Integrins: versatility, modulation, and signaling in cell adhesion.

One decade ago, vascular endothelium was commonly considered a "non-stick" lining of blood vessels that functioned only to prevent blood coagulation and to separate the vascular space from tissues. By comparison to many other cell types, endothelial cells were thought to be less active, less complex, and less interesting. Since that time, research concerning the endothelium has expanded dramatically and produced a new image of the vascular lining as an active participant in a wide variety of pathophysiological processes, including inflammation and immunity. Nowhere has the excitement been more intense than in the study of the molecular mechanisms of leukocyte adhesion to endothelium. Recent efforts resulted in the identification, characterization, and cloning of multiple endothelial cell-surface glycoproteins that support adhesion through an interaction with specific ligands (or counter-receptors) on leukocytes. The selectins, two of which are found on endothelium and one on leukocytes, support adhesion through the recognition of carbohydrates. Endothelial members of the immunoglobulin superfamily including ICAM-1 and VCAM-1/INCAM-110 bind to leukocyte cell-surface integrins. In various combinations, these and other molecules support leukocyte adhesion to the vessel wall and extravasation, key steps in our response to infection and tissue injury.

Endothelial- leukocyte adhesion molecules

▪ Abstract Von Willebrand factor (VWF) is a blood glycoprotein that is required for normal hemostasis, and deficiency of VWF, or von Willebrand disease (VWD), is the most common inherited bleeding disorder. VWF mediates the adhesion of platelets to sites of vascular damage by binding to specific platelet membrane glycoproteins and to constituents of exposed connective tissue. These activities appear to be regulated by allosteric mechanisms and possibly by hydrodynamic shear forces. VWF also is a carrier protein for blood clotting factor VIII, and this interaction is required for normal factor VIII survival in the circulation. VWF is assembled from identical ≈250 kDa subunits into disulfide-linked multimers that may be >20,000 kDa. Mutations in VWD can disrupt this complex biosynthetic process at several steps to impair the assembly, intracellular targeting, or secretion of VWF multimers. Other VWD mutations impair the survival of VWF in plasma or the function of specific ligand binding sites. This growing...

Biochemistry and Genetics of von Willebrand Factor

The dystrophin-glycoprotein complex was tested for interaction with several components of the extracellular matrix as well as actin. The 156-kD dystrophin-associated glycoprotein (156-kD dystroglycan) specifically bound laminin in a calcium-dependent manner and was inhibited by NaCl (IC50 = 250 mM) but was not affected by 1,000-fold (wt/wt) excesses of lactose, IKVAV, or YIGSR peptides. Laminin binding was inhibited by heparin (IC50 = 100 micrograms/ml), suggesting that one of the heparin-binding domains of laminin is involved in binding dystroglycan while negatively charged oligosaccharide moieties on dystroglycan were found to be necessary for its laminin-binding activity. No interaction between any component of the dystrophin-glycoprotein complex and fibronectin, collagen I, collagen IV, entactin, or heparan sulfate proteoglycan was detected by 125I-protein overlay and/or extracellular matrix protein-Sepharose precipitation. In addition, laminin-Sepharose quantitatively precipitated purified dystrophin-glycoprotein complex, demonstrating that the laminin-binding site is accessible when dystroglycan is associated with the complex. Dystroglycan of nonmuscle tissues also bound laminin. However, the other proteins of the striated muscle dystrophin-glycoprotein complex appear to be absent, antigenically dissimilar or less tightly associated with dystroglycan in nonmuscle tissues. Finally, we show that the dystrophin-glycoprotein complex cosediments with F-actin but does not bind calcium or calmodulin. Our results support a role for the striated muscle dystrophin-glycoprotein complex in linking the actin-based cytoskeleton with the extracellular matrix. Furthermore, our results suggest that dystrophin and dystroglycan may play substantially different functional roles in nonmuscle tissues.

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A role for the dystrophin-glycoprotein complex as a transmembrane linker between laminin and actin

The laminins are a family of glycoproteins that provide an integral part of the structural scaffolding of basement membranes in almost every animal tissue. Each laminin is a heterotrimer assembled from alpha, beta, and gamma chain subunits, secreted and incorporated into cell-associated extracellular matrices. The laminins can self-assemble, bind to other matrix macromolecules, and have unique and shared cell interactions mediated by integrins, dystroglycan, and other receptors. Through these interactions, laminins critically contribute to cell differentiation, cell shape and movement, maintenance of tissue phenotypes, and promotion of tissue survival. Recent advances in the characterization of genetic disruptions in humans, mice, nematodes and flies have revealed developmental roles for the different laminin subunits in diverse cell types, affecting differentiation from blastocyst formation to the post-natal period. These genetic defects have challenged some of the previous concepts about basement membranes and have shed new light on the diversity and complexity of laminin functions as well as established the molecular basis of several human diseases.

https://anatomypubs.onlinelibrary.wiley.com/doi/pdf/10.1002/%28SICI%291097-0177%28200006%29218%3A2%3C213%3A%3AAID-DVDY1%3E3.0.CO%3B2-R

Form and function: the laminin family of heterotrimers.

The α6β4 Integrin Is a Receptor for both Laminin and Kalinin

The alpha 6 beta 4 complex is a member of the integrin superfamily of adhesion receptors. A human keratinocyte lambda gt11 cDNA library was screened using a monoclonal antibody directed against the beta 4 subunit. Two cDNAs were selected and subsequently used to isolate a complete set of overlapping cDNA clones. The beta 4 subunit consists of 1778 amino acids with a 683 amino acid extracellular domain, a 23 amino acid transmembrane domain and an exceptionally long cytoplasmic domain of 1072 residues. The deduced amino-terminal sequence is in good agreement with the published amino-terminal sequence of purified beta 4. The extracellular domain contains five potential N-linked glycosylation sites and four cysteine-rich homologous repeat sequences. The extracellular part of the beta 4 subunit sequence shows 35% identify with other integrin beta subunits, but is the most different among this class of molecules. The transmembrane region is poorly conserved, whereas the cytoplasmic domain shows no substantial identity in any region to the cytoplasmic tails of the known beta sequences or to other protein sequences. The exceptionally long cytoplasmic domain suggests distinct interactions of the beta 4 subunit with cytoplasmic proteins.

Cloning and sequence analysis of beta‐4 cDNA: an integrin subunit that contains a unique 118 kd cytoplasmic domain.

Glycoproteins V and Ib-IX form a noncovalent complex in the platelet membrane.

We have isolated cDNAs encoding the α6 subunit from a γ gt11 expression library from human keratinocytes by combined screening with a rabbit polyclonal anti-α6 antibody and the polymerase chain reaction. The α6 subunit encoded by this cDNA consists of 1050 amino acids with a 991-amino-acid extracellular, a 23-amino-acid transmembrane and a 36-amino-acid cytoplasmic domain. The extracellular domain contains three putative divalent cation-binding sites and nine potential N-linked glycosylation sites. From a cDNA library from normal human mammary gland cells two different cDNAs for α6 were isolated, one of which is identical to the above cDNA. The two α6 subunits, called α6A and α6B, encoded by the two cDNAs each have a unique cytoplasmic domain, that of α6B being 18 amino acids longer than that of α6A.



Different carcinoma cell lines contain transcripts for both α6 subunits. K 562 leukemic cells have little α6A or α6B mRNAs. The overall level of expression varies in the carcinoma cell lines, but reflects α6 cell surface expression. In A375 melanoma cells, however, cell surface expression of α6 was low in spite of a high level of mRNA. This suggests that other mechanisms may be involved in regulating the expression of α6 on the surface of these cells. The mRNA for both α6 subunits is around 6 kb.



The α6 subunits are similar to other α subunits (26–31% identity with cleaved α subunits) of the integrin family but they are more similar to the α3 subunit (40% identity). This high degree of similarity may be the basis for their functional resemblance since both α3 and α6 subunits, when associated with β1, function as laminin receptors and bind to the long arm of laminin. The genes for α6 and β4, the alternative β subunit with which α6 combines on certain epithelial cells, were mapped to chromosome 2 and 17q11-qter, respectively.

Molecular cloning of the human alpha 6 integrin subunit. Alternative splicing of alpha 6 mRNA and chromosomal localization of the alpha 6 and beta 4 genes.

A. Sonnenberg

Papers

The α6β4 Integrin Is a Receptor for both Laminin and Kalinin

Cloning and sequence analysis of beta‐4 cDNA: an integrin subunit that contains a unique 118 kd cytoplasmic domain.

Glycoproteins V and Ib-IX form a noncovalent complex in the platelet membrane.

Molecular cloning of the human alpha 6 integrin subunit. Alternative splicing of alpha 6 mRNA and chromosomal localization of the alpha 6 and beta 4 genes.

P-selectin mediates Ca(2+)-dependent adhesion of activated platelets to many different types of leukocytes: detection by flow cytometry.