Identification of a 60‐kDa phosphoprotein that binds stored messenger RNA of Xenopus oocytes

Abstract

Rapidly labeled, polyadenylated RNA is contained in 3 distinct fractions isolated from homogenized amphibian oocytes; in ribonucleoprotein particles that are associated with a fibrillar matrix, the complexes sedimenting at > 1500 S [Svedberg units]; in ribonucleoprotein particles that sediment at 20-120 S and have the characteristics of stored (maternal) messenger ribonucleoprotein (mRNP) and in polyribosomes that sediment at 120-360 S. The RNA and protein components of the first 2 of these RNP fractions were compared. The polydenylated RNA extracted from the 2 RNP fractions differs in that the RNA from fibril-associated RNP contains a much higher content of repeat sequences than does the RNA from mRNP. In other words, the RNA from fibril-associated RNP is largely unprocessed and may constitute a premessenger state, which for convenience is referred to as premessenger RNP (pre-mRNP). RNA-binding experiments demonstrate that the polypeptide most tightly bound in pre-mRNP is a 54-kDA [kdalton] component (p54); the polypeptide most tightly bound in mRNP is a 60-kDa component (p60). Antibodies raised against p60 are used to show that this polypeptide is a common major component of pre-mRNP and mRNP and that is also located in oocyte nuclei. The state of p60 is modified between the premessenger and stored message levels: the polypeptide in mRNP is heavily phosphorylated whereas the equivalent polypeptide in pre-mRNP is completely unphosphorylated. The relative roles of the presence of repeat sequences and phosphorylation of mRNA-associated protein in blocking translation are discussed.