LEVEL OF HGCL2-MEDIATED PHOSPHORYLATION OF INTRACELLULAR PROTEINS DETERMINES DEATH OF THYMIC T-LYMPHOCYTES WITH OR WITHOUT DNA FRAGMENTATION

Exposure to Hg2+ at a wide range of concentrations(approximately 1-100 mu M) more or less caused the death of murine thymic T-lymphocytes, a nd exposure to 1 mu M but not 10 mu M (or more) of Hg2+ induced DNA fr agmentation. Exposure of cells to Hg2+ caused phosphorylation of multi ple cellular proteins at the tyrosine residue in a concentration-depen dent manner. We found that not only the DNA fragmentation induced by 1 mu M Hg2+ but also the cell death bypassing DNA fragmentation caused by 10 mu M or more Hg2+ was partly inhibited by protein kinase inhibit ors such as staurosporine and herbimycin A. This result suggested the involvement of. protein phosphorylation-linked signal in the mechanism of the Hg2+-mediated cell death with or without DNA fragmentation. An alysis of proteins by both one- and two-dimensional electrophoresis an d immunoblot showed that a 52-kDa Shc protein was heavily phosphorylat ed by an early signal delivered by a high concentration of Hg2+, which also phosphorylated extracellular signal-regulated kinase 1 (ERK1; p4 4) and ERK2 (p42) of the mitogen-activated protein kinase (MAPK) famil y in a concentration- and time-dependent manner. The c-Jun amino termi nal kinase (p54), which is a distant relative of the MAPK family was a lso phosphorylated by the treatment with Hg2+. This eventually formed the signaling cascade that ended with a nuclear target by phosphorylat ing c-jun at the serine 73. This phosphorylation of c-jun was inhibite d by staurosporine. These results suggest that a high level of Hg2+-me diated protein phosphorylation-linked signal induces rapid cell death bypassing DNA fragmentation, whereas a lower level induces cell death accompanying DNA fragmentation. This conclusion in turn implies that D NA fragmentation is not always a prerequisite for the signal transduct ion-dependent cell death of T-lymphocytes. J. Cell. Biochem. 71 :243-2 53, 1998. (C) 1998 Wiley-Liss, Inc.