Nucleotide excision repair (NER) of DNA-yeast as a eukaryotic model

NER of DNA in eukaryotic cells requires a large number of gene products. The genetic and biochemical versatility of the yeast S. cerevisiae has provided an informative model system to explore this biochemical complexity. Some of this complexity derives from the fact that most (presumably all) of the subunits of the RNA polymerase II basal transcription factor IIH (core TFIIH) are also indispensable for NER. This been directly demonstrated using cell-free extracts of mutants defective in the TFIIH components Ssl1, Ssl2, Rad3, Tfb1 and Tfb2. The NER function of the Tfb3 and Tfb4 subunits of core TFIIH is under investigation. The Kin28 and Ccl1 kinase subunits of the transcription factor TFIIK, which associates with core TFIIH, do not appear to be involved in NER. No other basal transcription factors have been identified which participate in this process. A supercomplex (repairosome) has been identified in extracts of yeast cells. This includes core TFIIH plus the Rad1, Rad10, Rad2, Rad4, Rad14 and Rad23 NER proteins, all of which are indispensable for NER in vitro. The products of the RAD7 and RAD16 genes are also required for NER in vitro. However Rad7 and Rad16 proteins have not been detected in the repairosome. Evidence will presented that some of the plasmid substrate DNA molecules in the cell-free NER system are apparently packaged into nucleosomes and that these molecules are preferentially repaired in vitro, consistent with the known requirement for Rad7 and Rad16 for NER of transcriptionally-repressed DNA.