Identification of a 60-kDa phosphoprotein that binds stored messenger RNA of Xenopus oocytes.
Rapidly labelled, polyadenylated RNA is contained in three distinct fractions isolated from homogenized amphibian oocytes: (a) in ribonucleoprotein particles that are associated with a fibrillar matrix, the complexes sedimenting at greater than 1500S; (b) in ribonucleoprotein particles that sediment at 20-120S and have the characteristics of stored (maternal) messenger ribonucleoprotein (mRNP) and (c) in polyribosomes that sediment at 120-360S. We have compared the RNA and protein components of the first two of these RNP fractions. The polyadenylated RNA extracted from the two 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 component (p54), whereas 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 it is also located in oocyte nuclei. However 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.
A L
AL
Dearsly
R M
RM
Johnson
P
P
Barrett
J
J
Sommerville
D000818:Animals; D002499:Centrifugation, Density Gradient; D011232:Chemical Precipitation; D004527:Egg Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D007120:Immunochemistry; D008970:Molecular Weight; D009865:Oocytes / Q000378:metabolism*; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D011061:Poly A / Q000378:metabolism; D011485:Protein Binding; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000302:isolation & purification / Q000378:metabolism*; D016601:RNA-Binding Proteins; D012091:Repetitive Sequences, Nucleic Acid; D012261:Ribonucleoproteins / Q000302:isolation & purification / Q000378:metabolism*; D014982:Xenopus laevis
D004527:Egg Proteins; D010750:Phosphoproteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; C046091:phosphoprotein p60 (Xenopus oocyte); D011061:Poly A; D012313:RNA