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Transcript
RACK1, A RECEPTOR FOR ACTIVATED C KINASE, LINKS METABOTROPIC GLUTAMATE RECEPTOR ACTIVATION WITH DENDRITIC TRANSLATIONAL CONTROL. F. Angenstein1*, J. E. Kacharmina2, B. Settlage3, S. T. Moran1, S.-C. Ling1, A. Klintsova1, D. Hunt3, J. Eberwine2, and W.T.Greenough1. 1Beckman Institute, University of Illinois, UrbanaIL 61801; 2Dept. of Pharmacology, University of Pennsylvania; 3Dept. of Chemistry, University of Virginia, Charlottesville VA 22901 Local protein synthesis is activated by glutamate in synaptoneurosomes (Weiler, Greenough PNAS, 90:7168, 1993). To search for transmitter receptor triggered mechanisms involved in translational control of dendritically localized mRNAs, we focussed on mRNPs that might be affected by second messenger systems. PolyAmRNAs were purified using oligo(d)T-cellulose. The associated proteins were then released and identified by mass spectrometry. Among the proteins that form the mRNP-complexes, we could detect RACK1 (receptor for activated C kinase). RACK1 appears to be linked via the polyA binding protein (PABP1) to mRNAs. Furthermore, in hippocampal slices a mGluR dependent binding of PKC 2 to this mRNA/PABP1/RACK1 complex could be observed. PKC2 is able to phosphorylate a set of mRNPs in vitro; three of them are phosphorylated under in situ conditions as well. To confirm that this mechanism – mGluR induced translocation of PKC to mRNA bound RACK1 – is in fact involved in dendritic translational control, we monitored the translation of a reporter mRNA construct into c-myc tagged protein in isolated dendrites. Stimulation of mGluR1,5 by 50 µM DHPG increased the translation of this construct. However, if this c-myc sequence-tagged mRNA was administered with a peptide that competed with the binding of PKC to RACK1, this receptor triggered enhancement in translation was impaired, an effect that was most clearly evident if the mRNA possessed a polyA tail. A random rearrangement of the same amino acids had no effect. Based on these findings we conclude that mGluR induced binding of activated PKC to mRNAs can trigger postsynaptic protein synthesis. (supported by FRAXA Research Foundation and NICHD)