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Status |
Public on Aug 02, 2010 |
Title |
The Role of the Rad4-Rad23 Complex and Rad4 Ubiquitination in UV-Responsive Transcription |
Organism |
Saccharomyces cerevisiae |
Experiment type |
Expression profiling by array
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Summary |
The Rad23/Rad4 protein complex plays a major role in DNA damage recognition during nucleotide excision repair (NER) in yeast. We recently showed that two distinct pathways contribute to efficient NER in yeast. The first operates independently of de novo protein synthesis and requires a nonproteolytic function of the 19S regulatory complex of the 26S proteasome and Rad23. The second pathway requires de novo protein synthesis, and relies on the activity of a newly identified Rad7-containing E3 ubiquitin ligase that ubiquitinates Rad4 in response to UV. Surprisingly, we found that cells deleted of either Rad23 or Rad4 caused reduced Rad4 and Rad23 mRNA levels respectively. We considered the possibility of an unexpected role of Rad23 and Rad4 in regulating the expression of genes involved in the transcriptional response to DNA damage. Gene expression profiling has suggested that Rad23 and Rad4 may function as a complex to affect transcription of a small subset of genes in response to UV damage. To determine how Rad4 and Rad23 contribute to the regulation of these genes, we have examined the occupancy of Rad4/Rad23 in their promoter regions by chromatin immunoprecipitation (ChIP), both in the presence and absence of UV damage. Our preliminary ChIP data suggests that the Rad4/Rad23 complex regulates a set of genes in response to UV light. We also proposed that the transcriptional regulatory activity of the Rad4-Rad23 complex required Rad4 ubiquitination. These arrays test this theory using the psocs mutant strain, which is unable to facilitate Rad4 ubiquitination after UV irradiation.
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Overall design |
*** This Series represents the gene expression component of the study.
Expression analysis was performed on the pRAD7 strain, which served as the WT control, and harboured the WT RAD7 sequence on the pRS313 plasmid, the psocs strain, which harboured the same plasmid with 2 point mutations in the RAD7 sequence that prevented post-UV Rad4 ubiquitination. Expression analysis was also conducted on a pRAD7 and psocs Strain with RAD23 deleted. Analysis was performed using untreated strains, and strains 15 minutes and 60 minutes after 100Jm-2 UV irradiation. mRNA was extracted from logaritmically growing cells.
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Contributor(s) |
Humphryes N, Reed S |
Citation(s) |
26150418 |
Submission date |
Jul 27, 2010 |
Last update date |
Jul 01, 2016 |
Contact name |
Simon Reed |
E-mail(s) |
ReedSH1@cf.ac.uk
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Organization name |
Cardiff University
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Department |
School of Medicine
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Street address |
Heath Park
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City |
CARDIFF |
ZIP/Postal code |
CF14 4XN |
Country |
United Kingdom |
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Platforms (1) |
GPL90 |
[YG_S98] Affymetrix Yeast Genome S98 Array |
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Samples (30)
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GSM570869 |
psocs Untreated, biological rep1 |
GSM570870 |
psocs Untreated, biological rep2 |
GSM570871 |
psocs Untreated, biological rep3 |
GSM570872 |
pRAD7 Δrad23 Untreated, biological rep1 |
GSM570873 |
pRAD7 Δrad23 Untreated, biological rep2 |
GSM570874 |
psocs Δrad23 Untreated, biological rep1 |
GSM570875 |
psocs Δrad23 Untreated, biological rep2 |
GSM570876 |
pRAD7 15, biological rep1 |
GSM570877 |
pRAD7 15, biological rep2 |
GSM570878 |
pRAD7 15, biological rep3 |
GSM570879 |
psocs 15, biological rep1 |
GSM570880 |
psocs 15, biological rep2 |
GSM570881 |
psocs 15, biological rep3 |
GSM570882 |
pRAD7 Δrad23 15, biological rep1 |
GSM570883 |
pRAD7 Δrad23 15, biological rep2 |
GSM570884 |
psocs Δrad23 15, biological rep1 |
GSM570885 |
psocs Δrad23 15, biological rep2 |
GSM570886 |
pRAD7 60, biological rep1 |
GSM570887 |
pRAD7 60, biological rep2 |
GSM570888 |
pRAD7 60, biological rep3 |
GSM570889 |
psocs 60, biological rep1 |
GSM570890 |
psocs 60, biological rep2 |
GSM570891 |
psocs 60, biological rep3 |
GSM570892 |
pRAD7 Δrad23 60, biological rep1 |
GSM570893 |
pRAD7 Δrad23 60, biological rep2 |
GSM570894 |
psocs Δrad23 60, biological rep1 |
GSM570895 |
psocs Δrad23 60, biological rep2 |
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Relations |
BioProject |
PRJNA131529 |
Supplementary file |
Size |
Download |
File type/resource |
GSE23204_RAW.tar |
42.8 Mb |
(http)(custom) |
TAR (of CEL, CHP) |
Processed data included within Sample table |
Processed data provided as supplementary file |
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