A polymer-dependent increase in phosphorylation of β-tubulin accompanies differentiation of a mouse neuroblastoma cell line

Abstract

We have examined the phosphorylation of cellular microtubule proteins during differentiation and neurite outgrowth in N115 mouse neuroblastoma cells. N115 differentiation, induced by serum withdrawal, is accompanied by a fourfold increase in phosphorylation of a 54,000-mol-wt protein identified as a specific isoform of β-tubulin by SDS PAGE, twodimensional isoelectric focusing/SDS PAGE, and immunoprecipitation with a specific monoclonal antiserum. Isoelectric focusing/SDS PAGE of [35S]methionine-labeled cell extracts revealed that the phosphorylated isoform of β-tubulin, termed β2, is one of three isoforms detected in differentiated Nl15 cells, and is diminished in amounts in the undifferentiated cells. Taxol, a drug which promotes microtubule assembly, stimulates phosphorylation of β-tubulin in both differentiated and undifferentiated Nl15 cells. In contrast, treatment of differentiated cells with either colcemid or nocodazole causes a rapid decrease in β-tubulin phosphorylation. Thus, the phosphorylation of β-tubulin in N115 cells is coupled to the levels of cellular microtubules. The observed increase in β-tubulin phosphorylation during differentiation then reflects developmental regulation of microtubule assembly during neurite outgrowth, rather than developmental regulation of a tubulin kinase activity. © 1985, Rockefeller University Press., All rights reserved.