Promoter recognition in archaea is mediated by transcription factors: identification of transcription factor aTFB fromMethanococcus thermolithotrophicusas archaeal TATA-binding protein

Abstract

At least two transcription factors, aTFB and aTFA, are required for accurate and faithful in vitro transcription of homologous templates in cell-free extracts from the methanogenic Archaeon Methanococcus thermolithotrophicus. We have recently shown that the function of aTFB can be replaced by eucaryal TATA-binding proteins. Here we demonstrate using template commitment experiments that promoter recognition in an Archaeon is mediated by transcription factors. The archaeal TATA box was identified as recognition site for binding of aTFB by gel shift analyses. aTFB binds also to the TATA box of adenovirus 2 major late promoter suggesting homology of eucaryal and archaeal TATA boxes. Our analyses provide evidence for a common molecular mechanism of transcription initiation by eucaryal RNA polymerases and archaeal RNA polymerase. They indicate also an evolutionary homology for aTFB and TBP.

Two Transcription Factors Related with the Eucaryal Transcription Factors TATA-binding Protein and Transcription Factor IIB Direct Promoter Recognition by an Archaeal RNA Polymerase

“…Methanococcus aTFB, Pc-TBP, and Sulfolobus-TBP were shown to bind to DNA fragments harboring an archaeal 2 C. Hethke and M. Thomm, unpublished data. TATA box in gel shift assays (7,13,18,19), and the TATA box of a Pyrococcus promoter was identified in this study as the binding site for Pc-TBP by DNase I protection studies (Fig. 3).…”

“…Highly purified Methanococcus aTFB showed striking similarities to eucaryal TATA-binding proteins (TBP). It exists as a dimer in solution (5), can be replaced by yeast and human TBPs in cell-free transcription reactions (6), and binds in mobility gel shift assays (7) to DNA fragments harboring an archaeal (8,9) or eucaryal TATA box. Furthermore, the protein translation of a putative TBP homologue encoded in the genome of Thermococcus (10) was able to substitute for Methanococcus aTFB in cell-free transcription reactions and showed serological crossreaction with this polypeptide (11).…”

“…Furthermore, the protein translation of a putative TBP homologue encoded in the genome of Thermococcus (10) was able to substitute for Methanococcus aTFB in cell-free transcription reactions and showed serological crossreaction with this polypeptide (11). Owing to its low stability purification of the aTFA activity thus far was not possible, but the incubation of aTFB or eucaryal TBPs in combination with aTFA results in template commitment (7,6), suggesting that it binds to and stabilizes the aTFB-promoter complex.…”

Occurrence and Characterization of Mercury Resistance in the Hyperthermophilic Archaeon Sulfolobus solfataricus by Use of Gene Disruption

“…The regulation of gene expression is an active area of research in archaea because of this evolutionary overlap and the attraction of a less complex experimental system. Examples of this overlap include homologous promoter structure (21,41), orthologs of TATA binding protein (29,37,48), TFIIB (referred to as TFB in archaea [18,38,39]), TFIIE␣ (TFE␣ [6,22]), TFIIS (TFS [25]), and the 12-subunit RNA polymerase II (28). They appear, however, to lack homologs of TFIIA, TFIIF, and TFIIH.…”

Events during Initiation of Archaeal Transcription: Open Complex Formation and DNA-Protein Interactions

“…This finding is in line with the data derived from analyses of archaeal genomes that indicate the absence of additional eucaryotic-like initiation factors like TFIIA, TFIIF, and TFIIH in archaea. Competition experiments using templates of different length and gel shift and footprinting experiments have shown that aTBP binds to the archaeal TATA box and that TFB stabilizes binding of aTBP to the promoter (8,12,23,26,30). This preinitiation complex seems to recruit the archaeal RNA polymerase to the promoter.…”