SEARCH · PubMed Health

Results for “Integrin”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,495 records · Page 83Linked to original sources

PubMed2005

VEGF-integrin interplay controls tumor growth and vascularization.

Cross-talk between the major angiogenic growth factor, VEGF, and integrin cell adhesion receptors has emerged recently as a critical factor in the regulation of angiogenesis and tumor development. However, the molecular mechanisms and consequences of this intercommunication remain unclear. Here, we define a mechanism whereby integrin alpha v beta3, through activation, clustering, and signaling by means of p66 Shc (Src homology 2 domain containing), regulates the production of VEGF in tumor cells expressing this integrin. Tumors with "activatable" but not "inactive" beta3 integrin secrete high levels of VEGF, which in turn promotes extensive neovascularization and augments tumor growth in vivo. This stimulation of VEGF expression depends upon the ability of alpha v beta3 integrin to cluster and promote phosphorylation of p66 Shc. These observations identify a link between beta3 integrins and VEGF in tumor growth and angiogenesis and, therefore, may influence anti-integrin as well as anti-VEGF therapeutic strategies.

Adaptor Proteins, Signal Transducing↗

PubMed1995

Integrin-mediated tyrosine phosphorylation and cytokine message induction in monocytic cells. A possible signaling role for the Syk tyrosine kinase.

Activation of cytoplasmic tyrosine kinases is an important aspect of signal transduction mediated by integrins. In the human monocytic cell line THP-1, either integrin-dependent cell adhesion to fibronectin or ligation of beta 1 integrins with antibodies causes a rapid and intense tyrosine phosphorylation of two sets of proteins of about 65-75 and 120-125 kDa. In addition, integrin ligation leads to nuclear translocation of the p50 and p65 subunits of the NF-kappa B transcription factor, to activation of a reporter gene driven by a promoter containing NF-kappa B sites, and to increased levels of mRNAs for immediate-early genes, including the cytokine interleukin (IL)-1 beta. The tyrosine kinase inhibitors genistein and herbimycin A block both integrin-mediated tyrosine phosphorylation and increases in IL-1 beta message levels, indicating a causal relationship between the two events. The components tyrosine phosphorylated subsequent to cell adhesion include paxillin, pp125FAK, and the SH2 domain containing tyrosine kinase Syk. In contrast, integrin ligation with antibodies induces tyrosine phosphorylation of Syk but not of FAK or paxillin. In adhering cells, pre-treatment with cytochalasin D suppresses tyrosine phosphorylation of FAK and paxillin but not of Syk, while IL-1 beta message induction is unaffected. These observations indicate that the Syk tyrosine kinase may be an important component of an integrin signaling pathway in monocytic cells, leading to activation of NF-kappa B and to increased levels of cytokine messages.

Benzoquinones↗

PubMed1998

Identification of pactolus, an integrin beta subunit-like cell-surface protein preferentially expressed by cells of the bone marrow.

We have sought to develop methodologies to identify genes that are preferentially expressed during the differentiation of mast cells from their hematopoietic stem cell precursors. By using a modified differential display protocol, we compared a subset of transcripts expressed in bone marrow cells differentiated into immature mast cells with the exogenous addition of stem cell factor (SCF) or interleukin 3. One gene was identified that was preferentially expressed in the SCF-derived cells and encodes a novel murine integrin beta subunit-like molecule, dubbed Pactolus-1 (Pactolus). Two distinct forms of Pactolus mRNA were detected which, via alternative splicing, are predicted to encode a membrane-bound form and truncated version of the protein. The full-length Pactolus gene product is very similar to a number of beta subunit integrin chains, particularly beta2, with the notable exceptions of the apparent deletion of the metal-binding site within the putative metal ion-dependent adhesion site-like domain of the Pactolus gene product and a cytoplasmic domain that shares no obvious homology to similar domains of the other beta subunit integrin proteins. Although the Pactolus sequence was first identified in immature mast cell samples, screening of murine tissues indicated the highest level of Pactolus expression was found in the bone marrow, suggesting that the expression of Pactolus is confined to immature and maturing bone marrow-derived cells, and that the SCF-derived mast cells are more representative of this state than are the interleukin 3-derived mast cells. Immunoprecipitation of Pactolus revealed a cell-surface protein with an apparent molecular mass of about 95 kDa. Surprisingly, no associating alpha integrin subunit could be identified suggesting that either Pactolus does not associate with another integrin subunit or the association is too weak to be identified. These data suggest that Pactolus represents a gene and gene product related to those of the integrin beta subunits but whose function(s) may be quite distinct from those of the integrin beta subunits.

Alternative Splicing↗

PubMed1998

The human WD repeat protein WAIT-1 specifically interacts with the cytoplasmic tails of beta7-integrins.

Integrins of the beta7 subfamily, alpha4 beta7 and alphaE beta7, contribute to lymphocyte homing and to the development of protective or autoreactive immune responses at mucosal sites. The beta subunits of integrins are considered important for regulation of stimulated cell adhesion and adhesion-dependent signal transduction. Using a yeast interaction trap screen, a human WD repeat protein, termed WAIT-1, was isolated that interacts with the integrin beta7 cytoplasmic tail and is homologous to mouse EED and Drosophila ESC proteins. WAIT-1 also binds to the cytoplasmic domains of alpha4 and alphaE but not to those of integrin beta1, beta2, and alphaL subunits. Association of WAIT-1 and beta7-integrin was confirmed by coprecipitation from transiently transfected 293 cells. The binding site for WAIT-1 was mapped to a short membrane-proximal region of the beta7 cytoplasmic tail with Tyr-735 being of critical importance. Northern blot analysis revealed multiple WAIT-1-related transcripts with differential expression in circulating leukocytes, tissue-resident cells of diverse origin, and lymphoid malignancies. These results suggest that WAIT-1, together with the recently identified RACK1, may define a novel subfamily of WD repeat proteins that interact with distinct subsets of integrin cytoplasmic tails and may act as specific regulators of integrin function.

Amino Acid Sequence↗

PubMed1999

Activation of p53 function in carcinoma cells by the alpha6beta4 integrin.

The interaction of integrins with extracellular matrix is known to promote cell survival by inhibiting apoptotic signaling. In contrast, we demonstrate here that the alpha6beta4 integrin induces apoptosis in carcinoma cells by stimulating p53 function. Specifically, we show that expression of alpha6beta4 in carcinoma cells that lack this integrin stimulates an increase in the transactivating function of p53 as demonstrated by the ability of this integrin to up-regulate the expression of a p53-sensitive reporter gene as well as the endogenous p53 response gene, bax. In addition, we report that alpha6beta4 triggers apoptosis in carcinoma cells that express wild-type but not mutant p53 and that these alpha6beta4 functions are inhibited by a dominant negative p53 construct. Importantly, we provide a link between integrin signaling and p53 activation by demonstrating that the clustering of alpha6beta4 with a beta4 integrin-specific antibody promotes p53-dependent apoptosis in cells that express both alpha6beta4 and wild-type p53. These studies are the first to demonstrate that a specific integrin can promote apoptosis by activating p53. Moreover, given the ability of alpha6beta4 to stimulate invasion (Shaw, L. M., Rabinovitz, I., Wang, H. F., Toker, A., and Mercurio, A. M. (1997) Cell 91, 949-960), these studies suggest that the ability of alpha6beta4 to promote carcinoma progression will be enhanced in tumor cells that express mutant, inactive forms of p53.

Antigens, Surface↗

PubMed2004

A novel adaptation of the integrin PSI domain revealed from its crystal structure.

Integrin beta-subunits contain an N-terminal PSI (for plexin-semaphorin-integrin) domain that contributes to integrin activation and harbors the PI(A) alloantigen associated with immune thrombocytopenias and susceptibility to sudden cardiac death. Here we report the crystal structure of PSI in the context of the crystallized alphaVbeta3 ectodomain. The integrin PSI forms a two-stranded antiparallel beta-sheet flanked by two short helices; its long interstrand loop houses Pl(A) and may face the EGF2 domain. The integrin PSI contains four cysteine pairs connected in a 1-4, 2-8, 3-6, 5-7 pattern. An unexpected feature of the structure is that the final, eighth cysteine is located C-terminal to the Ig-like hybrid domain and is thus separated by the hybrid domain from the other seven cysteines of PSI. This architecture may be relevant to the evolution of integrins and should help refine the current models of integrin activation.

Amino Acid Sequence↗

PubMed2001

An alternative processing of integrin alpha(v) subunit in tumor cells by membrane type-1 matrix metalloproteinase.

Membrane type-1 matrix metalloproteinase (MT1-MMP) and alpha(v)beta(3) integrin are both essential to cell invasion. Maturation of integrin pro-alpha(v)chain (pro-alpha(v)) involves its cleavage by proprotein convertases (PC) to form the disulfide-bonded 125-kDa heavy and 25-kDa light alpha chains. Our report presents evidence of an alternative pathway of pro-alpha(v) processing involving MT1-MMP. In breast carcinoma MCF7 cells deficient in MT1-MMP, pro-alpha(v) is processed by a conventional furin-like PC, and the mature alpha(v) integrin subunit is represented by the 125-kDa heavy chain and the 25-kDa light chain commencing from the N-terminal Asp(891). In contrast, in cells co-expressing alpha(v)beta(3) and MT1-MMP, MT1-MMP functions as an integrin convertase. MT1-MMP specifically cleaves pro-alpha(v), generating a 115-kDa heavy chain with the truncated C terminus and a 25-kDa light chain commencing from the N-terminal Leu(892). PC-cleavable alpha(3) and alpha(5) but not the PC-resistant alpha(2) integrin subunit are also susceptible to MT1-MMP cleavage. These novel mechanisms involved in the processing of integrin alpha subunits underscore the significance and complexity of interactions between MT1-MMP and adhesion receptors and suggest that regulation of integrin functionality may be an important role of MT1-MMP in migrating tumor cells.

Amino Acid Sequence↗

PubMed2002

Fas activation induces renal tubular epithelial cell beta 8 integrin expression and function in the absence of apoptosis.

Cell fate following Fas (CD95) ligand or agonistic anti-Fas antibody stimulation is determined by multiple factors, including Fas expression level, microdomain localization, and modulating cytokines. Highly expressed Fas clusters and activates a canonical apoptosis signaling pathway. In less susceptible cells, Fas transduces apoptosis-independent signals, which are not well defined, but have been linked to inflammation, angiogenesis, and fibrosis. To identify apoptosis-independent Fas pathways, cultured renal tubular epithelial cells were stimulated with agonistic anti-Fas antibodies under conditions that did not cause cell death. Analysis of filter cDNA microarrays revealed beta(8) integrin subunit mRNA induction in Fas-stimulated cells. beta(8) integrin mRNA expression increased within 3-6 h of Fas ligation due to enhanced mRNA stabilization, and mRNA increases were sustained for 48-72 h. Expression of plasma membrane beta(8) integrin, as well as its heterodimer partner alpha(v), was increased by Fas activation with a similar kinetic pattern. Fas-induced alpha(v)beta(8) expression correlated with increased migration to vitronectin, the ligand for alpha(v)beta(8). Results from studies with function-blocking antibodies against other alpha(v)beta integrins or suppression of beta(8) integrin expression by RNA interference demonstrated that induced beta(8) integrin expression mediated Fas-stimulated migration. We conclude that alpha(v)beta(8) integrin induction defines an unexpected role for Fas in cell migration, rather than as a cell death receptor.

Apoptosis↗

PubMed2002

A functionally relevant conformational epitope on the CD9 tetraspanin depends on the association with activated beta1 integrin.

Tetraspanins associate on the cell membrane with several transmembrane proteins, including members of the integrin superfamily. The tetraspanin CD9 has been implicated in cell motility, metastasis, and sperm-egg fusion. In this study we characterize the first CD9 conformation-dependent epitope (detected by monoclonal antibody (mAb) PAINS-13) whose expression depends on changes in the activation state of associated beta(1) integrins. MAb PAINS-13 precipitates CD9 under conditions that preserve the association of this tetraspanin with integrins, but not under conditions that disrupt these interactions. Induction of activation of beta(1) integrins by temperature, divalent cation Mn(2+), or mAb TS2/16 correlated with enhanced expression of the PAINS-13 epitope on a variety of cells. Through the use of different K562 myeloid leukemia transfectant cells expressing specific members of the beta(1) integrin subfamily we show that the expression of the PAINS-13 epitope depends on CD9 association with alpha(6)beta(1) integrin. The mAb PAINS-13 reactivity has been mapped to the CD9 region comprising residues 112-154 in the NH(2) half of the large extracellular loop. Also, we show that the CD9 conformation recognized by mAb PAINS-13 is functionally relevant in beta(1) integrin-mediated cellular processes including wound healing migration, tubular morphogenesis, cell adhesion and spreading and in signal transduction involving phosphatidylinositol 3-kinase activation.

Animals↗

PubMed2003

Identification of a novel integrin alpha 6 beta 1 binding site in the angiogenic inducer CCN1 (CYR61).

The angiogenic inducer CCN1 (cysteine-rich 61, CYR61), a secreted matricellular protein of the CCN family, is a ligand of multiple integrins, including alpha 6 beta 1. Previous studies have shown that CCN1 interaction with integrin alpha 6 beta 1 mediates adhesion of fibroblasts, endothelial cells, and smooth muscle cells, as well as migration of smooth muscle cells. Recently, we have reported that CCN1-induced tubule formation of unactivated endothelial cells is also mediated through integrin alpha 6 beta 1. In this study, we demonstrate that human skin fibroblasts adhere specifically to the T1 sequence (GQKCIVQTTSWSQCSKS) within domain III of CCN1, and this process is blocked by anti-alpha 6 and anti-beta 1 monoclonal antibodies. Alanine substitution mutagenesis of the T1 sequence further defines the sequence TTSWSQCSKS as the critical determinant for mediating alpha 6 beta 1-dependent adhesion. Soluble T1 peptide specifically inhibits fibroblast adhesion to CCN1 in a dose-dependent manner. Furthermore, T1 also inhibits cell adhesion to other alpha 6 beta 1 ligands, including CCN2 (CTGF), CCN3 (NOV), and laminin, but not to ligands of other integrins. In addition, T1 specifically inhibits alpha 6 beta 1-dependent tubule formation of unactivated endothelial cells in a CCN1-containing collagen gel matrix. To confirm that T1 binds integrin alpha 6 beta 1 directly, we perform affinity chromatography and show that integrin alpha 6 beta 1 is isolated from an octylglucoside extract of fibroblasts on T1-coupled Affi-gel. Taken together, these findings define the T1 sequence in CCN1 as a novel binding motif for integrin alpha 6 beta 1, providing the basis for the development of peptide mimetics to examine the functional role of alpha 6 beta 1 in angiogenesis.

Alanine↗

PubMed2004

Alphav-integrin utilization in human beta-cell adhesion, spreading, and motility.

The role of individual integrins in human beta-cell development and function is largely unknown. This study describes the contribution of alpha(v)-integrins to human beta-cell adhesion, spreading, and motility. Developmental differences in alpha(v)-integrin utilization are addressed by comparing the responses of adult and fetal beta-cells, and vitronectin is used as a substrate based on its unique pattern of expression in the developing pancreas. Fetal and adult beta-cells attached equally to vitronectin and integrin alpha(v)beta(5) was found to support the adhesion of both mature and immature beta-cell populations. Fetal beta-cells were also observed to spread and migrate on vitronectin, and integrin alpha(v)beta(1) was found to be essential for these responses. In contrast to their fetal counterparts, adult beta-cells failed to either spread or migrate and this deficit was associated with a marked down-regulation of alpha(v)beta(1) expression in adult islet preparations. The integrin alpha(v)beta(3) was not found to support significant beta-cell attachment or migration. Based on our findings, we conclude that integrins alpha(v)beta(5) and alpha(v)beta(1) are important mediators of human beta-cell adhesion and motility, respectively. By supporting fetal beta-cell migration, alpha(v)beta(1) could play an important role in early motile processes required for islet neogenesis.

Adult↗

PubMed2004

Heparin II domain of fibronectin uses alpha4beta1 integrin to control focal adhesion and stress fiber formation, independent of syndecan-4.

Co-signaling events between integrins and cell surface proteoglycans play a critical role in the organization of the cytoskeleton and adhesion forces of cells. These processes, which appear to be responsible for maintaining intraocular pressure in the human eye, involve a novel cooperative co-signaling pathway between alpha5beta1 and alpha4beta1 integrins and are independent of heparan sulfate proteoglycans. Human trabecular meshwork cells isolated from the eye were plated on type III 7-10 repeats of fibronectin (alpha5beta1 ligand) in the absence or presence of the heparin (Hep) II domain of fibronectin. In the absence of the Hep II domain, cells had a bipolar morphology with few focal adhesions and stress fibers. The addition of the Hep II domain increased cell spreading and the numbers of focal adhesions and stress fibers. Cell spreading and stress fiber formation were not mediated by heparan sulfate proteoglycans because treatment with chlorate, heparinase, or soluble heparin did not prevent Hep II domain-mediated cell spreading. Cell spreading and stress fiber formation were mediated by alpha4beta1 integrin because soluble anti-alpha4 integrin antibodies inhibited Hep II domain-mediated cell spreading and soluble vascular cell adhesion molecule-1 (alpha4beta1 ligand)-induced cell spreading. This is the first demonstration of the Hep II domain mediating cell spreading and stress fiber formation through alpha4beta1 integrin. This novel pathway demonstrates a cooperative, rather than antagonistic, role between alpha5beta1 and alpha4beta1 integrins and suggests that interactions between the Hep II domain and alpha4beta1 integrin could modulate the strength of cytoskeleton-mediated processes in the trabecular meshwork of the human eye.

Binding Sites↗

PubMed2005

A protein kinase C/Ras/ERK signaling pathway activates myeloid fibronectin receptors by altering beta1 integrin sialylation.

Here we report that myeloid cells differentiating along the monocyte/macrophage lineage down-regulate the ST6Gal-I sialyltransferase via a protein kinase C/Ras/ERK signaling cascade. In consequence, the beta1 integrin subunit becomes hyposialylated, which stimulates the ligand binding activity of alpha5beta1 fibronectin receptors. Pharmacologic inhibitors of protein kinase C, Ras, and MEK, but not phosphoinositide 3-kinase, block ST6Gal-I down-regulation, integrin hyposialylation, and fibronectin binding. In contrast, constitutively active MEK stimulates these same events, indicating that ERK is both a necessary and sufficient activator of hyposialylation-dependent integrin activation. Consistent with the enhanced activity of hyposialylated cell surface integrins, purified alpha5beta1 receptors bind fibronectin more strongly upon enzymatic desialylation, an effect completely reversed by resialylation of these integrins with recombinant ST6Gal-I. Finally, we have mapped the N-glycosylation sites on the beta1 integrin to better understand the potential effects of differential sialylation on integrin structure/function. Notably, there are three N-glycosylated sites within the beta1 I-like domain, a region that plays a crucial role in ligand binding. Our collective results suggest that variant sialylation, induced by a specific signaling cascade, mediates the sustained increase in cell adhesiveness associated with monocytic differentiation.

Down-Regulation↗

PubMed2000

The tetraspanin CD9 associates with the integrin alpha6beta4 in cultured human epidermal keratinocytes and is involved in cell motility.

Integrins are involved in several ways in keratinocyte physiology, including cell motility. CD9 is a member of the tetraspanin protein family which is found in association with other transmembrane proteins like the integrins. CD9 is expressed in the epidermal tissue, but this expression is not regulated by differentiation. The present work focuses on association of CD9 with the integrin alpha6beta4 in keratinocytes. In vivo, CD9 does not co-localize with alpha6beta4, and is not internalized with the integrin upon basal detachment with dispase. In vitro, CD9 is found partly in co-localization with alpha6beta4 and is internalized with the integrin after keratinocyte detachment with dispase. Using blocking antibodies in a phagokinetic tracks assay, we show that CD9, and to a lesser extent alpha6beta4, but not the tetraspanin CD82, promote motility of subconfluent keratinocytes on collagen I. Our observations also suggest that CD9 is involved in the formation of lamellipodia. We also report that the phorbol ester TPA has no effect on CD9 expression and association with alpha6beta4, but increases keratinocyte motility, possibly through modulation of integrin subunits expression, or through upregulation of collagenase-1 expression. Together, these results confirm that CD9 associates with alpha6beta4 in cultured keratinocytes, possibly in order to regulate the function of the integrin, and that CD9 is involved in keratinocyte motility on collagen. The data suggest that regulation of adhesion characteristics by CD9 in keratinocytes may play a role in epidermal repair.

Adult↗

PubMed1992

A synthetic peptide derived from the carboxy terminus of the laminin A chain represents a binding site for the alpha 3 beta 1 integrin.

The purpose of this study was to identify the binding site(s) within laminin for the alpha 3 beta 1 integrin receptor. It has been previously shown, using proteolytic fragments and anti-laminin antibodies, that the region in laminin for alpha 3 beta 1 integrin binding is localized to the carboxy-terminal region at the end of the long arm (Gehlsen, K. R., E. Engvall, K. Dickerson, W. S. Argraves, and E. Ruoslahti. 1989. J. Biol. Chem. 264:19034-19038; Tomaselli, K. J., D. E. Hall, L. T. Reichardt, L. A. Flier, K. R. Gehlsen, D. C. Turner, and S. Carbonetto. 1990. Neuron. 5:651-662). Using synthetic peptides, we have identified an amino acid sequence within the carboxy-terminal region of the laminin A chain that is recognized by the alpha 3 beta 1 integrin. The amino acid sequence represented by the synthetic peptide GD-6 (KQNCLSSRASFRGCVRNLRLSR residues numbered 3011 to 3032) of the globular domain of the murine A chain supports cell attachment and inhibits cell adhesion to laminin-coated surfaces. By affinity chromatography, peptide GD-6-Sepharose specifically bound solubilized alpha 3 beta 1 from extracts of surface-iodinated cells in a cation-dependent manner, while it did not bind other integrins. In addition, exogenous peptide GD-6 specifically eluted bound alpha 3 beta 1 from laminin-Sepharose columns but did not elute the alpha 3 beta 1 integrin from a fibronectin-Sepharose column. Using integrin subunit-specific monoclonal antibodies, only those antibodies against the alpha 3 and beta 1 subunits inhibited cell adhesion to peptide GD-6-coated surfaces. Finally, a polyclonal antibody made against peptide GD-6 reacted specifically with both murine and human laminin and significantly inhibited cell adhesion to laminin-coated surfaces but not those coated with other matrix proteins. These results identify the laminin A chain amino acid sequence of peptide GD-6 as representing a binding site in laminin for the alpha 3 beta 1 integrin.

Amino Acid Sequence↗

PubMed1992

Identification of amino acid sequences in the integrin beta 1 cytoplasmic domain implicated in cytoskeletal association.

Wild-type and mutant chicken integrin beta 1 subunit (beta 1c) cDNAs were expressed in NIH 3T3 cells and assayed for localization in focal adhesions of cells plated on fibronectin substrates. Focal adhesion localization in stable transfected cells was assayed by indirect immunofluorescent staining with chicken-specific anti-beta 1c antibodies. Mutant beta 1c integrins containing internal deletions of 13 amino acids adjacent to the membrane, delta 759-771, and 20 centrally located amino acids, delta 771-790, localized in focal adhesions demonstrating that sequences required for direction to focal adhesion structures were not limited to one region of the cytoplasmic domain. Point mutations revealed three clusters of amino acids which contribute to localization in focal adhesions. These three clusters or signals are: cyto-1 (764-774), cyto-2 (785-788), and cyto-3 (797-800). The 11-residue cyto-1 signal is only found on integrin beta subunit sequences, except beta 4. Four residues within this region, D764, F768, F771, and E774, could not be altered without reducing focal adhesion staining intensities, and likely form a signal that occupies one side of an alpha helix. Mutations involving two cyto-1 residues, K770 and F771, also appeared to affect heterodimer affinity and specificity. Cyto-2 (785-788,), NPIY, is an NPXY signal that forms a tight turn motif. Cyto-2 provides a structural conformation, which when perturbed by proline removal or addition, inhibits integrin localization in focal adhesions. Cyto-3 (797-800), NPKY, resembles cyto-2, however, the nonconserved proline residue can be replaced without alteration of the localization phenotype. Cyto-3, therefore, constitutes a unique integrin signal, NXXY. Both serine and tyrosine residues at positions 790 and 788, respectively, which have been implicated in integrin phosphorylation/regulation, were conservatively replaced without detectable effect on focal adhesion localization. However, acidic replacements for these amino acids reduced focal adhesion staining intensities, suggesting that phosphorylation at these sites may negatively regulate integrin function.

3T3 Cells↗

PubMed1993

Regulation of alpha 6 beta 1 integrin laminin receptor function by the cytoplasmic domain of the alpha 6 subunit.

The alpha 6 beta 1 integrin is expressed on the macrophage surface in an inactive state and requires cellular activation with PMA or cytokines to function as a laminin receptor (Shaw, L. M., J. M. Messier, and A. M. Mercurio. 1990. J. Cell Biol. 110:2167-2174). In the present study, the role of the alpha 6 subunit cytoplasmic domain in alpha 6 beta 1 integrin activation was examined. The use of P388D1 cells, an alpha 6-integrin deficient macrophage cell line, facilitated this analysis because expression of either the alpha 6A or alpha 6B subunit cDNAs restores their activation responsive laminin adhesion (Shaw, L. S., M. Lotz, and A. M. Mercurio. 1993. J. Biol. Chem. 268:11401-11408). A truncated alpha 6 cDNA, alpha 6-delta CYT, was constructed in which the human cytoplasmic domain sequence was deleted after the GFFKR pentapeptide. Expression of this cDNA in P388D1 cells resulted in the surface expression of a chimeric alpha 6-delta CYT beta 1 integrin that was unable to mediate laminin adhesion or increase this adhesion in response to PMA under normal conditions, i.e., in medium that contained physiological concentrations of Ca++ and Mg++. The alpha 6A-delta CYT transfectants adhered to laminin, however, when Ca++/Mg++ was replaced with 150 microM Mn++. We also assessed the role of serine phosphorylation in the regulation of alpha 6A beta 1 integrin function by site-directed mutagenesis of the two serine residues present in the alpha 6A cytoplasmic domain because this domain is phosphorylated on serine residues in response to stimuli that activate the laminin receptor function of alpha 6 A beta 1. Point mutations were introduced in the alpha 6A cDNA that changed either serine residue #1064 (M1) or serine residue #1071 (M2) to alanine residues. In addition, a double mutant (M3) was constructed in which both serine residues were changed to alanine residues. P388D1 transfectants which expressed these serine mutations adhered to laminin in response to PMA to the same extent as cells transfected with wild-type alpha 6A cDNA. These findings provide evidence for a novel mode of integrin regulation that is distinct from that reported for other regulated integrins (O'Toole, T. E., D. Mandelman, J. Forsyth, S. J. Shattil, E. F. Plow, and M. H. Ginsberg. 1991. Science (Wash. DC). 254:845-847. Hibbs, M. L., H. Xu, S. A. Stacker, and T. A. Springer. 1991. Science (Wash. DC). 251:1611-1613), and they demonstrate that serine phosphorylation of the alpha 6A cytoplasmic domain is not involved in this regulation.

Amino Acid Sequence↗

PubMed1995

Membrane-anchored heparin-binding EGF-like growth factor (HB-EGF) and diphtheria toxin receptor-associated protein (DRAP27)/CD9 form a complex with integrin alpha 3 beta 1 at cell-cell contact sites.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF family of growth factors, which interact with EGF receptor to exert mitogenic activity. The membrane-anchored form of HB-EGF, proHB-EGF, is biologically active, providing mitogenic stimulation to neighboring cells in a juxtacrine mode. ProHB-EGF forms a complex with diphtheria toxin receptor-associated protein (DRAP27)/CD9, a tetra membrane-spanning protein that upregulates the juxtacrine mitogenic activity of proHB-EGF. We explored whether other proteins associate with DRAP27/CD9 and proHB-EGF. Immunoprecipitation with anti-DRAP27/CD9 resulted in preferential coprecipitation of integrin alpha 3 beta 1 from Vero cell, A431 cell and MG63 cell lysates. Anti-integrin alpha 3 or anti-integrin beta 1 coprecipitated DRAP27/CD9 from the same cell lysates. Chemical cross-linking confirmed the physical association of DRAP27/CD9 and integrin alpha 3 beta 1. Using Vero-H cells, which overexpress HB-EGF, we also demonstrated the association of proHB-EGF with DRAP27/CD9 and integrin alpha 3 beta 1. Moreover, colocalization of proHB-EGF, DRAP27/CD9, and integrin alpha 3 beta 1 at cell-cell contact sites was observed by double-immunofluorescence staining. At cell-cell contact sites, DRAP27/CD9 was highly coincident with alpha-catenin and vinculin, suggesting that DRAP27/CD9, proHB-EGF, and integrin alpha 3 beta 1 are colocalized with adherence junction-locating proteins. These results indicate that direct interaction of growth factors and cell adhesion molecules may control cell proliferation during the cell-cell adhesion process.

Animals↗