Effect of vanadate on protein phosphorylation and on acid phosphatase activity in the canine prostate

Abstract

To evaluate the possible role of intracellular phosphatases in the local regulation of prostatic functions, the effect of sodium orthovanadate (VO 4), an inhibitor of phosphotyrosyl protein phosphatases, was studied on both protein phosphorylation and acid phosphatase activity. Secretory and non-secretory epithelial cells were isolated from normal and metaplastic prostates and incubated with [ 32P]phosphate in the presence and in the absence of VO 4; the phosphoproteins were separated by electrophoresis and the gels were either directly submitted to autoradiography or after an alkali treatment to reveal those proteins enriched in phosphotyrosine. Prior to alkali treatment, several phosphoproteins were evidenced and in less than half of the cell preparations a slight increase in labeling intensity under vanadate (<75%) was observed in two phosphoproteins, p57 and p44. After alkali treatment: (1) the effect of VO 4 on p57 remained in the order of 44–45% and it was restricted to less than half of non-secretory cell preparations; (2) its effect on p44 was intensified (134–207%) and observed in all cell types and in more than 80% of all preparations; and (3) in half of non-secretory cell preparations from metaplastic glands, an effect of VO 4 on p35 (127%) became evident. In all instances, with normal and/or metaplastic prostates, protein phosphorylation activity, either total or alkali-resistant and in the presence or in the absence of VO 4, was always higher in non-secretory epithelial cells as compared to secretory cells. In normal glands, prostatic acid phosphatase (PAP) and phosphotyrosine (p-Tyr) phosphatase activities were elevated and higher in secretory cells compared to non-secretory cells; on the contrary, these activities were very low in secretory and non-secretory cells from metaplastic glands. The acid phosphatase activity on phosphoserine and phosphothreonine was negligible while phosphotyrosine was hydrolysed to the same extent as p-nitrophenyl phosphate in all cell types. The PAP and p-Tyr phosphatase activities were strongly inhibited by VO 4 ( > 85%) in all cell types. Tartrate was a more potent inhibitor of both PAP and p-Tyr phosphatase activities measured with cells from normal glands ( > 80%) as compared to those of cells from metaplastic prostates (35–60%). Thus, vanadate inhibits PAP and p-Tyr phosphatases in prostatic cells and enhances the level of specific phosphoproteins, namely among alkali-resistant phosphoproteins. It is proposed that intracellular phosphatases, and possibly PAP, regulate protein phosphorylation in the prostate and therefore participate in the local modulation of prostatic growth.