Abstract
Rad14p is a DNA damage recognition factor in nucleotide excision repair. Intriguingly, we show here that Rad14p associates with the promoter of a galactose-inducible GAL1 gene after transcriptional induction in the absence of DNA lesion. Such an association of Rad14p facilitates the recruitment of TBP, TFIIH, and RNA polymerase II to the GAL1 promoter. Furthermore, the association of RNA polymerase II with the GAL1 promoter is significantly decreased in the absence of Rad14p, when the coding sequence was deleted. These results support the role of Rad14p in transcriptional initiation. Consistently, the level of GAL1 mRNA is significantly decreased in the absence of Rad14p. Similar results are also obtained at other galactose-inducible GAL genes such as GAL7 and GAL10. Likewise, Rad14p promotes transcription of other non-GAL genes such as CUP1, CTT1, and STL1 after transcriptional induction. However, the effect of Rad14p on the steady-state levels of transcription of GAL genes or constitutively active genes such as ADH1, PGK1, PYK1, and RPS5 is not observed. Thus, Rad14p promotes initial transcription but does not appear to regulate the steady-state level. Collectively, our results unveil a new role of Rad14p in stimulating transcription in addition to its well-known function in nucleotide excision repair.
Highlights
Rad14p has well known functions in NER, its role in transcription is not clearly known
Rad14p Associates with the Promoter and Coding Sequence of the Transcriptionally Active, but Not Inactive, GAL1 Gene—To determine whether Rad14p associates with active gene in the absence of lesion, we employed the chromatin immunoprecipitation (ChIP) assay to analyze the level of Rad14p at the promoter and coding sequence of a galactose-regulated gene, GAL1, under inducible and non-inducible conditions
Rad14p was not associated with the transcriptionally inactive region of chromosome V in both raffinose and galactose-containing growth media (Fig. 1, B and C). These results support that Rad14p associates with the promoter and coding sequence of the transcriptionally active, but not inactive, GAL1 gene in the absence of DNA lesion
Summary
Rad14p has well known functions in NER, its role in transcription is not clearly known. We show here that Rad14p associates with the promoter of a galactose-inducible GAL1 gene after transcriptional induction in the absence of DNA lesion. The association of RNA polymerase II with the GAL1 promoter is significantly decreased in the absence of Rad14p, when the coding sequence was deleted These results support the role of Rad14p in transcriptional initiation. Our results unveil a new role of Rad14p in stimulating transcription in addition to its well-known function in nucleotide excision repair. Human XPA recognizes DNA lesion and plays a central role in NER (21, 24 –28). The steady-state or constitutive transcription is not altered in the absence of Rad14p These results unveil a new role of Rad14p in transcriptional regulation in addition to its well-known function in NER
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