On the relation between distinct components of the cytoskeleton: an epitope shared by intermediate filaments, microfilaments and cytoplasmic foci.
Monoclonal antibodies were generated against detergent-insoluble cytoskeletal proteins isolated from low-density membrane fractions of rat liver. By immunofluorescence, one of the antibodies stains three distinct structures in cultured rat fibroblast and hepatocyte lines as well as the PtK2 rat-kangaroo kidney epithelial line. These structures are: i) many tangled filaments similar to intermediate filaments (IFs), ii) fewer and variable numbers of straight filaments, and iii) punctate cytoplasmic foci, often most intense around the nucleus. All three of these structures are resistant to extraction by non-ionic detergent. Close examination reveals that the tangled and straight filaments are not stained uniformly, but as a series of bright patches. In cells treated with nocodazole, the antibody reacts strongly with a perinuclear filamentous cage. Very few tangled filaments are detected in these cells, however, the straight filaments and punctate cytoplasmic staining are resistant to nocodazole treatment. Double-label immunofluorescence shows that, even though tangled filament distribution and nocodazole sensitivity are similar to the behavior of vimentin IFs, there is only partial coincidence of staining with either vimentin or cytokeratin IFs. The straight filaments coincide with some actin stress fibers, but the punctate cytoplasmic staining is not related to IFs, actin, or tubulin. Thus, this monoclonal antibody stains a novel group of three seemingly unrelated cytoskeletal structures, including a previously undescribed insoluble nonfilamentous pool. Taken as a whole, two hypotheses are consistent with these data. i) The antigen recognized may be a protein which has a large insoluble cytoplasmic pool and binds both IFs and actin, but only binds to a subset of each class of filaments.(ABSTRACT TRUNCATED AT 250 WORDS)
Turner
J R
JR
Institute of Pathology, Case Western Reserve University, Cleveland, OH 44106.
Tartakoff
A M
AM
D008841:Actin Cytoskeleton / Q000737:chemistry*; D000199:Actins / Q000032:analysis; D000818:Animals; D000911:Antibodies, Monoclonal; D002460:Cell Line; D003429:Cross Reactions; D003593:Cytoplasm / Q000737:chemistry*; D003599:Cytoskeleton / Q000737:chemistry* / Q000187:drug effects; D003902:Detergents / Q000494:pharmacology; D000939:Epitopes / Q000032:analysis; D007382:Intermediate Filaments / Q000737:chemistry*; D007633:Keratins / Q000032:analysis; D007614:Macropodidae; D008856:Microscopy, Fluorescence; D008938:Mitosis / Q000502:physiology; D015739:Nocodazole / Q000494:pharmacology; D051381:Rats; D014404:Tubulin / Q000032:analysis; D014746:Vimentin / Q000032:analysis
D000199:Actins; D000911:Antibodies, Monoclonal; D003902:Detergents; D000939:Epitopes; D014404:Tubulin; D014746:Vimentin; D007633:Keratins; D015739:Nocodazole