Doctor of Philosophy (PhD)
IRS24 is a strain of Deinococcus radiodurans carrying mutations in two loci, uvrA and irrE, rendering it sensitive to the lethal effects of UV and ionizing radiation. These sensitivities can be reversed by introducing the wild type irrE allele back into IRS24 via natural transformation. The irrE allele was localized to a 970bp region of D. radiodurans R1 Chromosome I containing one putative open reading frame, DR0167, and 179bp of upstream sequence. Subsequent sequence analysis of the irrE allele in IRS24 revealed a transition mutation at codon 111 of DR0167 (IrrE) resulting in an arginine to cysteine amino acid substitution. IrrE was also inactivated by transposon mutagenesis in the wild type strain, R1. The insertion mutant, LSU2030, has a more pronounced sensitivity to both UV and ionizing radiation suggesting that IRS24's IrrE retains some activity. BLASTp analysis of IrrE reveals only minimal similarity to proteins currently available in protein sequence databases. A "weak" helix-turn-helix motif was identified within this protein that may indicate a capacity to bind DNA and, perhaps, a potential role for IrrE in gene regulation. To test whether the mutation in IrrE causes a regulatory deficiency, we examined the pattern of transcription following ionizing radiation, comparing LSU2030 and R1 using DNA microarray technology. Our analysis has determined that IrrE is a transcriptional activator that controls expression of many genes including recA. A recent investigation has shown that, as in E. coli's response to stress, RecA is necessary for proteolytic cleavage of the LexA repressor in D. radiodurans. However, unlike the E. coli paradigm, deinococcal RecA is not controlled by LexA. Functional IrrE appears to be necessary for recA induction and in mounting an effective response to exogenous stress. This analysis along with examination of the transcriptional changes exhibited by R1 and a lexA-defective strain following ionizing radiation has focused our attention to a subset of 12 genes that are induced during D. radiodurans' response to ionizing radiation and recovery from prolonged desiccation. These genes appear to be critical to this species' ability to survive both stresses and may be involved in DNA double strand break repair.
Document Availability at the Time of Submission
Release the entire work immediately for access worldwide.
Earl, Ashlee M., "Global expression analysis of Deinococcus radiodurans' response to ionizing radiation: irrE is a novel regulator of this response" (2003). LSU Doctoral Dissertations. 3527.
John R. Battista