Rpb4, a Non-essential Subunit of Core RNA Polymerase II ofSaccharomyces cerevisiae Is Important for Activated Transcription of a Subset of Genes

Abstract

A major role in the regulation of eukaryotic protein-coding genes is played by the gene-specific transcriptional regulators, which recruit the RNA polymerase II holoenzyme to the specific promoter. Several components of the mediator complex within the holoenzyme also have been shown to affect activation of different subsets of genes. Only recently has it been suggested that besides the largest subunit of RNA polymerase II, smaller subunits like Rpb3 and Rpb5 may have regulatory roles in expression of specific sets of genes. We report here, the role of Rpb4, a non-essential subunit of core RNA polymerase II, in activation of a subset of genes in Saccharomyces cerevisiae. We have shown below that whereas constitutive transcription is largely unaffected, activation from various promoters tested is severely compromised in the absence of RPB4. This activation defect can be rescued by the overexpression of cognate activators. We have localized the region of Rpb4 involved in activation to the C-terminal 24 amino acids. We have also shown here that transcriptional activation by artificial recruitment of the TATA-binding protein (TBP) to the promoter is also defective in the absence of RPB4. Surprisingly, the overexpression of RPB7 (the interacting partner of Rpb4) does not rescue the activation defect of all the promoters tested, although it rescues the activation defect of the heat shock element-containing promoter and the temperature sensitivity associated with RPB4 deletion. Overall, our results indicate that Rpb4 and Rpb7 play independent roles in transcriptional regulation of genes.

Whole Genome Expression Profiles of Yeast RNA Polymerase II Core Subunit, Rpb4, in Stress and Nonstress Conditions

“…Studies using promoter reporter constructs have also shown that various unrelated genes are affected to different extents in rpb4⌬ (8). On the other hand, the stress-related phenotypes of rpb4⌬ cells like temperature sensitivity and lethality during the stationary phase point toward a stress-specific role for RPB4 in transcription.…”

“…We have previously reported that the activation defect of rpb4⌬ strain can be partially rescued by overexpression of the cognate transcriptional activator (8). Msn2 (transcriptional activator in heat shock response) and some other proteins (that are not transcriptional activators) have been shown to partially rescue the temperature sensitivity of rpb4⌬ cells (7).…”

“…Strains and Plasmids-The rpb4⌬ and wild type strains (SY10-1: Mat a, his3⌬-200, ura3-52, leu2-3, 112, lys2-1, rpb4⌬::HIS3/pPS2; SY10-2: Mat a, his3⌬-200, ura3-52, leu2-3, 112, lys2, rpb4⌬::HIS3/pNS114) have been described before (8). The corresponding diploid rpb4⌬/rpb4⌬ strain was generated by crossing SY10 (Mat a,112,rpb4⌬::HIS3) to its Mat␣ sibling.…”

“…An rpb4⌬ mutant is unable to survive at extreme temperatures (Ͼ34°C and Ͻ12°C) and dies rapidly during a prolonged stationary phase (3,6). Recently, several groups have shown that Rpb4 plays an important role in the activation of many genes (7,8) but has a milder effect on the basal expression of these genes. Expression analysis of specific genes showed that RNA polymerase II from the mutant lacking Rpb4 cannot transcribe some genes (9).…”

The RNA Pol II sub-complex hsRpb4/7 is required for viability of multiple human cell lines

“…Second, Rpb4 (Kimura et al, 2001) and Rpb7 (Khazak et al, 1995;Na et al, 2003) can be found in the cytoplasm with Rpb4 mediating RNA export from the nucleus under stress conditions (Farago et al, 2003) and both Rpb4 and Rpb7 are involved in selective mRNA decay (Lotan et al, 2005;Goler-Baron et al, 2008). In terms of their biological roles, while scRpb4 and scRpb7 may not be identical (scRpb4 is conditional (Woychik and Young, 1989) and scRpb7 is essential (McKune et al, 1993)) both proteins are nonetheless involved in particular stress responses (Choder and Young, 1993;Sheffer et al, 1999;Maillet et al, 1999;Pillai et al, 2001) and are implicated in morphological changes that accompany nutrient starvation (Khazak et al, 1995;Pillai et al, 2003). Consistent with the conditional nature of scRpb4, global gene expression profiling in yeast indicates that a limited number of genes (~2% of the genome) are affected by Rpb4 (Pillai et al, 2003), although other evidence points to a more global effect (Miyao et al, 2001).…”