Department of Biotechnology, BMS Block-1, South Campus, Panjab University, Chandigarh, India.
Department of Biotechnology, BMS Block-1, South Campus, Panjab University, Chandigarh 160014, India.
Gene. 2021 Jul 30;791:145720. doi: 10.1016/j.gene.2021.145720. Epub 2021 May 18.
Mycobacterium tuberculosis has distinct cell wall composition that helps in intracellular survival of bacteria. Rv1900c, a two domain protein, has been grouped in lip gene family. The expression of rv1900c was upregulated under acidic, nutritive and iron stress conditions in M. tuberculosis H37Ra. To investigate the biological effect of Rv1900c in mycobacterium physiology, rv1900c gene was cloned in M. smegmatis, a surrogate host. Its counterpart MSMEG_4477 in M. smegmatis demonstrated 38% protein similarity with Rv1900c. MSMEG_4477 gene was knocked out in M. smegmatis by homologous recombination. rv1900c and MSMEG_4477 genes, cloned in pVV16, were expressed in the M. smegmatis knockout strain (M. smegmatis ΔMSMEG_4477). Gene knockout significantly altered colony morphology and growth kinetics of M. smegmatis. M. smegmatis ΔMSMEG_1900 (pVV16::rv1900c) colonies were less wrinkled and had smooth surface as compared to M. smegmatis ΔMSMEG_4477. The changes were reverted back to normal upon expression of MSMEG_4477 in knockout strain M. smegmatis ΔMSMEG_4477 (pVV16::MSMEG_4477). The expression of rv1900c enhanced the biofilm formation and survival of bacteria under various in vitro stresses like acidic, nutritive stress, including lysozyme, SDS and multiple antibiotics treatment in comparison to control. On the other hand the expression of rv1900c decreased the cell wall permeability. The resistance provided by M. smegmatis ΔMSMEG_4477 (pVV16::MSMEG_4477) was comparable to M. smegmatis having vector alone (MS_vec). The lipid content of M. smegmatis ΔMSMEG_1900 (pVV16::rv1900c) was observed to be different from M. smegmatis ΔMSMEG_4477 (pVV16::MSMEG_4477). M. smegmatis ΔMSMEG_1900 (pVV16::rv1900c) was more tolerant to stress conditions in comparison to M. smegmatis ΔMSMEG_4477 (pVV16::MSMEG_4477). Expression of rv1900c enhanced the intracellular survival of mycobacteria. Therefore, the present study suggested an association of Rv1900c to the stress tolerance by cell wall modification that might have resulted in enhanced intracellular survival of the mycobacteria.
结核分枝杆菌具有独特的细胞壁组成，有助于细菌在细胞内存活。Rv1900c 是一种具有两个结构域的蛋白质，已被归入脂基因家族。在结核分枝杆菌 H37Ra 中，rv1900c 在酸性、营养和铁胁迫条件下的表达上调。为了研究 Rv1900c 在分枝杆菌生理学中的生物学效应，将 rv1900c 基因克隆到了替代宿主耻垢分枝杆菌中。耻垢分枝杆菌中的 MSMEG_4477 与 Rv1900c 具有 38%的蛋白质相似性。通过同源重组敲除了耻垢分枝杆菌中的 MSMEG_4477 基因。rv1900c 和 MSMEG_4477 基因被克隆到 pVV16 中，并在耻垢分枝杆菌的敲除菌株（M. smegmatisΔMSMEG_4477）中表达。基因敲除显著改变了耻垢分枝杆菌的菌落形态和生长动力学。与 M. smegmatisΔMSMEG_4477 相比，M. smegmatisΔMSMEG_1900（pVV16::rv1900c）的菌落较少起皱，表面光滑。在敲除菌株 M. smegmatisΔMSMEG_4477（pVV16::MSMEG_4477）中表达 MSMEG_4477 后，这些变化恢复正常。与对照相比，rv1900c 的表达增强了细菌在各种体外应激条件下（如酸性、营养应激）的生物膜形成和存活能力，包括溶菌酶、SDS 和多种抗生素处理。另一方面，rv1900c 的表达降低了细胞壁通透性。与携带空载体的耻垢分枝杆菌（MS_vec）相比，耻垢分枝杆菌ΔMSMEG_4477（pVV16::MSMEG_4477）提供的抗性相当。与耻垢分枝杆菌ΔMSMEG_4477（pVV16::MSMEG_4477）相比，耻垢分枝杆菌ΔMSMEG_1900（pVV16::rv1900c）的脂质含量有所不同。与耻垢分枝杆菌ΔMSMEG_4477（pVV16::MSMEG_4477）相比，耻垢分枝杆菌ΔMSMEG_1900（pVV16::rv1900c）对压力条件更耐受。rv1900c 的表达增强了分枝杆菌的细胞内存活。因此，本研究表明 Rv1900c 与细胞壁修饰有关，与应激耐受性有关，这可能导致分枝杆菌的细胞内存活能力增强。
