title: A potential target gene for the host-directed therapy of mycobacterial infection in murine macrophages.
abstract: Mycobacterium tuberculosis (MTB), one of the major bacterial pathogens for lethal infectious diseases, is capable of surviving within the phagosomes of host alveolar macrophages; therefore, host genetic variations may alter the susceptibility to MTB. In this study, to identify host genes exploited by MTB during infection, genes were non-selectively inactivated using lentivirus-based antisense RNA methods in Raw264.7 macrophages, and the cells that survived virulent MTB infection were then screened. Following DNA sequencing of the surviving cell clones, 26 host genes affecting susceptibility to MTB were identified and their pathways were analyzed by bioinformatics analysis. In total, 9 of these genes were confirmed as positive regulators of collagen α-5(IV) chain (Col4a5) expression, a gene encoding a type IV collagen subunit present on the cell surface. The knockdown of Col4a5 consistently suppressed intracellular mycobacterial viability, promoting the survival of Raw264.7 macrophages following mycobacterial infection. Furthermore, Col4a5 deficiency lowered the pH levels of intracellular vesicles, including endosomes, lysosomes and phagosomes in the Raw264.7 cells. Finally, the knockdown of Col4a5 post-translationally increased microsomal vacuolar-type H+-ATPase activity in macrophages, leading to the acidification of intracellular vesicles. Our findings reveal a novel role for Col4a5 in the regulation of macrophage responses to mycobacterial infection and identify Col4a5 as a potential target for the host-directed anti-mycobacterial therapy.
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Zhang
authors.initials: Z
authors.lastname: Bao
authors.affiliation: Zhejiang JFK Biological Technology Co., Ltd., Hangzhou, Zhejiang 310052, P.R. China.
authors.forename: Ran
authors.initials: R
authors.lastname: Chen
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Pei
authors.initials: P
authors.lastname: Zhang
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Shan
authors.initials: S
authors.lastname: Lu
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Xing
authors.initials: X
authors.lastname: Chen
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Yake
authors.initials: Y
authors.lastname: Yao
authors.affiliation: Zhejiang JFK Biological Technology Co., Ltd., Hangzhou, Zhejiang 310052, P.R. China.
authors.forename: Xiaozheng
authors.initials: X
authors.lastname: Jin
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Yilan
authors.initials: Y
authors.lastname: Sun
authors.affiliation: Department of Respiratory Medicine, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang 310003, P.R. China.
authors.forename: Jianying
authors.initials: J
authors.lastname: Zhou
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival / Q000235:genetics; D002478:Cells, Cultured; D024141:Collagen Type IV / Q000235:genetics* / Q000378:metabolism; D022162:Cytoplasmic Vesicles / Q000737:chemistry / Q000378:metabolism; D015870:Gene Expression; D054884:Host-Pathogen Interactions; D006863:Hydrogen-Ion Concentration; D008264:Macrophages / Q000378:metabolism* / Q000382:microbiology*; D008297:Male; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D009169:Mycobacterium tuberculosis / Q000502:physiology*; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism
chemical_list: C478159:COL4A5 protein, human; D024141:Collagen Type IV; D025262:Vacuolar Proton-Translocating ATPases