Characterization of Transcription from TATA-Less Promoters: Identification of a New Core Promoter Element XCPE2 and Analysis of Factor Requirements

Abstract

BackgroundMore than 80% of mammalian protein-coding genes are driven by TATA-less promoters which often show multiple transcriptional start sites (TSSs). However, little is known about the core promoter DNA sequences or mechanisms of transcriptional initiation for this class of promoters.Methodology/Principal FindingsHere we identify a new core promoter element XCPE2 (X core promoter element 2) (consensus sequence: A/C/G-C-C/T-C-G/A-T-T-G/A-C-C/A+1-C/T) that can direct specific transcription from the second TSS of hepatitis B virus X gene mRNA. XCPE2 sequences can also be found in human promoter regions and typically appear to drive one of the start sites within multiple TSS-containing TATA-less promoters. To gain insight into mechanisms of transcriptional initiation from this class of promoters, we examined requirements of several general transcription factors by in vitro transcription experiments using immunodepleted nuclear extracts and purified factors. Our results show that XCPE2-driven transcription uses at least TFIIB, either TFIID or free TBP, RNA polymerase II (RNA pol II) and the MED26-containing mediator complex but not Gcn5. Therefore, XCPE2-driven transcription can be carried out by a mechanism which differs from previously described TAF-dependent mechanisms for initiator (Inr)- or downstream promoter element (DPE)-containing promoters, the TBP- and SAGA (Spt-Ada-Gcn5-acetyltransferase)-dependent mechanism for yeast TATA-containing promoters, or the TFTC (TBP-free-TAF-containing complex)-dependent mechanism for certain Inr-containing TATA-less promoters. EMSA assays using XCPE2 promoter and purified factors further suggest that XCPE2 promoter recognition requires a set of factors different from those for TATA box, Inr, or DPE promoter recognition.Conclusions/SignificanceWe identified a new core promoter element XCPE2 that are found in multiple TSS-containing TATA-less promoters. Mechanisms of promoter recognition and transcriptional initiation for XCPE2-driven promoters appear different from previously shown mechanisms for classical promoters that show single “focused” TSSs. Our studies provide insight into novel mechanisms of RNA Pol II transcription from multiple TSS-containing TATA-less promoters.

Identification of a Secondary Promoter within the Human B Cell Receptor Component Gene hCD79b

“…3A). The heterogeneity of these 5Ј termini agreed with prior studies (9) and was consistent with the GCrich and TATA-less structure of the promoter region (35). Of note, ϳ10% of the 5Ј termini in this 5Ј-RACE assay mapped to a site within exon 2 (Fig.…”

Novel Core Promoter Elements and a Cognate Transcription Factor in the Divergent Unicellular Eukaryote Trichomonas vaginalis

“…As no previously described eukaryotic core promoter elements were identified by the MEME program, with the exception of the Inr element, we used custom Perl scripts to directly search the URDB for the core promoter elements: TATA box, TFIIB, BRE u , BRE d , DPE, MTE, DCE, XCPE1, and XCPE2 (1,5,32,37,50,66). Although all are represented, with the exception of MTE and XCPE2, little to no positional conservation is observed.…”

“…An array of core promoter elements that direct accurate initiation of transcription at the TSS have been identified in metazoans and include the TATA box, the initiator (Inr) element, the transcription factor II B (TFIIB), recognition elements (BRE u and BRE d ), the downstream promoter element (DPE), the motif 10 element (MTE), the downstream core element (DCE), and the X core promoter elements (XCPEs; XCPE1 and XCPE2) (1,5,16,21,32,33,37,50,58,66). These can function independently, as in the case of the TATA box, Inr, and XCPEs, or in varied combinations to direct basal transcription.…”

Promoting developmental transcription

“…Additional elements identified and studied in individual human or viral genes include two TFIIB recognition elements (BRE U and BRE D ), the downstream core element (DCE), and the X core promoter element (XCPE) ( Table 1) (Lewis et al, 2000;Tokusumi et al, 2007;Anish et al, 2009) (Fig. 3A).…”