Expression of a gene for mouse eucaryotic elongation factor Tu during murine erythroleukemic cell differentiation.
The eucaryotic elongation factor Tu (eEF-Tu) is a single polypeptide with an approximate Mr of 53,000. During protein synthesis eEF-Tu promotes the binding of aminoacyl-tRNA to the ribosome. To study the expression of the gene(s) for this factor, a genomic clone was isolated that contains a mouse eEF-Tu gene. We screened a phage genomic library with a synthetic oligonucleotide probe complementary to a region of the Saccharomyces cerevisiae and Artemia sp. eEF-Tu genes which codes for an area that is highly conserved between both yeast and Artemia sp. eEF-Tu. From approximately 75,000 phage plaques we obtained five isolates with apparently identical inserts. All five clones contained a 3.8-kilobase EcoRI fragment that hybridized to additional oligonucleotide probes corresponding to different conserved regions of eEF-Tu. We sequenced the 5' end of one genomic clone and determined the length of the cloned fragment that was protected by eEF-Tu mRNA in S1 nuclease protection assays. A quantitative S1 nuclease protection assay was used to compare the relative steady-state levels of eEF-Tu mRNA in total mRNA in total RNA isolated from hexamethylene-bisacetamide-induced murine erythroleukemia cells. The results show a dramatic reduction in the steady-state level of eEF-Tu mRNA as differentiation proceeds. A similar reduction in transcription of eEF-Tu mRNA was observed in isolated nuclei. Finally, we examined the in vivo synthesis of eEF-Tu during differentiation and found that it declined in a manner parallel to the decline in the steady-state level of eEF-Tu mRNA. In addition, we have isolated and sequenced a cDNA clone for mouse eEF-Tu. The derived amino acid sequence is compared with sequences from other eucaryotes.
D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D003062:Codon; D005796:Genes; D004915:Leukemia, Erythroblastic, Acute / Q000235:genetics*; D007942:Leukemia, Experimental / Q000235:genetics* / Q000473:pathology; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010444:Peptide Elongation Factor Tu / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification; D014158:Transcription, Genetic
D003062:Codon; D012333:RNA, Messenger; D010444:Peptide Elongation Factor Tu