Turnover and processing of poly(A) in full-grown oocytes and during progesterone-induced oocyte maturation in Xenopus laevis

Abstract

Changes in the amount and the size of poly(A) sequences during progesterone-induced oocyte maturation were determined by [3H]poly(U) hybridization and by labeling with microinjected [α-32P]ATP and [3H]ATP. Following exposure to progesterone total poly(A) length increases by 10–20 nucleotides, largely due to extension of preexisting poly(A) although there is an increase of 5 to 10% in the number of poly(A) sequences. Immediately following germinal vesicle breakdown (GVBD), poly(A) is degraded with first-order kinetics (t1/2 = 5.7 hr) until about 10 hr after GVBD. This degradation results in the loss of 35–50% of the poly(A) sequences but the size distribution of the remaining poly(A) is unchanged. These changes occur only in oocytes which undergo GVBD, but all the changes in poly(A) content also occur in oocytes enucleated before exposure to progesterone. The poly(A) of ovulated eggs is similar in amount and size to that ofin vitro matured oocytes. The incorporation of [3H]ATP into poly(A) confirms that most of the poly(A) synthesis in both maturing and control oocytes is due to chain extension and turnover rather than tode novo synthesis. The amount of newly synthesized poly(A) is sufficient to allow for the net increase in larger poly(A) sequences by chain extension but not byde novo synthesis. Newly synthesized poly(A) has a size distribution similar to that of steady-state poly(A), and no precursor-like32P-poly(A) molecules are detected. The lengths of poly(A) sequences labeled with [3H]ATP, determined by [3H]AMP:[3H]adenosine ratios, are smaller than those indicated by their electrophoretic mobilities in both control and maturing oocytes. It is argued that the size and stability of oocyte poly(A) sequences are regulated by balanced synthetic and degradative activities of cytoplasmic poly(A) polymerase which result in turnover of the 3′-terminal portion of the poly(A). During progesterone-induced maturation there is a relative increase in poly(A) synthesis until the time of GVBD, when a different degradative activity results in destruction of complete poly(A) sequences. These changes must be cytoplasmically programmed because they occur in the absence of the nucleus.