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1.
Ryoko Nishiuchi Junichi Takagi Maria Hayashi Hiroyuki Ido Yoshiko Yagi Noriko Sanzen Tsutomu Tsuji Masashi Yamada Kiyotoshi Sekiguchi 《Matrix biology》2006,25(3):189-197
The interactions of cells with basement membranes are primarily mediated via the engagement of laminins by a group of integrin family proteins, including integrins alpha3beta1, alpha6beta1, alpha7beta1 and alpha6beta4. To explore the ligand-binding specificities of these laminin-binding integrins, we produced these integrins, including two alpha7beta1 splice variants (alpha7X1beta1 and alpha7X2beta1), as soluble recombinant proteins and determined their binding specificities and affinities toward a panel of purified laminin isoforms containing distinct alpha chains. Among the five laminin-binding integrins investigated, alpha3beta1 and alpha6beta4 exhibited a clear specificity for laminin-332 (alpha3beta3gamma2) and laminin-511 (alpha5beta1gamma1)/521 (alpha5beta2gamma1), while integrin alpha6beta1 showed a broad specificity, binding to all laminin isoforms with a preference for laminin-111 (alpha1beta1gamma1), laminin-332 and laminin-511/521. The two alpha7beta1 variants were distinct from alpha3beta1, alpha6beta1 and alpha6beta4 in that they did not bind to laminin-332. alpha7X1beta1 bound to all laminins, except laminin-332, with a preference for laminin-211 (alpha2beta1gamma1)/221 (alpha2beta2gamma1) and laminin-511/521, while alpha7X2beta1 bound preferentially to laminin-111 and laminin-211/221. Laminin-511/521 was the most preferred ligand for all the laminin-binding integrins, except for alpha7X2beta1, whereas laminin-411 was the poorest ligand, capable of binding to alpha6beta1 and alpha7X1beta1 with only modest binding affinities. These comprehensive analyses of the interactions between laminin-binding integrins and a panel of laminins clearly demonstrate that the isoforms of both integrins and laminins differ in their binding specificities and affinities, and provide a molecular basis for better understanding of the adhesive interactions of cells with basement membranes of defined laminin compositions. 相似文献
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Antti Tuori Hannu M. Uusitalo Robert E. Burgeson Jeffrey H. Miner Joshua R. Sanes Ismo Virtanen 《The Histochemical journal》1998,30(5):375-381
The distribution of a novel laminin alpha5-chain in the basement membranes of the anterior segment of rat eye was studied. Frozen sections of embryonic day (E)16--17, post-natal day (P)2, 5, 10, 15 and 30 and adult rat eyes were immunostained for laminin chains alpha2, alpha5, beta1, beta2 and gamma1 and for laminin-5, as well as for EHS-laminin, to visualize all basement membranes. Laminin alpha5-, beta1- and gamma1-chain immunoreactivities were found in the basement membranes of the inner and outer layers of optic cup, lens epithelium, further corneal epithelium and skin of the eyelids in E16--17 rat eyes. In P2 and older rat eyes, laminin alpha5-, beta1- and gamma1-chains were all seen in the basement membranes of the corneal and conjunctival epithelium, Descemet's membrane, lens epithelium, ciliary processes, blood vessels and skin of the eyelids. There was a change in the expression pattern of laminin alpha5, beta1- and gamma1-chains in Descemet's membrane from the endothelial side of the membrane (P2--P15 eyes) to both sides of the membrane after P30. Immunoreactivity for laminin-5 was weak in the basement membrane of E16--17 epidermis, but strong in the basement membrane of corneal, conjunctival and eyelid epithelium in P2 and older rat eyes. Laminin alpha2- and beta2-chains were seen in conjunctival and uveal blood vessels in P15 and older rat eyes. The laminin beta2-chain emerged into the basement membrane of conjunctival epithelium in P30 and older rat eyes, suggesting a role for the laminin beta2-chain in the maturation of conjunctiva. The results suggest that laminin alpha5-chain, possibly in laminin-10 (alpha5beta1gamma1), is early and widely expressed in the basement membranes of developing and adult rat eye and, further, that laminin alpha5-chain is a major laminin alpha-chain, partly in coexpression with the alpha3-chain of laminin-5 in the basement membranes of the anterior segment of the eye in developing and adult rats. © 1998 Chapman & Hall 相似文献
4.
James R McMillan Masashi Akiyama Hideki Nakamura Hiroshi Shimizu 《The journal of histochemistry and cytochemistry》2006,54(1):109-118
Multiple laminin isoforms including laminins 5 (alpha3 beta3 gamma2), 6 (alpha3 beta1 gamma1), 10 (alpha5 beta1 gamma1), and possibly laminins 7 (alpha3 beta2 gamma1) and 11 (alpha5 beta2 gamma1) are present in the epidermal basement membrane. However, only the precise epidermal ultrastructural localization of laminin 5 (alpha3 beta3 gamma2) has been elucidated. We therefore determined the precise expression and ultrastructural localization of the alpha5, beta1, beta2, and gamma1 chains in the epidermis. The expression of laminin chains in skin samples was analyzed from patients with epidermolysis bullosa (EB, n=15) that harbor defects in specific hemidesmosome (HD)-associated components. The expression of the alpha5, beta1, and gamma1 chains (present in laminins 10/11) and beta2 chain (laminins 7/11) was unaffected in all intact (unseparated) skin of EB patients including Herlitz junctional EB with laminin-5 defects (n=6). In the basement membrane of human epidermis, the alpha5, beta1, beta2, and gamma1 chains were expressed but also localized to the dermal vessels. Immunogold electron microscopy of normal human epidermis localized the alpha5, beta1, beta2, and gamma1 chains to the upper lamina densa, with between 84% and 92% of labeling restricted to beneath the HDs, similar to laminin 5 (n> or =200 gold particles per sample, sample number n=4) but distinct from collagen IV labeling (with only 63% labeling beneath HDs, p<0.001). Taken together, the majority of the alpha5beta1/beta2gamma1 laminin chains are located beneath HDs. This suggests that laminin-10-associated chains have specific functions or molecular interactions beneath HDs in the epidermal basement membrane. 相似文献
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Fang-Xu Jiang Gaetano Naselli Leonard C Harrison 《The journal of histochemistry and cytochemistry》2002,50(12):1625-1632
Tissue function is regulated by the extracellular microenvironment including cell basement membranes, in which laminins are a major component. Previously, we found that laminin-1 promotes differentiation and survival of pancreatic islet cells. Here we characterize the expression pattern of laminins and their integrin receptors in adult pancreas. Although they are expressed in the basement membrane of acinar cells and duct epithelium, no laminin chains examined were detected extracellularly in the pancreatic islets. In contrast to laminin beta(1)- and gamma(1)-chains, the alpha(1)-chain, unique to laminin-1, was not detected. Laminin-10 (alpha(5)beta(1)gamma(1)) was expressed in acinar tissue, whereas laminins-2 (alpha(2)beta(1)gamma(1)) and -10 were expressed in the blood vessels. The laminin connector molecule, nidogen-1, had a distribution similar to that of laminin beta(1) and gamma(1), whereas fibulin-1 and -2, which compete with nidogen-1, were mostly confined to blood vessels. Integrin subunits alpha(6) and alpha(3) were detected in acinar cells and duct epithelial cells, but alpha(6) was absent in islet cells. Integrin alpha(6)beta(4) was detected only in duct cells, alpha(6)beta(1) in both acinar and ductal cells, and alpha(3)beta(1) in acinar, duct, and islet cells. These findings are a basis for further investigation of the role of extracellular matrix molecules and their receptors in pancreas function. 相似文献
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Kariya Y Mori T Yasuda C Watanabe N Kaneko Y Nakashima Y Ogawa T Miyazaki K 《Journal of molecular histology》2008,39(4):435-446
The basement membrane (BM) proteins laminins, which consist of alpha, beta and gamma chains, play critical roles in the maintenance of tissue structures. One of laminin alpha chains, alpha3 has two isoforms, the truncated form alpha3A and the full-sized form alpha3B. In contrast to alpha3A laminins, little is known about alpha3B laminins. To show the histological distribution of the laminin alpha3B chain, we prepared alpha3B-specific monoclonal antibodies. Immunohistochemical analysis showed that the alpha3B chain was colocalized with the alpha3A, beta3 and gamma2 chains in the epithelial BMs of the skin, esophagus, breast and lung, suggesting the presence of laminin-3B32 (laminin-5B) and laminin-3A32 (laminin-5A). In the lung alveoli, laminin-3B32 was dominant over laminin-3A32, but vice versa in other epithelial BMs. In contrast, the BMs of blood vessels including capillaries were strongly positive for alpha3B, but almost or completely negative for alpha3A, beta3 and gamma2. alpha3B was colocalized with beta1 and gamma1 in these BMs. The alpha3B chain was scarcely detected in the vessels of malignant skin cancers, though the gamma2 and beta3 chains were highly expressed in the cancer cells. These results strongly suggest that the laminin alpha3B chain is widely expressed in vascular BMs of normal tissues, probably as laminin-3B11/3B21 (laminin-6B/7B). 相似文献
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Nishiuchi R Murayama O Fujiwara H Gu J Kawakami T Aimoto S Wada Y Sekiguchi K 《Journal of biochemistry》2003,134(4):497-504
Integrins alpha3beta1 and alpha6beta1 are two major laminin receptors expressed on the surface of mammalian cells. Interactions of cells with laminins through these integrins play important roles in cell adhesion, differentiation, motility, and matrix assembly. To determine the binding specificity and affinity of these integrins toward various types of laminins at the level of direct protein-protein interactions, we purified integrins alpha3beta1 and alpha6beta1 from human placenta, and examined their binding to a panel of laminin isoforms, each containing distinct alpha chains (i.e., laminin-1, laminin-2/4, laminin-5, laminin-8, and laminin-10/11). Integrin alpha3beta1 showed clear specificity for laminin-5 and laminin-10/11, with no significant binding to laminin-1, laminin-2/4, and laminin-8. In contrast, integrin alpha6beta1 showed a broad spectrum of specificity, with apparent binding affinity in the following order: laminin-10/11 > laminin-5 > laminin-1 > laminin-2/4 congruent with laminin-8. Integrin titration assays demonstrated that laminin-10/11 was the most preferred ligand among the five distinct laminin isoforms for both alpha3beta1 and alpha6beta1 integrins. Given that laminin-10/11 is the major basement membrane component of many adult tissues, the interaction of laminin-10/11 with these integrins should play a central role in the adhesive interactions of epithelial cells with underlying basement membranes. 相似文献
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Champliaud MF Virtanen I Tiger CF Korhonen M Burgeson R Gullberg D 《Experimental cell research》2000,259(2):326-335
Laminins assemble into trimers composed of alpha, beta, and gamma chains which posttranslationally are glycosylated and sometimes proteolytically cleaved. In the current paper we set out to characterize posttranslational modifications and the laminin isoforms formed by laminin alpha1 and alpha5 chains. Comparative pulse-chase experiments and deglycosylation studies in JAR cells established that the M(r) 360,000 laminin alpha1 chain is glycosylated into a mature M(r) 400,000 band while the M(r) 370,000 laminin alpha5 chain is glycosylated into a M(r) 390,000 form that upon secretion is further processed into a M(r) 380,000 form. Hence, despite the shorter peptide length of alpha1 chain in comparison with the alpha5 chain, secreted alpha1 assumes a larger size in SDS-PAGE due to a higher degree of N-linked glycosylation and due to the lack of proteolytic processing. Immunoprecipitations and Western blotting of JAR laminins identified laminin alpha1 and laminin alpha5 chains in laminin-1 and laminin-10. In placenta laminin alpha1 chain (M(r) 400,000) and laminin alpha5 chain (M(r) 380, 000/370,000 doublet) were found in laminin-1/-3 and laminin-10/-11. Immunohistochemically we could establish that the laminin alpha1 chain in placenta is deposited in the developing villous and trophoblast basement membrane, also found to contain laminin beta2 chains. Surprisingly, a fraction of the laminin alpha1 chain from JAR cells and placenta could not be precipitated by antibodies to laminin beta1-beta3 chains, possibly pointing to an unexpected complexity in the chain composition of alpha1-containing laminin isoforms. 相似文献
9.
Genersch E Ferletta M Virtanen I Haller H Ekblom P 《European journal of cell biology》2003,82(3):105-117
Human ECV304 cells respond reproducibly by tube formation to complex basement membrane matrices. Laminins are major glycoproteins of basement membranes. We therefore studied the ability of ECV304 cells to attach to defined laminin isoforms and to fibronectin, and identified the involved laminin receptors. The cells bound poorly to fibronectin, to some extent to laminin-1, whereas laminin-2/4 and -10/11 were strong adhesive substrates. Antibody perturbation assays showed that adhesion to laminin-1 was mediated by integrin alpha6beta1, and adhesion to laminin-2/4 by cooperative activity of integrins alpha3beta1 and alpha6beta1. Adhesion of ECV 304 cells to laminin-10/11 was mainly mediated by integrins alpha3beta1, with minor involvement of alpha6beta1/4 and alphavbeta3. Solid-phase binding assays confirmed that integrin alphavbeta3 binds human laminin-10/11 and -10, in an RGD-dependent fashion. Although integrin alphavbeta3 played a very minor role in cell adhesion to laminin-10/11, this interaction facilitated growth factor-induced proliferation of ECV304 cells. In response to FGF-2 or VEGF, the cells proliferated better when attached on laminin-10/11 than on laminin-1, -2/4, or gelatin. The proliferation induced by the joint application of laminin-10/11 and either one of the growth factors could be blocked by antibodies against integrin alphavbeta3. Fragments of several other basement membrane components are known to interact with alphavbeta3. The current data show that that integrin alphavbeta3 can bind intact alpha5-containing laminin trimers. Since the laminin alpha5 chain is broadly expressed in adult basement membranes, this interaction could be physiologically important. Our data suggest that this interaction is involved in the regulation of cellular responses to growth factors known to be involved in epithelial and endothelial development. 相似文献
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Laminins are a family of glycoproteins which are ubiquitous components of basement membranes and play key structural and functional roles. Eleven isoforms have been identified to date; each is an alpha beta gamma heterotrimer assembled from a repertoire of five alpha, three beta and two gamma chains. Studies of laminin-11 (alpha 5 beta 2 gamma 1) illustrate the unique expression patterns and distinct functions that can be displayed by laminin isoforms. Laminin-11 is found in the glomerular basement membrane in kidney, in the neuromuscular synaptic cleft in skeletal muscle and in other tissues such as placenta and lung. Mice lacking laminin-11 exhibit defective glomerular filtration and developmental defects in neuromuscular synapse formation, with Schwann cells invading the synaptic cleft. Consistent with these observations, both motoneurons and Schwann cells distinguish laminin-11 from other isoforms in vitro. These results suggest that laminin-11 is a structural component of the basement membrane which influences cell behavior in physiologically relevant ways. A greater understanding of laminin-11 assembly and basement membrane incorporation could provide a logical basis for therapy in merosin-deficient congenital muscular dystrophy. 相似文献
12.
Laminin represents a macromolecule family. The heterotrimeric isoforms of laminin can be determined by immunohistochemical demonstration of the single chains. The laminin chain heterogeneity of the basement membrane in adult human major salivary glands was evaluated in relation to cellular differentiation of the epithelia and the stromal compartment. Monoclonal antibodies to the laminin alpha1, alpha3 (BM165) chains and epiligrin reacted with the basement membranes of serous and mucous acini and of intercalated, striated and excretory ducts. As evidenced by a double-labelling technique, the alpha2 chain showed a spatial association with the myoepithelium of the acini, whereas the ductal basement membranes containing no myoepithelial cells were negative. Almost exclusively, beta1 chain was detected in acinar basement membrane, beta2 chain whereas in ductal basement membrane. gamma2 chain is a unique chain belonging to the laminin-5 isoform. It was restricted to the ductal basement membrane. alpha1, alpha2, beta1, beta2 and gamma1 chains were detected in nerves of salivary tissue and alpha1, alpha3, beta1, beta2 and gamma1 chains and epiligrin in blood vessels. Our results indicate that the acinar ductal unit contains basement membranes with different isoforms, which relate to cell differentiation and cell function. © 1998 Chapman & Hall 相似文献
13.
Kikkawa Y Moulson CL Virtanen I Miner JH 《The Journal of biological chemistry》2002,277(47):44864-44869
The Lutheran blood group glycoprotein (Lu), also known as basal cell adhesion molecule, is an Ig superfamily transmembrane receptor for laminin alpha5. Lu is expressed on the surface of a subset of muscle and epithelial cells in diverse tissues and is thought to be involved in both normal and disease processes, including sickle cell disease and cancer. Here we investigated the binding of Lu to laminin alpha5 in vivo and in vitro. We prepared a soluble recombinant Lu (sol-Lu) composed of the Lu extracellular domain and a His(6) tag. Sol-Lu bound specifically to laminin-10/11 (alpha5beta1/beta2gamma1) in enzyme-linked immunosorbent assays and bound to bona fide basement membranes containing laminin alpha5 in tissue sections. Sol-Lu did not bind to tissue sections of laminin alpha5 knockout embryos, despite the fact that the four other alpha chains were present. To identify the Lu-binding site on laminin alpha5, we prepared modified alpha5 cDNAs encoding chimeric laminins containing all or part of the laminin alpha1 G domain in place of the analogous alpha5 regions. These constructs were used to generate transgenic mice. Proteins derived from transgenes were detected in basement membranes and were assayed for their ability to bind Lu by examining the localization of endogenous Lu and the binding of sol-Lu applied to tissue sections. Our results demonstrate that the alpha5 LG3 module is essential for Lu binding to laminin alpha5. 相似文献
14.
Antibodies against domain E3 of laminin-1 and integrin alpha 6 subunit perturb branching epithelial morphogenesis of submandibular gland, but by different modes 总被引:5,自引:2,他引:3
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《The Journal of cell biology》1995,129(2):521-534
Branching epithelial morphogenesis requires interactions between the surrounding mesenchyme and the epithelium, as well as interactions between basement membrane components and the epithelium. Embryonic submandibular gland was used to study the roles of two mesenchymal proteins, epimorphin and tenascin-C, as well as the epithelial protein laminin-1 and one of its integrin receptors on branching morphogenesis. Laminin-1 is a heterotrimer composed of an alpha 1 chain and two smaller chains (beta 1 and gamma 1). Immunofluorescence revealed a transient expression of laminin alpha 1 chain in the epithelial basement membrane during early stages of branching morphogenesis. Other laminin-1 chains and alpha 6, beta 1, and beta 4 integrin subunits seemed to be expressed constitutively. Expression of epimorphin, but not tenascin-C, was seen in the mesenchyme during early developmental stages, but a mAb against epimorphin did not perturb branching morphogenesis of this early epithelium. In contrast, inhibition of branching morphogenesis was seen with a mAb against the carboxy terminus of laminin alpha 1 chain, the E3 domain. An inhibition of branching was also seen with a mAb against the integrin alpha 6 subunit. The antibodies against laminin alpha 1 chain and integrin alpha 6 subunit perturbed development in distinct fashions. Whereas treatment with the anti-E3 resulted in discontinuities of the basement membrane at the tips of the branching epithelium, treatment with the mAb against alpha 6 integrin subunit seemed to leave the basement membrane intact. We suggest that the laminin E3 domain is involved in basement membrane formation, whereas alpha 6 beta 1 integrin binding to laminin-1 may elicit differentiation signals to the epithelial cells. 相似文献
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Immunohistochemical localization of laminin and fibronectin isoforms in human placental villi. 总被引:4,自引:0,他引:4
We studied the localization of laminin alpha1, alpha2, alpha3, alpha5, beta1, beta2, and gamma1 chains and extradomain A- (EDA), EDB-, and oncofetal fibronectin by immunohistochemistry in human placental villi during placental development. The laminin alpha2, alpha5, beta1, beta2, and gamma1 chains were detected in the trophoblastic basement membrane (BM) at all stages of gestation, suggesting the presence of laminin-2, -4, -10, and -11 trimers. The laminin alpha1 chain was selectively found at sites where the villous BM was in contact with proliferating cells in trophoblastic islands or columns. EDA-Fn, but not other Fn isoforms, was found in the trophoblastic BM during the first trimester. The laminin alpha2, beta1, beta2, and gamma1 chains were detected in the villous stroma and capillaries throughout placental development, while the laminin alpha5 chain emerged distinctly during development. Extensive EDA-Fn immunoreactivity was found in first-trimester villous stroma, but distinctly fewer Fn isoforms were seen in the villous stroma during the later stages of gestation. Our results also suggest that, during the formation of new villi, laminins are not found in trophoblastic sprouts before the ingrowth of the villous mesenchyme. Rather, laminins may be deposited at the villous epithelial-mesenchymal interface. Furthermore, the results show that distinct changes occur in the localization of various laminin and Fn isoforms during the maturation of villous trophoblastic and capillary BMs. 相似文献
17.
Inhibition of laminin alpha 1-chain expression leads to alteration of basement membrane assembly and cell differentiation 总被引:6,自引:0,他引:6
《The Journal of cell biology》1996,133(2):417-430
The expression of the constituent alpha 1 chain of laminin-1, a major component of basement membranes, is markedly regulated during development and differentiation. We have designed an antisense RNA strategy to analyze the direct involvement of the alpha 1 chain in laminin assembly, basement membrane formation, and cell differentiation. We report that the absence of alpha 1-chain expression, resulting from the stable transfection of the human colonic cancer Caco2 cells with an eukaryotic expression vector comprising a cDNA fragment of the alpha 1 chain inserted in an antisense orientation, led to (a) an incorrect secretion of the two other constituent chains of laminin-1, the beta 1/gamma 1 chains, (b) the lack of basement membrane assembly when Caco2-deficient cells were cultured on top of fibroblasts, assessed by the absence of collagen IV and nidogen deposition, and (c) changes in the structural polarity of cells accompanied by the inhibition of an apical digestive enzyme, sucrase-isomaltase. The results demonstrate that the alpha 1 chain is required for secretion of laminin-1 and for the assembly of basement membrane network. Furthermore, expression of the laminin alpha 1-chain gene may be a regulatory element in determining cell differentiation. 相似文献
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Paola Spessotto Anna Gronkowska Rainer Deutzmann Roberto Perris Alfonso Colombatti 《Matrix biology》2003,22(4):351-361
To identify the laminin isoforms of the basement membranes that could be implicated in the extravasation process of neoplastic lymphocytes, a number of purified laminins and one native renal laminin complex were comparatively investigated for their ability to promote migration of neoplastic lymphocytes in vitro. The identity/composition of a human placental laminin complex was asserted by combining immunochemical assays, sequence determination of tryptic peptides, and ultrastructural analysis to be composed predominantly of laminin-10 in which the coiled-coil C-terminal regions and the G globular domain of the alpha5 chain were preserved intact despite the enzymatic treatment used for its isolation. Lymphoma and leukemic cell lines failed to migrate towards laminin-4, -9, -11, moved poorly in response to laminin-1, -2/4, -5 and the renal laminin complex, but markedly locomoted towards the subendothelial laminin-8 and -10. The motility-promoting interaction with these latter laminins was interchangeably mediated by the alpha3beta1 and alpha6beta1 integrins. Lymphocyte locomotion on laminins assayed in the presence of cytokines was either reduced or enhanced suggesting that local cytokine milieu could further influence motility response. 相似文献
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Kariya Y Ishida K Tsubota Y Nakashima Y Hirosaki T Ogawa T Miyazaki K 《Journal of biochemistry》2002,132(4):607-612
Laminin-5, a heterotrimer of laminin alpha3, beta3, and gamma2 chains, is an essential component of various epithelial basement membranes, and it strongly promotes cellular adhesion and motility in vitro. In this study, we established an efficient expression system of human recombinant laminin-5 (rLN5), in which full-length cDNAs encoding the human laminin alpha3, beta3, and gamma2 chains were introduced into the human embryonic kidney cell line HEK293. rLN5 was purified from the conditioned medium of the HEK293 transfectant (LN5-HEK) by immuno-affinity chromatography in a yield of 1 mg protein/liter, about 10 times higher than that of a natural LN5 from human gastric cancer cells. rLN5 was indistinguishable from the natural LN5 in its protein composition and biological activity. In addition, analysis of HEK293 transfectants expressing two exogenous LN5 subunits showed that the alpha3/gamma2 chains and the beta3/gamma2 chains, but not the alpha3/beta3 chains, were secreted as heterodimers, suggesting an important role of the gamma2 chain in the association of the three LN5 subunits. The expression system of rLN5 can be used as an important tool to understand the biological functions of this laminin and may be applicable to future regenerative medicine. 相似文献