Heat-induced nuclear accumulation of hsc70 proteins is regulated by phosphorylation and inhibited in confluent cells

Stress affects the general organization of cells and, in particular, the subcellular localization of molecules. Proteins of the hsp70/hsc70 family relocate to nuclei in response to heat shock, when classical nuclear protein import is inhibited. We have now further characterized the effect of stress on hsc70 protein localization in HeLa cells. Heat-induced nuclear concentration of hsc70 proteins depends on cell density, and low-density cultures efficiently imported hsc70 proteins into nuclei when exposed to heat. However, high-density cultures failed to accumulate hsc70 proteins in nuclei during heat shock. In low-density cultures, heat-induced hsc70 nuclear accumulation was insensitive to inhibitors of tyrosine kinases, Ser/Thr protein kinases, and rapamycin, which indicates that activation of mitogen-activated protein kinase family members or p70 S6 kinase is not required for this process. In contrast, inhibitors of Ser/Thr protein phosphatase and Tyr protein phosphatase abolished the nuclear concentration of hsc70 proteins. Likewise, inhibitors of protein phosphatases affected classical nuclear protein import in unstressed cells, but these transport pathways differed drastically in their sensitivities toward inhibitors. Although protein phosphorylation negatively regulates hsc70 nuclear accumulation in response to heat, protein kinase inhibitors were unable to overcome the block of hsc70 nuclear accumulation in high-density cultures, indicating that additional components control hsc70 nuclear transport.