Fluorescence recovery after photobleaching studies on ATR kinase dynamics in live cells

Abstract

ATR (Ataxia-telangiectasia-mutated-and-Rad3-related) kinase is a key sensor of DNA damage in eukaryotic cells. Cells that express disease-causing mutations in lamin A, a major structural protein in the nucleus, are unable to repair damaged DNA due to degradation of ATR kinase. In order to understand why ATR kinase is predisposed to degradation in laminopathic cells, we have studied the dynamics of ATR kinase by fluorescence recovery after photobleaching experiments in normal HeLa cells and cells expressing lamin mutants. In normal cells subjected to UV irradiation, ATR kinase is localized in DNA repair foci, leading to reduction in its mobility. However, in cells expressing deleterious lamin mutants, ATR kinase mobility is reduced in the absence of formation of DNA repair foci. We propose that reduction of ATR kinase mobility in laminopathic cells may be due to aberrant interactions and mislocalization in the nucleus, and this may eventually lead to degradation of ATR kinase.