Induction of protein methylases on immune responses in splenic lymphocytes froM balb/c MICE

Abstract

The induction of protein methylase I preceded that of protein methylase III during in vivo immunizing period by SRBC and in vitro mitotic stimulation of splenic lymphocytes by lipopolysaccharide (LPS) and concanavalin A (Con A). The cellular protein methylation of arginyl and lysyl residues increased in proportion to the enzymes`` activities. The methylation of these residues, and the proliferative, the polyclonal antibody and the T-independent antibody responses were markedly inhibited by sinefungin (SF) and 5``-deoxy-5``-S-isobutyl-adenosine (SIBA), two potent inhibitors of the enzymes. the Ki values of SF for the enzymes in vitro were lower than those of SIBA, however the cellular responses of the protein N-methylations and the immune responses (except T-independent response) were inhibited more sensitively by SIBA. Regardless of when added, SIBA immediately showed its effects on LPS-induced immune responses. The effects of SF were no longer observed when added later than the first 12 hrs in the 48 hrs experiment, however those of SF did not appear until 24 hrs after addition. These results showed that the rate-limiting factor to show its toxicity might be the step of transport into the cell. Protein methylase I and III were assayed at indicated time during incubation of splenic lymphocytes with SF and SIBA. The enzymes activities in all tested groups were significantly inhibited by SIBA, but the enzymes activities were not inhibited by SF in cells harvested within 24 hrs after incubation in spite of higher inhibitory effect for the enzymes in vitro. This confirmed the hypothesis. Therefore, the magnitude of inhibition by SF varied in proportion to the duration of incubation. That is, the effects of SF were lower than those of SIBA before 72 hrs of incubation, and the effects of SF increased so rapidly thereafter that LPS-induced immune responses were more potently blocked by SF. In this condition, it showed a correspondence between Ki value of SF and SIBA...