Sonnenberg A, de Melker AA, Martinez de Velasco AM et al.Formation of hemidesmosomes in cells of a transformed murine mammary tumor cell line and mechanisms involved in adherence of these cells to laminin and kalinin. J Cell Sci 106:1083-1102

The Netherlands Cancer Institute, Division of Cell Biology, Amsterdam.
Journal of Cell Science (Impact Factor: 5.43). 01/1994; 106 ( Pt 4)(4):1083-102.
Source: PubMed


Keratinocytes attach to an underlying basement membrane by adhesion junctions called hemidesmosomes. We have characterized a cell line, RAC-11P/SD, established from a murine mammary tumor, which differentiates into squamous epithelium and forms well defined hemidesmosomes. These hemidesmosomes contain the integrin alpha 6 beta 4 as well as the hemidesmosomal plaque proteins BP230 and HD1 and are associated with a matrix containing kalinin and laminin. We examined how these cells adhere to laminin and to kalinin present in matrices as well as immunopurified kalinin. We show that adhesion to laminin is energy dependent but does not require an intact actin-containing cytoskeleton. The affinity for kalinin proved to be greater and binding to kalinin was still observed when cells had been treated with deoxyglucose and azide to inhibit metabolic energy. Binding to laminin (or fragment E8), but not to kalinin was partially blocked by a monoclonal antibody specific for the integrin alpha 6 subunit, and only in the initial phase of adhesion. The antibody efficiently blocked adhesion to laminin of cells treated with the microfilament disrupting drug cytochalasin B, but only partially blocked the adhesion of cytochalasin B-treated cells to kalinin, while adherence of cells treated with deoxyglucose and azide to kalinin was blocked completely. The integrin alpha 6 beta 4 is redistributed to the basal surface during adhesion and then is organized into ring-like structures when cells are bound to laminin and localized into hemidesmosomes in cells adhered to kalinin. We suggest that anti-alpha 6 hinders the binding of the alpha 6 beta 4 integrins to its ligands laminin and kalinin, but cannot prevent adhesion when clustering of the integrin has become complete. In addition, there is evidence that adhesion to kalinin is mediated by a second receptor, which associates with the actin-containing cytoskeleton. The presence of such a second receptor is suggested because the cells can spread on kalinin, but not when they have been treated with cytochalasin B. On laminin spreading does not occur, irrespective of whether cells have been treated with cytochalasin B or not. The integrin alpha 3 beta 1, which has been identified as a receptor for kalinin but not for laminin, is strongly expressed in RAC-11P/SD cells and it seems likely that this integrin is responsible for spreading of cells on kalinin.

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    • "Laminin-5 (Laminin γ2 chain): Lam-5 consists of extracellular proteins and is among the components of the basement membrane. The major functions of Lam-5 include binding of epithelial cells to the basement membrane through the formation of hemidesmosomes [4] and the migration of epithelial cells during wound repair [5,6]. In addition, it has been reported that there is a correlation between its expression and tumor progression in various kinds of malignant tumors [7-20]. "
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    Diagnostic Pathology 08/2012; 7(1):105. DOI:10.1186/1746-1596-7-105 · 2.60 Impact Factor
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    • "Confluent Rac 11P cells (Sonnenberg et al., 1993) were allowed to secrete laminin-5 rich extracellular matrix (ECM) for 24 h in a 96 wells plate. Wells were three times washed with PBS and incubated with 20 mM EDTA overnight at 4 • C. Next the wells were washed three times with PBS and blocked for 1 h with 0.35% BSA in PBS at room temperature and washed twice with PBS. "
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    • "To evaluate the involvement of integrins, cell adhesion assays were carried out with anti-integrin-specific antibodies. As previously shown for many cell types (Carter et al., 1991; Sonnenberg et al., 1993; Rousselle and Aumailley, 1994), NHK and HT1080 cell adhesion to LN332 was substantially inhibited by the anti-a3 integrin subunit mAb but not by the anti-a6 subunit mAb (Fig. 2D,G). Combining both mAbs completely abrogated adhesion, demonstrating the involvement of a3b1 and a6b4 integrins in NHK adhesion to LN332, or a3b1, and a6b1 in HT1080 cell adhesion. "
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