Expression of antisense RNA against initiation factor eIF-4E mRNA in HeLa cells results in lengthened cell division times, diminished translation rates, and reduced levels of both eIF-4E and the p220 component of eIF-4F.
HeLa cells were transformed to express antisense RNA against initiation factor eIF-4E mRNA from an inducible promoter. In the absence of inducer, these cells (AS cells) were morphologically similar to control cells but grew four- to sevenfold more slowly. Induction of antisense RNA production was lethal. Both eIF-4E mRNA and protein levels were reduced in proportion to the degree of antisense RNA expression, as were the rates of protein synthesis in vivo and in vitro. Polysomes were disaggregated with a concomitant increase in ribosomal subunits. Translation in vitro was restored by addition of the initiation factor complex eIF-4F but not by eIF-4E alone. Immunological analysis revealed that the p220 component of eIF-4F was decreased in extracts of AS cells and undetectable in AS cells treated with inducer, suggesting that p220 and eIF-4E levels are coordinately regulated. eIF-4A, another component of eIF-4F, was unaltered.
De Benedetti
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A
Department of Biochemistry, University of Kentucky Medical Center, Lexington 40536.
Joshi-Barve
S
S
Rinker-Schaeffer
C
C
Rhoads
R E
RE
D000199:Actins / Q000235:genetics; D001483:Base Sequence; D015152:Blotting, Northern; D002455:Cell Division; D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense; D010448:Peptide Initiation Factors / Q000235:genetics*; D010641:Phenotype; D016133:Polymerase Chain Reaction; D011132:Polyribosomes / Q000378:metabolism; D016372:RNA, Antisense / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D014162:Transfection
D000199:Actins; D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D046911:Macromolecular Substances; D016376:Oligonucleotides, Antisense; D010448:Peptide Initiation Factors; D016372:RNA, Antisense; D012333:RNA, Messenger