Expression and heat-responsive regulation of a TFIIB homologue from the archaeon Haloferax volcanii.
Multiple divergent genes encoding the eukaryal-like TFIIB (TFB) transcription initiation factor have been identified in the archaeon Haloferax volcanii. Expression of one of these TFB-encoding genes, referred to here as tfb2, was induced specifically in response to heat shock at the transcription level. A time course for tfb2 induction demonstrated that mRNA levels increased as much as eightfold after 15 min at 60 degrees C. A transcription fusion of the tfb2 promoter region with a stable RNA reporter gene confirmed the heat responsiveness of the tfb2 core promoter, and immunoblot analysis using antibodies generated against a recombinant His-tagged TFB2 showed that the protein levels of one TFB increased slightly in response to elevated temperatures. An archaeal consensus TATA element (5'-TTTATA-3') was located 110 bp upstream of the translation start site and appeared to be used for both basal and heat shock-induced expression. The long DNA leader region (79 bp) preceding the predicted AUG translation start codon for TFB2 contained a T-rich sequence element located 22 bp downstream of the transcription start site. Using an in vivo transcription termination assay, we demonstrated that this T-rich element can function as a sequence-dependent transcription terminator, which may serve to downregulate expression of the tfb2 gene under both non-heat shock and heat shock conditions.
Thompson
D K
DK
Department of Microbiology, The Ohio State University, 484 West 12th Avenue, Columbus, OH 43210, USA.
Palmer
J R
JR
Daniels
C J
CJ
D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D019848:Gene Expression Regulation, Archaeal; D017930:Genes, Reporter; D019622:Haloferax volcanii / Q000235:genetics* / Q000502:physiology; D018869:Heat-Shock Response; D006358:Hot Temperature; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D035581:Transcription Factor TFIIB; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
D019843:Archaeal Proteins; D011994:Recombinant Proteins; D035581:Transcription Factor TFIIB; D014157:Transcription Factors