title: Regulation of c-myc mRNA decay by translational pausing in a coding region instability determinant.
abstract: A 249-nucleotide coding region instability determinant (CRD) destabilizes c-myc mRNA. Previous experiments identified a CRD-binding protein (CRD-BP) that appears to protect the CRD from endonuclease cleavage. However, it was unclear why a CRD-BP is required to protect a well-translated mRNA whose coding region is covered with ribosomes. We hypothesized that translational pausing in the CRD generates a ribosome-deficient region downstream of the pause site, and this region is exposed to endonuclease attack unless it is shielded by the CRD-BP. Transfection and cell-free translation experiments reported here support this hypothesis. Ribosome pausing occurs within the c-myc CRD in tRNA-depleted reticulocyte translation reactions. The pause sites map to a rare arginine (CGA) codon and to an adjacent threonine (ACA) codon. Changing these codons to more common codons increases translational efficiency in vitro and increases mRNA abundance in transfected cells. These data suggest that c-myc mRNA is rapidly degraded unless it is (i) translated without pausing or (ii) protected by the CRD-BP when pausing occurs. Additional mapping experiments suggest that the CRD is bipartite, with several upstream translation pause sites and a downstream endonuclease cleavage site.
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
authors.forename: Ira
authors.initials: I
authors.lastname: Lemm
authors.forename: Jeff
authors.initials: J
authors.lastname: Ross
mesh_terms: D000818:Animals; D001120:Arginine / Q000235:genetics; D003062:Codon; D014176:Protein Biosynthesis; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics* / Q000378:metabolism; D020871:RNA Stability; D012333:RNA, Messenger / Q000378:metabolism*; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D012156:Reticulocytes; D012270:Ribosomes / Q000235:genetics
chemical_list: D003062:Codon; C498634:Igf2bp1 protein, rat; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012343:RNA, Transfer; D001120:Arginine