Analysis Of Deinococcus Radiodurans's Transcriptional Response To Ionizing Radiation, Hydrogen Peroxide, And Some Important Transcript Regulators Disruption

Deinococcus radiodurans(DEIRA)has an extraordinary resistance to gamma-radiation, desiccation and oxidizing agents.DEIRA has the ability to reconstruct the chromosomal fragments from hundreds of double stand DNA breaks(DSBs)during the post-irradiation incubation.It is generally known that the ability of its resistant is due to a complex network of DNA repair and metabolic pathway,but the molecular mechanisms underlying this character remain poorly understand.The complete genome sequence of the DEIRA is composed of two chromosomes,a megaplasmid and a small plasmid,coding a total of 3187 ORFs.Among these ORFs,about fifty percent proteins' functions are still unknown.Remarkably,the number of genes identified in DEIRA that are known to be involved in DNA repair is less than the number reported for Escherichia coli.Therefore,clarifying the functions of these unknown function proteins may help us to better understanding the mechanisms of extreme radioresistance in this organism.We constructed whole-genome cDNA microarray for DEIRA,and finished experiments as follow:1.Detection of transcriptome dynamics of DEIRA recovering from ionizing radiation at dose of 2 kGy.From the expression profile,we found that the process of DNA repair was induced in order.Furthermore,the response of oxidative stress related genes under low dose irradiation showed a different pattern from that of the acute high-level irradiation,many anti-oxidative related genes were induced at low dose.2.Transcriptome dynamics were examined in three mutant strains,including pprI mutant, recX mutant,and recQ mutant.The expression data of pprI mutant showed that PprI protein is a key protein to induce DNA repair genes,but it is not a regulator to anti-oxidative genes.On the contrary,the expression data of recX mutant showed that drrecX might relate to anti-oxidative genes,but not to DNA repair genes(including recA).Global transcriptome profiles responding to drrecQ deletion demonstrated that about 9.1%genes of the genome changed at least 2-fold in drrecQ mutant strain,such as iron homeostasis,oxidative related genes,and cell division related proteins.Furthermore,under 20mM hydrogen peroxide stress, the profile expression pattern of many genes,e.g.,recA,pprA,ddrA,ddrC,ddrB in drrecQ mutant strain was similar to that of wild type strain in acute gamma-irradiation and prolonged desiccation.So,we deduced that besides DrRecQ possesses direct ability to repair DNA damage in D.radiodurans,it also could maintain cellular homeostasis.3.Global transcriptome profiles responding to wild type strains under 20mM hydrogen peroxide stress were carried out.The data showed that both anti-oxidative genes and DNA repair genes were induced.It is suggested that DNA are damaged under 20 mM hydrogen peroxide treatment.4.Three oxidative-related genes were testified.DR0615 is a special OxyR homolog with only one conserved cysteine.In addition,QRT PCR and global transcription profile results showed that OxyR is not only a transcriptional activator(e.g.,katE,drb0125),but also a transcriptional repressor(e.g.,dps,mntH).We also found that Mn/Fe ion ratio is changed in dr0615 mutant.Sequence alignment results showed that DR0865 and DR2539 are ferric uptake regulation protein.However,our data excluded these possibilities.Based on out phenotype data and QRT-PCR data,we deduced that DR0865 is a PerR homolog,which negatively control katE,and DR2539 is a MntR homolog,which negatively regulate ABC-type Mn transport genes.In these experiments,we intent to discover the network of DNA repair and anti-oxidative, and construct a database for future research.