Mycobacterium tuberculosis Methyltransferase Rv1515c Can Suppress Host Defense Mechanisms by Modulating Immune Functions Utilizing a Multipronged Mechanism

Abstract

Mycobacterium tuberculosis (M. tb) gene Rv1515c encodes a conserved hypothetical protein exclusively present within organisms of MTB complex and absent in non-pathogenic mycobacteria. In silico analysis revealed that Rv1515c contain S-adenosylmethionine binding site and methyltransferase domain. The DNA binding and DNA methyltransferase activity of Rv1515c was confirmed in vitro. Knock-in of Rv1515c in a model mycobacteria M. smegmatis (M. s_Rv1515c) resulted in remarkable physiological and morphological changes and conferred the recombinant strain with an ability to adapt to various stress conditions, including resistance to TB drugs. M. s_Rv1515c was phagocytosed at a greater rate and displayed extended intra-macrophage survival in vitro. Recombinant M. s_Rv1515c contributed to enhanced virulence by suppressing the host defense mechanisms including RNS and ROS production, and apoptotic clearance. M. s_Rv1515c, while suppressing the phagolysosomal maturation, modulated pro-inflammatory cytokine production and also inhibited antigen presentation by downregulating the expression of MHC-I/MHC-II and co-stimulatory signals CD80 and CD86. Mice infected with M. s_Rv1515c produced more Treg cells than vector control (M. s_Vc) and exhibited reduced effector T cell responses, along-with reduced expression of macrophage activation markers in the chronic phase of infection. M. s_Rv1515c was able to survive in the major organs of mice up to 7 weeks post-infection. These results indicate a crucial role of Rv1515c in M. tb pathogenesis.

Mycobacterium tuberculosis Essential Gene Thymidylate Synthase Is Involved in Immune Modulation and Survival inside the Host

“…Under standard tissue culture conditions of 37 °C and 5% CO 2 , cells were grown and maintained. 30 After the initial frozen stocks were seeded, eight passages later, all experiments were completed in the different cell lines. 30 4.5.…”

“…30 After the initial frozen stocks were seeded, eight passages later, all experiments were completed in the different cell lines. 30 4.5. In-Vitro Survival of M.s_Vc and M.s_ThyX under Normal Growth Conditions.…”

Mycobacterium tuberculosis-macrophage interaction: Molecular updates

“…Upregulation of these receptors activates STRING and p38MAPK-NOX2, inhibits pro-inflammatory cytokines (TNF-a, IL-1b, and ROS), and promotes anti-inflammatory cytokines (IFN-b and IL-10) (Dabla et al, 2022). Mtb effectors such as the early secreted protein target 12 (EST12), the methyltransferase (Rv1515c) encoded by the RD 6, or the dormancy regulator DosS, repress macrophage immune defenses (ROS, RNS, phagolysosomal maturation, proinflammatory response, inflammasome, antigen presentation) by activating the JAK2-STAT5a signaling pathway (Gautam et al, 2019;Yang et al, 2021;Rani et al, 2022;Yao et al, 2022). Mtb escape strategies are becoming increasingly non-negligible, and the aforementioned are recent and far to be exhaustive.…”

Rv2231c, a unique histidinol phosphate aminotransferase from Mycobacterium tuberculosis, supports virulence by inhibiting host-directed defense

“…For flow cytometric assessment of macrophage activation markers, cells were treated with an Fc block for 15 min at 4°C. Cells were then treated with cocktail of fluorescently labelled anti-mouse antibodies specific for cell-surface markers and incubated at 4°C for 15 min [37]. After staining procedure, cells were gently washed with FACS buffer followed by fixing in 4% paraformaldehyde.…”