| Tuberculosis （TB）, caused by Mycobacterium tuberculosis （M. tuberculosis）, remains one of the major global public health threats. It causes Tuberculosis among millions of people each year and ranks as the second mortality from an infectious disease worldwide, after the human immunodeficiency virus （HIV）.According to the Global tuberculosis report2013, there were8.6million new TB cases in2012and1.3million TB deaths[11].Due to the emergence of multiple resistant bacteria,multiple resistant bacteria and co-infection with HIV, the situation of Tuberculosis is more serious.so the further research of M. tuberculosis are imminent.Genomic integrity is critical for organism survival and reproduction under stresses. Tuberculosis （TB）, caused by Mycobacterium tuberculosis （M. tuberculosis）, remains one of the major global public health threats. Environmental M. tuberculosis is under frequent UV radiation. M. tuberculosis within host is assaulted by host immune response, such as oxidative stress. The outcome of UV infliction is usually the forming of pyrimidine dimmers in DNA, which will disrupt the normal replication of DNA.Oxidative stress, which will lead to DNA strand break through disturbing the intracellular Ca2+homeostasis, represents another challenge for pathogen within host. M. tuberculosis is exceptional for its capacity to maintain genome integrity under stresses.M. tuberculosis genome encodes most known systems for genome integrity maintenance,such as the Nucleotide Excision Repair （NER）, Base Excision Repair （BER）, recombinational repair, non-homologous end joining, and SOS repair. M tuberculosis OxyR protein is a critical component of the oxidative stress response. OxyR is both a sensor of reactive oxygen species and a transcriptional activator. M tuberculosis genes katG and ahpC are also involved in response to oxidative stress. Genomic instability is recognized as a characteristic when M. tuberculosis expose to UV or reactive oxygen. We speculated that there might be as-yet unkown mechanism underlying the exceptional genome stability of M. tuberculosis.Forkhead-associated domain （FHA）, is phosphopeptide recognition domain embedded in some regulatory proteins. Several roles of FHA domain in bacteria have been documented, such as regulating the glutamate and lipid production, regulation of cell shape, type Ⅲ secretion, ethambutol resistance, sporulation, signal transduction, carbohydrate storage and transport, pathogenic and symbiotic host-bacterium interactions. In eukaryotes, many FHA domain-containing proteins localize in the nucleus and are responsible for the establishment and maintenance of cell cycle checkpoints, DNA repair, or transcriptional regulation.This study focuses on FHA-containing protein GarA（Rv1827, MSMEG3647）. M. tuberculosis GarA is reported to interact with three metabolic enzyme complexes to regulate their catalytic activities:Rv1248c （a-ketoglutarate decarboxylase,KGD）, Rv2476c（nicotinamide adenine dinucleotide（NAD）-dependent glutamate dehydrogenase, GDH） and Rv3859c （a subunit （GltB） of the glutamate synthase, GS）[15]. Given the widespread role of garA in eukaryotes, we speculated that M. tuberculosis FHA-containing protein GarA might serve role beyond the metabolic modulation. In this study, a FHA-domain protein GarA encoding M. smegmatis mutant, found by transposon mutants library screening, was sensitive to fluoroquinolone antibiotics. M. smegmatis△GarA colonies had a smoother, flatter morphology. The cell hydrophobicity of M. smegmatis△GarA was significantly higher than M. smegmatis mc2155. Loss of GarA specifically interferes with biofilm maturation. So we suggest that mutant of GarA alters component of the cell wall.we found that GarA mutant results in reduced sensitivity to UV exposure and more susceptible to oxidative stress. This may be relat to genomic stability.we also found that GarA mutant selectively altered the intracellular pools of amino acids ratios. But the role of amino acids in the genomic integrity maintenance remains largely untapped. Our research found a novel molecular mechanism preserving genome stability has never been reported before. GarA can regulate amino acid composition in intracellular pools, especially glutamic acid and serine. This is the reason that alters the sensitivity to UV and oxidative stress. |
