasis of aggrecan metabolism, and thus, any disruption to this inhibition could lead to accelerated aggrecan breakdown. To date TIMP-3 (tissue inhibitor of matrix metalloproteinases-3) is the only endogenous inhibitor of ADAMTS-4/-5 that has been identified. In the present studies we identify ␣ 2-macroglobulin (␣ 2 M) as an additional endogenous inhibitor of ADAMTS-4 and ADAMTS-5. ␣ 2 M inhibited the activity of both ADAMTS-4 and ADAMTS-5 in a concentration-dependent manner, demonstrating 1:1 stoichiometry with second-order rate constants on the order of 10 6 and 10 5 M ؊1 s ؊1 , respectively. Inhibition of the aggrecanases was mediated by proteolysis of the bait region within ␣ 2 M, resulting in physical entrapment of these proteinases. Both ADAMTS-4 and ADAMTS-5 cleaved ␣ 2 M at Met 690 / Gly 691 , representing a novel proteinase cleavage site within ␣ 2 M and a novel site of cleavage for ADAMTS-4 and ADAMTS-5. Finally, the use of the anti-neoepitope antibodies to detect aggrecanase-generated ␣ 2 M-fragments in synovial fluid was investigated and found to be uninformative.
2007
Abbreviations used in this paper: ADAM, a disintegrin and metalloproteinase; EEA1, early endosomal antigen 1; HSC, hepatic stellate cell; PAI-1, plasmino-gen activator inhibitor-1; RD, Rhabdomyosarcoma; SARA, Smad anchor for receptor activation; shRNA, short hairpin RNA; TβRII, TGF-β type II receptor. The online version of this article contains supplemental material. on 202 ADAM12 REGULATES TGF-β RECEPTOR TRAFFICKING • ATFI ET AL. 207 independent colonies were screened as previously described (Colland et al., 2004). The prey fragments of the positive clones were PCR ampli-fi ed and sequenced. Cell culture and transfection The human embryonic kidney cell line 293T, HSCs, human RD cells, mouse C2C12 cells, monkey kidney COS7 cells, and mink lung MvLu1 cells were transfected using LipofectAMINE-Plus reagent (Invitrogen) according to the manufacturer's instructions. For experiments with ADAM12 antisense, cells were incubated with 2 μM of antisense oligonucleotides to ADAM12
2006
The molecular mechanisms of Idiopathic Pulmonary Fibrosis (IPF) remain elusive. Transforming Growth Factor beta 1(TGF-β1) is a key effector cytokine in the development of lung fibrosis. We used microarray and computational biology strategies to identify genes whose expression is significantly altered in alveolar epithelial cells (A549) in response to TGF-β1, IL-4 and IL-13 and Epstein Barr virus. A549 cells were exposed to 10 ng/ml TGF-β1, IL-4 and IL-13 at serial time points. Total RNA was used for hybridisation to Affymetrix Human Genome U133A microarrays. Each in vitro time-point was studied in duplicate and an average RMA value computed. Expression data for each time point was compared to control and a signal log ratio of 0.6 or greater taken to identify significant differential regulation. Using normalised RMA values and unsupervised Average Linkage Hierarchical Cluster Analysis, a list of 312 extracellular matrix (ECM) proteins or modulators of matrix turnover was curated via Onto-Compare and Gene-Ontology (GO) databases for baited cluster analysis of ECM associated genes. Interrogation of the dataset using ontological classification focused cluster analysis revealed coordinate differential expression of a large cohort of extracellular matrix associated genes. Of this grouping members of the ADAM (A disintegrin and Metalloproteinase domain containing) family of genes were differentially expressed. ADAM gene expression was also identified in EBV infected A549 cells as well as IL-13 and IL-4 stimulated cells. We probed pathologenomic activities (activation and functional activity) of ADAM19 and ADAMTS9 using siRNA and collagen assays. Knockdown of these genes resulted in diminished production of collagen in A549 cells exposed to TGF-β1, suggesting a potential role for these molecules in ECM accumulation in IPF.
Journal of Experimental Medicine, 2010
The protease a disintegrin and metalloprotease (ADAM) 17 cleaves tumor necrosis factor (TNF), L-selectin, and epidermal growth factor receptor (EGF-R) ligands from the plasma membrane. ADAM17 is expressed in most tissues and is up-regulated during inflammation and cancer. ADAM17-deficient mice are not viable. Conditional ADAM17 knockout models demonstrated proinflammatory activities of ADAM17 in septic shock via shedding of TNF. We used a novel gene targeting strategy to generate mice with dramatically reduced ADAM17 levels in all tissues. The resulting mice called ADAM17 ex/ex were viable, showed compromised shedding of ADAM17 substrates from the cell surface, and developed eye, heart, and skin defects as a consequence of impaired EGF-R signaling caused by failure of shedding of EGF-R ligands. Unexpectedly, although the intestine of unchallenged homozygous ADAM17 ex/ex mice was normal, ADAM17 ex/ex mice showed substantially increased susceptibility to inflammation in dextran sulfate sodium colitis. This was a result of impaired shedding of EGF-R ligands resulting in failure to phosphorylate STAT3 via the EGF-R and, consequently, in defective regeneration of epithelial cells and breakdown of the intestinal barrier. Besides regulating the systemic availability of the proinflammatory cytokine TNF, our results demonstrate that ADAM17 is needed for vital regenerative activities during the immune response. Thus, our mouse model will help investigate ADAM17 as a potential drug target.
Differential Induction of the ADAM17 Regulators iRhom1 and 2 in Endothelial Cells
Frontiers in Cardiovascular Medicine, 2020
Background: Endothelial function significantly depends on the proteolytic release of surface expressed signal molecules, their receptors and adhesion molecules via the metalloproteinase ADAM17. The pseudoproteases iRhom1 and 2 independently function as adapter proteins for ADAM17 and are essential for the maturation, trafficking, and activity regulation of ADAM17. Bioinformatic data confirmed that immune cells predominantly express iRhom2 while endothelial cells preferentially express iRhom1. Objective: Here, we investigate possible reasons for higher iRhom1 expression and potential inflammatory regulation of iRhom2 in endothelial cells and analyze the consequences for ADAM17 maturation and function. Methods: Primary endothelial cells were cultured in absence and presence of flow with and without inflammatory cytokines (TNFα and INFγ). Regulation of iRhoms was studied by qPCR, involved signaling pathways were studied with transcriptional inhibitors and consequences were analyzed by assessment of ADAM17 maturation, surface expression and cleavage of the ADAM17 substrate junctional adhesion molecule JAM-A. Results: Endothelial iRhom1 is profoundly upregulated by physiological shear stress. This is accompanied by a homeostatic phenotype driven by the transcription factor KLF2 which is, however, only partially responsible for regulation of iRhom1. By contrast, iRhom2 is most prominently upregulated by inflammatory cytokines. This correlates with an inflammatory phenotype driven by the transcription factors NFκB and AP-1 of which AP-1 is most relevant for iRhom2 regulation. Finally, shear stress exposure and inflammatory stimulation have independent and no synergistic effects on ADAM17 maturation, surface expression and JAM-A shedding. Conditions of shear stress and inflammation differentially upregulate iRhom1 and 2 in primary endothelial cells which then results in independent regulation of ADAM17.
ADAM17 mediates OSCC development in an orthotopic murine model
Molecular Cancer, 2014
Background: ADAM17 is one of the main sheddases of the cells and it is responsible for the cleavage and the release of ectodomains of important signaling molecules, such as EGFR ligands. Despite the known crosstalk between ADAM17 and EGFR, which has been considered a promising targeted therapy in oral squamous cell carcinoma (OSCC), the role of ADAM17 in OSCC development is not clear. Method: In this study the effect of overexpressing ADAM17 in cell migration, viability, adhesion and proliferation was comprehensively appraised in vitro. In addition, the tumor size, tumor proliferative activity, tumor collagenase activity and MS-based proteomics of tumor tissues have been evaluated by injecting tumorigenic squamous carcinoma cells (SCC-9) overexpressing ADAM17 in immunodeficient mice.
Regulation of ADAM12 Cell-surface Expression by Protein Kinase C
Journal of Biological Chemistry, 2004
The ADAM (a disintegrin and metalloprotease) family consists of multidomain cell-surface proteins that have a major impact on cell behavior. These transmembraneanchored proteins are synthesized as proforms that have (from the N terminus): a prodomain; a metalloprotease-, disintegrin-like-, cysteine-rich, epidermal growth factor-like, and transmembrane domain; and a cytoplasmic tail. The 90-kDa mature form of human ADAM12 is generated in the trans-Golgi through cleavage of the prodomain by a furin-peptidase and is stored intracellularly until translocation to the cell surface as a constitutively active protein. However, little is known about the regulation of ADAM12 cell-surface translocation. Here, we used human RD rhabdomyosarcoma cells, which express ADAM12 at the cell surface, in a temporal pattern. We report that protein kinase C (PKC) ⑀ induces ADAM12 translocation to the cell surface and that catalytic activity of PKC⑀ is required for this translocation. The following results support this conclusion: 1) treatment of cells with 0.1 M phorbol 12-myristate 13-acetate (PMA) enhanced ADAM12 cell-surface immunostaining, 2) ADAM12 and PKC⑀ could be co-immunoprecipitated from membrane-enriched fractions of PMA-treated cells, 3) RD cells transfected with EGFP-tagged, myristoylated PKC⑀ expressed more ADAM12 at the cell surface than did non-transfected cells, and 4) RD cells transfected with a kinase-inactive PKC⑀ mutant did not exhibit ADAM12 cell-surface translocation upon PMA treatment. Finally, we demonstrate that the C1 and C2 domains of PKC⑀ both contain a binding site for ADAM12. These studies show that PKC⑀ plays a critical role in the regulation of ADAM12 cell-surface expression.