Inhibition of myristoylated alanine-rich C kinase substrate phosphorylation by UV light and methyl methanesulfonate

Abstract

The possible involvement of ultraviolet light and methyl methanesulfonate in the intracellular signal transduction cascade was investigated. When NIH 3T3 cells were exposed to 254 nm short wavelength ultraviolet light, a typical physical DNA-damaging agent, there was a temporal cessation of DNA synthesis and phosphorylation of myristoylated alanine-rich C kinase substrate (MARCKS) protein was inhibited. This inhibition of MARCKS phosphorylation was correlated with decreased protein kinase C activity in the membrane fraction of ultraviolet light-damaged cells. The inositol-1,4,5-trisphosphate contents measured, by the competitive binding assay using bovine adrenal binding protein, showed 80 % reduction in the fibroblasts treated with 15 J/㎡ of ultraviolet light. The intracellular diacylgylcerol concentration was also markedly reduced in ultraviolet light-damaged cells. The results suggest that ultraviolet light causes acute reductions of inositol trisphosphate and diacylglycerol contents in cells along with the decrease in membrane protein kinase C activity, which leads to the inhibition of phosphorylation of MARCKS. The effect of a typical chemical mutagen, methyl methanesulfonate on the phosphorylation of MARCKS protein was also investigated. Methyl methanesulfonate inhibited membrane bound protein kinase C activity and the phosphorylation of myristoylated alanine-rich C kinase substrate. Treatment of the chemical also lowered the cellular amount of second messengers of inositol-1,4,5-trisphosphate and diacylglycerol. Data suggest that the perturbed production of second messenger molecules by methyl methanesulfonate is resulted from the decreased phosphorylation of phospholipase C of which the activity is influenced by the states of its phosphorylation. Therefore it was concluded that ultraviolet light and methyl methanesulfonate intervene in the signal cascade in NIH 3T3 cells by inhibiting the production of second messenger molecule, which leads to the inact...