Ssl2/TFIIH function in Transcription Start Site Scanning by RNA Polymerase II in Saccharomyces cerevisiae

Ssl2, as an essential subunit of the RNA Polymerase II (Pol II) general transcription factor TFIIH, promotes melting of promoter DNA through its ATPase-dependent DNA translocase activity within the Pol II pre-initiation complex. In Saccharomyces cerevisiae, after DNA melting, Ssl2 is proposed to drive Pol II scanning downstream for usable transcription start sites (TSSs) through this same ATP-dependent DNA translocase activity. However, how Pol II catalytic activity or activities of other general transcription factors integrate with Ssl2/TFIIH activity for promoter scanning and what ways Ssl2 modulates TSS selection by scanning have not been well studied. Here, we report the identification of ssl2 alleles conferring phenotypes in vivo consistent with altered Pol II initiation. We find that these ssl2 alleles alter TSS selection by scanning in ways that are distinct from how changes to Pol II or other GTF activities alter TSS selection by scanning. Specific predictions arise for interactions among initiation mutants depending on whether they function in promoter scanning through modulating initiation at an individual TSS or if they control the probability that a TSS is reached during the scanning process (scanning processivity). We test these predictions genetically and through analysis of transcription output at ADH1 and at promoters across the genome. Our data support a model whereby ssl2 alleles alter Ssl2 processivity and therefore the probability that TSSs are scanned. Examination of TSS usage genome-wide finds global effects of ssl2 alleles on TSS usage and the potential coupling of Pol II activity and Ssl2/TFIIH processivity during scanning. We propose that the initiation by promoter scanning is determined by the interaction of two functional networks, one controlling initiation efficiency and one controlling the processivity of scanning.