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Unmasking mRNA in clam oocytes: role of phosphorylation of a 3' UTR masking element-binding protein at fertilization.

Abstract：

:During meiotic maturation or after fertilization of invertebrate and vertebrate oocytes, many of the quiescent stored mRNAs are recruited into polysomes. In the clam, Spisula solidissima, such masked messages include the abundant mRNAs encoding cyclin A and the small subunit of ribonucleotide reductase. We have previously shown that mRNA-specific unmasking of these two messages can be achieved in vitro, in oocyte cell-free extracts, by the addition of antisense RNAs corresponding to a fairly short (130-140 nucleotides) segment in their cognate 3' untranslated regions. We postulated that the antisense RNAs prevented the binding of a masking repressor protein (Standart et al., 1990). Here we report UV-crosslinking and gel retardation studies which show that the masking portions of the translationally regulated mRNAs bind an oocyte protein of 82 kDa (p82), which is phosphorylated after fertilization. This modification was accompanied by altered RNP complex formation in gel retardation assays. These changes presumably reflect the activation of translation of the masked mRNAs. The role of p82 phosphorylation in maternal mRNA unmasking was assessed in a novel in vitro activation system developed from clam oocytes, based upon the natural rise in pH which accompanies fertilization. Concomitant with mRNA unmasking, several kinases, including cdc2 and MAP kinases were activated in this system, as was p82 phosphorylation. Inhibitors of serine/threonine kinases, including 6-DMAP, staurosporine, and H7 inhibited p82 phosphorylation, whereas inhibitors of tyrosine kinases, protein kinase C, cAMP-dependent protein kinase, and p70s6k did not prevent this modification. A specific inhibitor of cdc2 kinase, p27Kip1, prevented p82 phosphorylation and translational activation, strongly suggesting that p82 modification is required for unmasking.

journal_name

Dev Biol

journal_title

Developmental biology

authors

Walker J,Dale M,Standart N

doi

10.1006/dbio.1996.0024

subject

Has Abstract

pub_date

1996-01-10 00:00:00

pages

292-305

issue

eissn

0012-1606

issn

1095-564X

pii

S0012-1606(96)90024-X

journal_volume

173

pub_type

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Unmasking mRNA in clam oocytes: role of phosphorylation of a 3' UTR masking element-binding protein at fertilization.