Heat shock protein HSP90 and its association with the cytoskeleton: a morphological study.
To investigate the cellular localization of the 90-kilodalton heat shock protein (HSP90) and its interaction with the cytoskeleton, we performed single- and double-staining immunofluorescence microscopy of cytoskeletal proteins and HSP90 in the absence and presence of cytoskeletal inhibitors. As a model, we used a human endometrial adenocarcinoma cell line (Ishikawa cells), which expresses HSP90. We confirmed the recently reported colocalization of HSP90 with microtubules. However, Ishikawa cells treated with 10(-5) M of the antimicrotubule agents colchicine or triethyl lead showed residual filamentous structures stained with anti-HSP90 antibodies, while no microtubules were visualized with anti-tubulin antibodies. In the presence of 10(-5) M cytochalasin B, the microfilament staining of the cells disappeared, while residual filamentous structures were labeled with anti-HSP90 antibodies. Furthermore, Ishikawa cells treated with 10(-5) M triethyl lead and stained with anti-HSP90 antibodies demonstrated residual filamentous structures, clearly different from those of reorganized vimentin intermediate filaments. Conversely, similar reorganized morphology of filamentous structures stained with both anti-HSP90 and anti-cytokeratins antibodies were observed when Ishikawa cells were treated with 2 x 10(-5) M cytochalasin B and 2 x 10(-5) M colchicine. HSP90 was also present in Ishikawa cell preparations of the Triton X-100 insoluble cytoskeleton. In addition, Triton-insoluble cytoskeleton treated with 10(-5). M triethyl lead and double stained with anti-HSP90 and anti-vimentin antibodies demonstrated clearly different filamentous patterns, when exposed on the same photographic plaque.(ABSTRACT TRUNCATED AT 250 WORDS)
Fostinis
Y
Y
Department of Basic Sciences, School of Medicine, University of Crete, Heraklion, Greece.
Theodoropoulos
P A
PA
Gravanis
A
A
Stournaras
C
C
D000230:Adenocarcinoma; D003598:Cytoskeletal Proteins / Q000032:analysis* / Q000037:antagonists & inhibitors; D003902:Detergents; D016889:Endometrial Neoplasms; D005260:Female; D006360:Heat-Shock Proteins / Q000032:analysis*; D006801:Humans; D015151:Immunoblotting; D008856:Microscopy, Fluorescence; D009363:Neoplasm Proteins / Q000032:analysis; D017830:Octoxynol; D011092:Polyethylene Glycols; D012995:Solubility; D014407:Tumor Cells, Cultured
D003598:Cytoskeletal Proteins; D003902:Detergents; D006360:Heat-Shock Proteins; D009363:Neoplasm Proteins; D011092:Polyethylene Glycols; D017830:Octoxynol