High molecular weight polypeptides (270,000-340,000) from cultured cells are related to hog brain microtubule-associated proteins but copurify with intermediate filaments.

Abstract

High MW polypeptides (HMWP) 270,000 to 340,000 were major components of intermediate filaments prepared by Triton X-100 extraction after spreading of rat glioma C6, [human cervical carcinoma] HeLa, Chinese hamster ovary, and SV40-transformed Chinese hamster lung cells. C6 HMWP resembled high MW microtubule-associated proteins from hog brain by 4 criteria: comigration in electrophoresis on high-resolution sodium dodecyl sulfate/polyacrylamide gels, 1-dimensional peptide mapping, phosphorylation in vitro with [.gamma.-32P]ATP and ability to promote microtubule assembly in vitro. HMWP were also major components of 1-time polymerized C6 microtubule preparations, which contained a sizable amount of intermediate filaments. The predominant part of HMWP present in these microtubule preparations did not copurify with microtubules in cycles of temperature-dependent assembly/disassembly but remained with the cold-insoluble intermediate filaments. These results provide an explanation for the low yields that have hampered attempts to purify microtubule-associated proteins, in particular HMWP, from cultured cells in the past. HMWP might have a dual role in the cell, serving not only as regulators of microtubule assembly but also as linker components between microtubules and intermediate filaments.