Overexpression of initiation factor eIF-4E does not relieve the translational repression of ribosomal protein mRNAs in quiescent cells.
Translation of ribosomal protein (rp) mRNA is selectively repressed in mouse erythroleukemia (MEL) cells, which cease to proliferate upon differentiation, and in NIH 3T3 cells, for which growth is arrested by either serum starvation, contact inhibition, or treatment with the DNA polymerase inhibitor, aphidicolin. The efficiency of translation of rp mRNAs correlates with the expression of the gene encoding the cap binding protein, eIF-4E, as indicated by the fact that the abundance of the corresponding mRNA and protein also fluctuates in a growth-dependent manner. To examine the hypothesis that eIF-4E plays a role in regulation of the translation efficiency of rp mRNAs, we utilized an NIH 3T3-derived eIF-4E-overexpressing cell line. These cells overproduce eIF-4E to the extent that even under conditions of growth arrest, the abundance of the respective protein in its active (phosphorylated) form is higher than that found in exponentially growing NIH 3T3 cells. Nevertheless, this surplus amount of eIF-4E does not prevent the translational repression of rp mRNAs when the growth of these cells is arrested by blocking DNA synthesis with aphidicolin or hydroxyurea. In complementary experiments we used an in vitro translation system to compare the competitive potential of mRNAs, containing the translational cis-regulatory element (5' terminal oligopyrimidne tract) and mRNAs lacking such a motif, for the cap binding protein. Our results demonstrate that both types of mRNAs, regardless of their translational response to growth arrest, exhibit similar sensitivity to the cap analogue m7G(5')ppp(5')G. It appears, therefore, that the presence of the regulatory sequence at the 5' terminus of rp mRNAs does not lessen its competitive potential for the cap binding protein and that the growth-dependent decrease in the activity of eIF-4E does not play a key role in the repression of translation of rp mRNAs.
Shama
S
S
Department of Developmental Biochemistry, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Avni
D
D
Frederickson
R M
RM
Sonenberg
N
N
Meyuhas
O
O
D016475:3T3 Cells; D000818:Animals; D016590:Aphidicolin / Q000494:pharmacology; D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology; D002455:Cell Division / Q000235:genetics; D003432:Cross-Linking Reagents / Q000378:metabolism; D015226:Dinucleoside Phosphates / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D039561:Eukaryotic Initiation Factor-4E; D005786:Gene Expression Regulation; D006131:Growth Inhibitors / Q000235:genetics / Q000378:metabolism; D006918:Hydroxyurea / Q000494:pharmacology; D007525:Isoelectric Focusing; D004915:Leukemia, Erythroblastic, Acute; D051379:Mice; D010448:Peptide Initiation Factors / Q000096:biosynthesis* / Q000235:genetics; D010766:Phosphorylation; D014176:Protein Biosynthesis / Q000502:physiology*; D012314:RNA Cap Analogs / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics; D011817:Rabbits; D012097:Repressor Proteins / Q000235:genetics; D012269:Ribosomal Proteins / Q000235:genetics*; D014407:Tumor Cells, Cultured
D003432:Cross-Linking Reagents; D015226:Dinucleoside Phosphates; D039561:Eukaryotic Initiation Factor-4E; D006131:Growth Inhibitors; D010448:Peptide Initiation Factors; D012314:RNA Cap Analogs; D012333:RNA, Messenger; D012097:Repressor Proteins; D012269:Ribosomal Proteins; D016590:Aphidicolin; C062229:7-methyl-diguanosine triphosphate; D006918:Hydroxyurea