A plant poly(A) polymerase requires a novel RNA-binding protein for activity.
We have purified a novel factor (PAP-III) that is a component of a multisubunit poly(A) polymerase from pea seedlings. This factor consists of one or more polypeptides with molecular masses of about 105 kDa and of a population of associated RNAs that can serve as substrates for polyadenylation. When these RNAs are separated from the 105-kDa polypeptides, polyadenylation becomes dependent upon exogenously added RNA. This RNA-dependent activity does not require the presence of a polyadenylation signal in the substrate, indicating that the activity under study is a nonspecific polyadenylation activity. One or more of the 105-kDa polypeptides could be cross-linked to the products of polyadenylation labeled with [alpha-32P]ATP and to exogenously added labeled RNAs. Cross-linking of the 105-kDa polypeptides to the products of polyadenylation was not affected by the presence of exogenously added competitors, whereas cross-linking to exogenous RNAs was diminished by excesses of RNA homopolymers. Exogenous RNAs could be polyadenylated by the combination of PAP-I + PAP-III, and this activity was diminished if the binding of the exogenous RNAs to PAP-III was prevented. We conclude from these studies that PAP-III is an RNA binding protein, that polyadenylation by the poly(A) polymerase occurs while the substrate RNAs are associated with this protein, and that the pea poly(A) polymerase can only polyadenylate those RNAs that are associated with PAP-III.
Li
Q S
QS
Plant Physiology/Biochemistry/Molecular Biology Program, Department of Agronomy, University of Kentucky, Lexington, Kentucky 40546-0091, USA.
Gupta
J D
JD
Hunt
A G
AG
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D008970:Molecular Weight; D018532:Pisum sativum; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D011062:Polynucleotide Adenylyltransferase / Q000378:metabolism*; D012313:RNA / Q000378:metabolism*; D018749:RNA, Plant / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid; D012261:Ribonucleoproteins / Q000378:metabolism
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D010940:Plant Proteins; D018749:RNA, Plant; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D012313:RNA; D011062:Polynucleotide Adenylyltransferase