Overexpression of wild type p70 S6 kinase interferes with cytokinesis

Abstract

p70 S6 kinase (p70^s6k) is a serine/threonine kinase which is activated through an unidentified pathway by mitogenic stimuli. Here we demonstrate that stable- and transient-overexpression of wild type (WT)-p70^s6k results in perturbation of cytokinesis in NIH3T3 cells. Cells overexpressing WT-p70^s6k demonstrated a slow growth rate and lower viability, whereas cells overexpressing a mutant T229A-p70^s6k had a similar growth rate and viability as those of parental NIH3T3 cells. Moreover, WT-p70^s6k cells demonstrated several abnormalities in cellular morphology; both the cytoplasm and the nuclei were large and a proportion of the cells was multi-nucleated. Flow cytometric analysis of propidium iodide-stained cells demonstrated that about 50% of the cells had 4N or greater content of DNA. When cell division in WT-p70^s6k cells was monitored by time-lapse video microscopy, cytokines was often incomplete after mitosis, producing binucleated cells. Addition of rapamycin to WT-p70^s6k cells, which inactivates endogenous and transfected p70^s6k, failed to reverse any of the morphological abnormalities. These indicate that overexpression of p70^s6k but not increases in activity of p70^s6k, is responsible for the phenotype. A molecule which interacts with p70^s6k may be involed in this regulation of cytokinesis.