Cell localization and redistribution of the 67 kD laminin receptor and alpha 6 beta 1 integrin subunits in response to laminin stimulation: an immunogold electron microscopy study.
The 67 kDa laminin receptor (67LR), one of several cell surface laminin-binding proteins, is involved in the interactions between cancer cells and laminin during tumor invasion and metastasis. A 37 kDa polypeptide (37LRP), previously identified as the 67LR precursor, is abundantly present in the cytoplasm and has been implicated in polysome formation. To better understand the cellular localization of the 67LR and its precursor, transmission electron microscopic studies of human melanoma A2058 cells were carried out using immunogold labeling and a variety of antibodies: (a) affinity purified antibodies directed against 37LRP cDNA-derived synthetic peptides; (b) anti-67LR monoclonal antibodies raised against intact human small cell lung carcinoma cells; and (c) monoclonal antibodies against the subunits of the integrin laminin receptor, alpha 6 beta 1. Double-labeling immunocyto-chemistry revealed that anti-67LR monoclonal antibodies as well as anti-37 LRP antibodies recognized antigens that were localized in the cytoplasm in electron dense structures. As expected, cell membrane labeling was also observed. Surprisingly, alpha 6 and beta 1 integrin subunits were detected in the same cytoplasmic structures positive for the 67LR and the 37LRP. After addition of soluble laminin to A2058 cells in suspension, the number of labeled cytoplasmic structures increased especially in the vicinity of the plasma membrane, and were exported onto the cell surface. Neither fibronectin nor BSA induced such an effect. The data demonstrate that the 67 LR and the 37 LRP antibodies detect colocalized antigens that are in cytoplasmic structures with alpha 6 beta 1 integrin.(ABSTRACT TRUNCATED AT 250 WORDS)
Romanov
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Structural Biology Section, NCI-Frederick Cancer Research and Development Center, Maryland 21702-1201.
Sobel
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pinto da Silva
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Menard
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Castronovo
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D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D039121:Integrin alpha6beta1; D016023:Integrins / Q000378:metabolism*; D007797:Laminin / Q000494:pharmacology*; D008545:Melanoma / Q000378:metabolism / Q000556:secondary / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron; D008970:Molecular Weight; D009361:Neoplasm Invasiveness; D017474:Receptors, Laminin / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure
D039121:Integrin alpha6beta1; D016023:Integrins; D007797:Laminin; D017474:Receptors, Laminin