Peroxynitrite-induced tyrosine nitration and inhibition of protein kinase C.

Protein kinase C (PKC) is an important intracellular signaling molecule whose activity is essential for a number of aspects of neuronal function including synaptic plasticity. We investigated the regulation of PKC activity by reactive nitrogen species in order to examine whether such species regulate PKC in neurons. Neither autonomous nor cofactor-dependent PKC activity was altered when either hippocampal homogenates or rat brain purified PKC were incubated briefly with three different nitric oxide donor compounds. However, brief incubation of either hippocampal homogenates or purified PKC with peroxynitrite (ONOO(-)) inhibited cofactor-dependent PKC activity in a manner that correlated with the nitration of tyrosine residues on PKC, suggesting that this modification was responsible for the inhibition of PKC. Consistent with this idea, reducing agents had no effect on the inhibition of PKC activity caused by ONOO(-). Because there are numerous PKC isoforms that differ in the composition of the regulatory domain, we studied the effect of ONOO(-) on various PKC isoforms. ONOO(-) inhibited the cofactor-dependent activity of the alpha, betaII, epsilon, and zeta isoforms, indicating that inhibition of enzymatic activity by ONOO(-) was not PKC isoform-specific. We also were able to isolate nitrated PKCalpha and PKCbetaII from ONOO(-)-treated hippocampal homogenates via immunoprecipitation. Collectively, our findings support the hypothesis that ONOO(-) inhibits PKC activity via tyrosine nitration in neurons.

Lack of effect on cyclic GMP content of cells treated with mycophenolic acid.

Mycophenolic acid, an oncolytic agent and a known inhibitor of guanine ribonucleotide synthesis, has proven to be an effective drug against psoriasis. With reports of greater guantities of c-GMP in psoriatic tissues than in normal tissue, and with the correlation of c-GMP content of cells to proliferation, the effect of mycophenolic acid on cellular c-GMP was investigated. When HeLa, green monkey BSC-1, and mouse L-cells were treated with inhibitory concentrations of mycophenolic acid, no decrease in c-GMP was observed from that of untreated cells. Though mycophenolic acid inhibits guanine ribonucleotide synthesis, this inhibition does not extend to c-GMP synthesis. The inhibition of proliferation of cells by mycophenolic acid then does not include the inhibition of synthesis of c-GMP, but apparently resides solely in limiting the guanylate necessary for nucleic acid synthesis.