Effect of eukaryotic initiation factor 4F on AUG selection in a bicistronic mRNA.
Artificial bicistronic mRNAs based on rabbit beta-globin and bacterial chloramphenicol acetyltransferase protein-coding sequences were tested for translation activity in a mouse astrocytoma cell-free extract. This cell extract exhibited an apparent preference for 5'-distal or internal initiation over 5'-proximal ("first AUG") initiation. 5'-Distal initiation appeared to be 5'-cap independent, suggesting that nonstandard initiation was responsible. This conclusion was based on a lack of inhibition of internal initiation by added cap analog and insensitivity of internal initiation to the presence or absence of a 5'-cap structure. Exogenous reticulocyte initiation factors were tested for effect on 5'-proximal initiation. The only factor with a significant effect was found to be eukaryotic initiation factor 4F, or the cap-binding protein. Addition of this factor promoted 5'-end initiation as evident by a general increase in 5'-proximal open reading frame (ORF) product relative to 5'-distal ORF product. The relative expression of 5'-proximal to 5'-distal ORFs in bicistronic or multicistronic mRNAs may very well be dependent on activity levels of eukaryotic initiation factor 4F and possibly other mRNA-dependent initiation factors.
Tahara
S M
SM
Department of Microbiology, University of Southern California School of Medicine, Los Angeles 90033-1054.
Dietlin
T A
TA
Dever
T E
TE
Merrick
W C
WC
Worrilow
L M
LM
D000818:Animals; D001345:Autoradiography; D001483:Base Sequence; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D039562:Eukaryotic Initiation Factor-4F; D051379:Mice; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010448:Peptide Initiation Factors / Q000494:pharmacology*; D010957:Plasmids; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D014158:Transcription, Genetic
D039562:Eukaryotic Initiation Factor-4F; D010448:Peptide Initiation Factors; D012333:RNA, Messenger