Genetic Interactions Between TFIIF and TFIIS

Abstract

The eukaryotic transcript elongation factor TFIIS is encoded by a nonessential gene, PPR2, in Saccharomyces cerevisiae. Disruptions of PPR2 are lethal in conjunction with a disruption in the nonessential gene TAF14/TFG3. While investigating which of the Taf14p-containing complexes may be responsible for the synthetic lethality between ppr2D and taf14D, we discovered genetic interactions between PPR2 and both TFG1 and TFG2 encoding the two larger subunits of the TFIIF complex that also contains Taf14p. Mutant alleles of tfg1 or tfg2 that render cells cold sensitive have improved growth at low temperature in the absence of TFIIS. Remarkably, the amino-terminal 130 amino acids of TFIIS, which are dispensable for the known in vitro and in vivo activities of TFIIS, are required to complement the lethality in taf14D ppr2D cells. Analyses of deletion and chimeric gene constructs of PPR2 implicate contributions by different regions of this N-terminal domain. No strong common phenotypes were identified for the ppr2D and taf14D strains, implying that the proteins are not functionally redundant. Instead, the absence of Taf14p in the cell appears to create a dependence on an undefined function of TFIIS mediated by its N-terminal region. This region of TFIIS is also at least in part responsible for the deleterious effect of TFIIS on tfg1 or tfg2 cold-sensitive cells. Together, these results suggest a physiologically relevant functional connection between TFIIS and TFIIF.

The transcription elongation factor TFIIS is a component of RNA polymerase II preinitiation complexes

“…Second, a GSTfusion of TFIIS was used to purify initiation factors such as Mediator, TAFs, and some GTFs from HeLa nuclear extracts (20). Third, DST1 interacts genetically with numerous genes encoding factors with roles in initiation such as MED31, MED3, SIN4, SRB5, SPT8, SWI1, SNF2, SNF5, and KIN28 (21)(22)(23)45) and with genes encoding factors involved in both initiation and elongation such as RPB1, RPB4, TFG1, TFG2, TAF14, and HTZ1 (21,22,24,45). In light of the well characterized role of TFIIS in stimulating elongation in vitro, the results of some of these genetic studies were interpreted as evidence that TFIIS could function at a step after PIC assembly (21,22,24).…”

“…Although this domain is not present in the cocrystal structure available between TFIIS and RNA polymerase II (9), it is reasonable to suggest that it would be on the periphery of the Pol II structure and would thus be available for interaction with other Pol II factors such as SAGA and Mediator components (21). In addition, although this domain is dispensable for the known in vitro and in vivo activities of TFIIS, it was recently shown to be required to complement the lethality of yeast cells lacking TAF14 and DST1 (24). This and other evidence suggested an undefined function of TFIIS mediated by its N-terminal region.…”

“…Notably, however, DST1 deletion strains exhibit growth defects in the presence of nucleotide-depleting drugs (10), microtubule-destabilizing drugs (22), and at elevated temperature (46,47). In addition, TFIIS becomes essential for normal growth in strains carrying mutant alleles of factors with roles in initiation such as Med31, Sin4, Srb5, Tfg3, Rpb1, Rpb4, Htz1, and Swi/Snf components (21)(22)(23)(24)45). It is likely that TFIIS is required for normal transcription initiation under conditions where efficient PIC assembly is compromised.…”

“…Domain I (amino acids 1-130) is composed of a four-helix bundle that is not required for the known biochemical and biological functions of TFIIS (19). Intriguingly, however, a number of biochemical and genetic interactions between Domain I and initiation factors have been described (see below) (20)(21)(22)(23)(24). Domain II (amino acids 131-240) forms a three-helix bundle and is tethered to Domain III (amino acids 265-309) through a short, unstructured linker (9,18).…”

“…This study also showed that TFIIS was required for efficient recruitment of TATA box-binding protein (TBP) and Pol II to the promoter, and, at high temperature, efficient recruitment of SAGA and Mediator components to the UAS was TFIIS-dependent. In addition to these molecular and biochemical studies, a number of genetic studies have uncovered functional interactions between DST1 and genes encoding factors that can function during initiation such as components of Mediator, Swi/Snf, and TFIIF (22)(23)(24). Although these results suggest that TFIIS may play a role during early events in transcription, no direct biochemical evidence has been presented to support this claim.…”