When vertebrate cells are exposed to sodium arsenite, they respond by increased synthesis of a small number of polypeptides similar to the heat shock proteins, which are induced by exposure to 40-45°C. The 70,000 dalton inducible protein (hsp70) is the most commonly induced species and this protein is methylated at both lysyl and arginyl residues. In chicken fibroblasts, the hsp70 is composed of two major distinct isoelectric variants, as well as several minor components. The more acidic one (hsp70A; pI 5.6) is highly conserved and found in all vertebrate tissues and cultured cells examined, while the more basic one (hsp70B; pI 6.0) at present has only been found in avian cells.
The hsp70 is a prominent cytoplasmic constituent under normal growth conditions. In chicken fibroblasts, the induction by arsenite not only increases the synthesis of hsp70 but also results in an accumulation of this protein. In contrast, the induction in 3T3 and SR-RSV 3T3 cells results in increased synthesis, but no accumulation of this polypeptide.
In chicken cells, ε-N-trimethyl-lysine has been identified as the major component of methyl-lysine species in hsp70, but the contribution of ε-N-monomethyl-lysine and ε-N-dimethyl-lysine is evident. The methyl-arginine of hsp70 is exclusively NG-monomethyl-arginine; these methylations appear to be stoichiometric. Furthermore, these methylations can be modulated by arsenite. In particular, in hsp70A the amount of ε-N-trimethyl-lysine decreases and the ε-N-dimethyl-lysine significantly increases, while in hsp70B the quantity of NG-monomethyl-arginine is reduce fivefold in the presence of sodium arsenite. In 3T3 and SR-RSV 3T3 cells, ε-N-trimethyl-lysine appears to be the only methylated lysine species; both NG-monomethyl-arginine and NG,NG-dimethyl-arginine have been identified as the methylated arginyl residues. Similar to the homologous polypeptide in chicken fibroblasts, the level of arginyl methylation of hsp70 of 3T3 cells can be reduced in the presence of arsenite. This arsenite-induced reduction in arginyl methylation of hsp70 appears to be restricted to the polypeptides synthesized during the arsenite incubation. However, the arginyl methylation level of hsp70 of SR-RSV 3T3 is constitutively lower than their untransformed counterpart (3T3 cells), and the methylation cannot be reduced further by arsenite. In addition, the level of lysyl methylation of hsp70 remains the same after arsenite treatment and transformation. Since the basic amino acid methylation in total cellular proteins remains unchanged after arsenite incubation and Rous sarcoma virus transformation, the reduction in arginyl methylation of hsp70 in chicken fibroblasts and 3T3 cells by these treatments appears to be specific.
