Molecular Interactions in Intermediate‐Sized Filaments Revealed by Chemical Cross‐Linking

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Abstract
Certain glia cells, notably astrocytes and tumor cells derived from them, express simultaneously 2 types of proteins of intermediate-sized filaments, vimentin and glia filament protein (GFP). An established human glioma (astrocytoma) cell culture line (U 333 CG/343 MG) in which both proteins are seen in partly overlapping fibrillar structures was examined by immunofluorescence microscopy for the possible existence of heteropolymer filaments of these 2 proteins, using reversible oxidative cross-linking facilitated by the 1,10-phenanthroline-cupric ion complex. Dimeric cross-link products are characterized by 1-dimensional and 2-dimensional gel electrophoresis under non-reducing and reducing conditions as well as by peptide mapping. The relatively large proportions of heterodimers of vimentin and GFP obtained in cytoskeletal filaments cross-linked in this way, demonstrate the frequency of heteropolymer filaments in this cell as well as the frequency of face-to-face pairs of GFP and vimentin in such filaments. Together with related observations on heteropolymer filaments between vimentin and desmin in some smooth muscle cells, this is discussed as evidence for common principles of molecular arrangement of vimentin, GFP and desmin, at least in the cysteine-containing surface domains. The results are discussed in relation to cytoskeletal changes during glial differentiation.