Separation of "estrogen-induced" protein from phosphoprotein phosphatase activity of immature rat uterus.

Abstract

Preparations of the "induced protein" which appears in the rat uterus within 40 min of estradiol administration have recently been reported to contain phosphoprotein phosphatase (phosphoprotein phosphohydrolase, EC 3.1.3.16) activity. We found that these two proteins distribute differently on ammonium sulfate fractionation of uterine cytosol. Preparative cellulose acetate electrophoresis afforded complete (>99.9%) separation of phosphoprotein phosphatase activity from the induced protein. The specific activity of phosphoprotein phosphatase in uterine cytosol was unchanged 1, 4, 12, or 24 hr after estradiol administration. These results are incompatible with the view that the induced protein mediates estrogen action by virtue of an inherent phosphoprotein phosphatase activity. Administration of estradiol-17fl to immature rats, or mature ovariectomized rats, results in an increased rate of synthesis of a uterine protein which has been designated "induced protein" (IP) by Notides and Gorski (1). Synthesis of estrogen-induced protein can be detected within 40 min of estradiol injection into rats (2) The proposal of Liu and Greengard (10), that increases in PPPase activity may modulate the response of toad bladder to the steroid hormone aldosterone, lends plausibility to the idea that an increase in PPPase activity may also be involved in the mechanism of estrogen action. In view of the conceptual importance of the identification of the function of IP and the practical advantage of having an enzymic activity for the quantitation of IP in our uterine cell-free synthetic system (11), we investigated the relationship between IP and PPPase. We studied the distribution of IP and PPPase during fractionation by ammonium sulfate precipitation and electrophoresis on cellulose acetate gels and found it possible to separate IP and PPPase activity.