Self-initiation of translation of mRNAs devoid of translational initiators in Escherichia coli.
Recent studies have shown that the canonical SD-anti-SD interaction is dispensable for the initiation of translation of certain mRNAs in Escherichia coli. In this study the cat and tetR genes were modified to either destroy complementarity to E. coli 16S RNA or completely delete their 5' non-translated regions. Thus a series of cat- and tetR-derived genes were constructed, cloned under a strong constitutive promoter and expressed in E. coli cells. The efficiency of expression was evaluated by the yield of CAT (for the cat gene) and cell viability in increasing concentrations of antibiotic (for both cat and tetR genes). The obtained results show that the mRNAs transcribed from both series of reporter genes (cat and tetR) were active in vivo. Their activity was preserved even in the cases when the length of their 5' non-translated leader sequences was reduced to one nucleotide for the cat gene and eight nucleotides for the tetR gene. The yield of protein obtained with the latter constructs was detectable and sufficient for bacteria to survive at 50-100 microg/ml chloramphenicol and 20 microg/ml tetracycline, respectively.
Kolev
V
V
Department of Gene Regulations, Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia.
Berzal-Herranz
A
A
Ivanov
I
I
D020121:5' Untranslated Regions / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002470:Cell Survival / Q000187:drug effects; D002701:Chloramphenicol / Q000494:pharmacology; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics; D010442:Peptide Chain Initiation, Translational; D010448:Peptide Initiation Factors / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D012336:RNA, Ribosomal, 16S / Q000235:genetics / Q000378:metabolism; D013752:Tetracycline / Q000494:pharmacology; D013753:Tetracycline Resistance / Q000235:genetics
D020121:5' Untranslated Regions; D010448:Peptide Initiation Factors; D012333:RNA, Messenger; D012336:RNA, Ribosomal, 16S; D002701:Chloramphenicol; D015500:Chloramphenicol O-Acetyltransferase; D013752:Tetracycline