RRM
2006
Rustbelt RNA Meeting
RRM
|
|
|
|
|
Talk abstracts
Abstract:
PMR1 is an mRNA endonuclease that catalyzes the selective decay of a discrete subset of mRNAs. In contrast to the generalized pathway of mRNA decay, which acts on non-translating mRNA, PMR1 acts by forming a complex with its translating substrate mRNA, where it initiates decay by endonuclease cleavage within the body of the mRNA. We previously showed that PMR1 is phosphorylated at Y650, and tyrosine phosphorylation at this site is required for targeting to polysomes and mRNA decay. Our current work identifies c-Src as the responsible kinase. Most tyrosine kinases form a complex with their substrate, and in this context can phosphorylate both the target protein and the kinase itself. In vitro kinase assays of PMR1-TAP complexes recovered from transfected cells identified 2 tyrosine-phosphorylated proteins, one of which disappears when recovery was performed with the Y650F mutant form of the protein. The remaining ~60 kDa protein was similar in size to c-Src, and its activity was blocked by the Src inhibitor PP2. Reciprocal co-ip experiments confirmed Src and PMR1 are recovered in the same complex, and more Src is recovered with the Y650F mutant than wild-type protein. In addition recombinant Src phosphorylates both PMR1 and a peptide with the tyrosine phosphorylation site in vitro. PMR1 does not target to polysomes in Src-deficient cells or cells expressing either kinase-inactive Src or a dominant negative form of Src. However, virtually all of the PMR1 targets to polysomes in cells expressing constitutively active Src. Similarly, Src activates PMR1-mediated mRNA decay whereas dominant-negative Src or catalytically-inactive Src do not. These data provide the first link between mRNA decay and signal transduction by tyrosine kinases.
Keywords: polysomal ribonuclease 1, c-Src kinase, mRNA decay
