Carter W G, Kaur P, Gil S G, Gahr P J, Wayner E A
Fred Hutchinson Cancer Research Center, University of Washington, Seattle 98104.
J Cell Biol. 1990 Dec;111(6 Pt 2):3141-54. doi: 10.1083/jcb.111.6.3141.
Basal cells of stratified epidermis are anchored to the basement membrane zone (BMZ) of skin via hemidesmosomes. We previously identified integrin alpha 3 beta 1, in focal adhesions (FAs), of cultured human keratinocytes (HFKs) as a mediator of HFK adhesion to secreted BMZ-like extracellular matrix (ECM; Carter, W.G., E.A. Wayner, T.S. Bouchard, and P. Kaur. 1990. J. Cell Biol. 110: 1387-1404). Here, we have examined the relation of integrins alpha 6 beta 4 and alpha 3 beta 1, to bullous pemphigoid antigen (BPA), a component of hemidesmosomes. We conclude that alpha 6 beta 4 in HFKs localizes in a new stable anchoring contact (SAC) that cooperates with alpha 3 beta 1-FAs to mediate adhesion to ECM, based on the following. (a) Comparison of secreted ECM, with exogenous laminin, fibronectin and collagen identified ECM as the preferred ligand for HFK adhesion and spreading and for formation of both alpha 6 beta 4-SACs and alpha 3 beta 1-FAs. (b) Inhibition of HFK adhesion with combined anti-alpha 3 beta 1 (P1B5) and anti-alpha 6 beta 4 (GoH3) antibodies indicated that both receptors were functional in adhesion to ECM while alpha 3 beta 1 played a dominant role in spreading. (c) alpha 6 beta 4 colocalized with BPA in SACs that were proximal to but excluded from FAs. Both alpha 6 beta 4-SACs and alpha 3 beta 1-FAs were in contact with the adhesion surface as indicated by antibody exclusion and interference reflection microscopy. (d) In contrast to alpha 3 beta 1-FAs, alpha 6 beta 4-SACs were present only in nonmotile cells, not associated with stress fibers, and were relatively stable to detergents and urea, suggesting a nonmotile, or anchoring function for SACs and motility functions for alpha 3 beta 1-FAs. (e) alpha 6 beta 4 formed a detergent-insoluble complex with exogenous ECM in an affinity isolation procedure, confirming the ability of an unidentified ECM ligand to interact with alpha 6 beta 4. (f) We suggest that alpha 6 beta 4/BPA-SACs in culture restrict migration of HFKs on ECM while alpha 3 beta 1-FAs form dynamic adhesions in spreading and migrating cells. alpha 6 beta 4/BPA-SACs in culture bear functional and compositional similarities to hemidesmosomes in skin.
复层表皮的基底细胞通过半桥粒锚定在皮肤的基底膜带（BMZ）上。我们之前已确定，培养的人角质形成细胞（HFKs）粘着斑（FAs）中的整合素α3β1是HFKs与分泌的类BMZ细胞外基质（ECM）黏附的介质（Carter, W.G., E.A. Wayner, T.S. Bouchard, and P. Kaur. 1990. J. Cell Biol. 110: 1387 - 1404）。在此，我们研究了整合素α6β4和α3β1与大疱性类天疱疮抗原（BPA，半桥粒的一种成分）之间的关系。基于以下几点，我们得出结论：HFKs中的α6β4定位于一种新的稳定锚定接触（SAC），它与α3β1-FAs协同介导对ECM的黏附。（a）将分泌的ECM与外源性层粘连蛋白、纤连蛋白和胶原蛋白进行比较，确定ECM是HFKs黏附、铺展以及形成α6β4-SACs和α3β1-FAs的首选配体。（b）用抗α3β1（P1B5）和抗α6β4（GoH3）抗体联合抑制HFKs黏附表明，两种受体在与ECM黏附中均有功能，而α3β1在铺展中起主导作用。（c）α6β4与BPA在紧邻FAs但不包含在FAs内的SACs中共定位。抗体排除和干涉反射显微镜显示，α6β4-SACs和α3β1-FAs均与黏附表面接触。（d）与α3β1-FAs不同，α6β4-SACs仅存在于非运动细胞中，与应力纤维无关，并且对去污剂和尿素相对稳定，这表明SACs具有非运动或锚定功能，而α3β1-FAs具有运动功能。（e）在亲和分离实验中，α6β4与外源性ECM形成了去污剂不溶性复合物，证实了一种未鉴定的ECM配体与α6β4相互作用的能力。（f）我们认为，培养中的α6β4/BPA-SACs限制了HFKs在ECM上的迁移，而α3β1-FAs在铺展和迁移细胞中形成动态黏附。培养中的α6β4/BPA-SACs在功能和组成上与皮肤中的半桥粒相似。
