EsxA membrane-permeabilizing activity plays a key role in mycobacterial cytosolic translocation and virulence: effects of single-residue mutations at glutamine 5.
EsxA is required for virulence of Mycobacterium tuberculosis (Mtb) and plays an essential role in phagosome rupture and translocation to the cytosol of macrophages. Recent biochemical studies have demonstrated that EsxA is a membrane-permeabilizing protein. However, evidence that link EsxA membrane-permeabilizing activity to Mtb cytosolic translocation and virulence is lacking. Here we found that mutations at glutamine 5 (Q5) could up or down regulate EsxA membrane-permeabilizing activity. The mutation Q5K significantly diminished the membrane-permeabilizing activity, while Q5V enhanced the activity. By taking advantage of the single-residue mutations, we tested the effects of EsxA membrane-permeabilizing activity on mycobacterial virulence and cytosolic translocation using the esxA/esxB knockout strains of Mycobacterium marinum (Mm) and Mtb. Compared to wild type (WT), the Q5K mutant exhibited significantly attenuated virulence, evidenced by intracellular survival and cytotoxicity in mouse macrophages as well as infection of zebra fish embryos. The attenuated virulence of the Q5K mutant was correlated to the impaired cytosolic translocation. On the contrary, the Q5V mutant had a significantly increased cytosolic translocation and showed an overall increased virulence. This study provides convincing evidence that EsxA contributes to mycobacterial virulence with its membrane-permeabilizing activity that is required for cytosolic translocation.
Department of Biological Sciences, Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX 79968, USA.|School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, 45001, China.
Qi
Q
Zhang
Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA.|Medical College of China Three Gorges University, Yichang, 443002, China.
Decheng
D
Wang
Department of Biological Sciences, Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX 79968, USA.
Guozhong
G
Jiang
Department of Biological Sciences, Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX 79968, USA.
Wei
W
Liu
Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA.
Qing
Q
Deng
Department of Chemistry, University of Texas at El Paso, El Paso, TX, 79968, USA.
Xiujun
X
Li
Sino-Duth Biomedical Information Engineering School of Northeastern University, Shenyang, China.
Wei
W
Qian
Department of Biological Sciences, Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX 79968, USA.
Hugues
H
Ouellet
Department of Biological Sciences, Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX 79968, USA.|Sino-Duth Biomedical Information Engineering School of Northeastern University, Shenyang, China.
Jianjun
J
Sun
D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001422:Bacterial Adhesion / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002463:Cell Membrane Permeability; D003600:Cytosol / Q000378:metabolism*; D015536:Down-Regulation / Q000235:genetics; D004625:Embryo, Nonmammalian / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D005973:Glutamine / Q000235:genetics*; D059765:Homologous Recombination / Q000235:genetics; D006801:Humans; D051379:Mice; D050296:Microbial Viability / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity*; D021381:Protein Transport; D015854:Up-Regulation / Q000235:genetics; D014774:Virulence; D015027:Zebrafish / Q000196:embryology
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D005973:Glutamine