The Phosphatidyl- myo -Inositol Dimannoside Acyltransferase PatA Is Essential for Mycobacterium tuberculosis Growth In Vitro and In Vivo

Abstract

Mycobacterium tuberculosis comprises an unusual cell envelope dominated by unique lipids and glycans that provides a permeability barrier against hydrophilic drugs and is central for its survival and virulence. Phosphatidyl-myo-inositol mannosides (PIMs) are glycolipids considered not only key structural components of the cell envelope but also the precursors of lipomannan (LM) and lipoarabinomannan (LAM), important lipoglycans implicated in host-pathogen interactions. Here, we focus on PatA, a membrane associated acyltransferase that transfers a palmitoyl moiety from palmitoyl–CoA to the 6-position of the mannose ring linked to 2-position of inositol in PIM1/PIM2. We validate that the function of PatA is vital for M. tuberculosis in vitro and in vivo. We constructed a patA conditional mutant and showed that silencing patA is bactericidal in batch cultures. This phenotype was associated with significant reduced levels of Ac1PIM2, an important structural component of the mycobacterial inner membrane. The requirement of PatA for viability was also demonstrated during macrophage infection and in a mouse model of infection, where a dramatic decrease in viable counts was observed upon silencing of the patA gene. This is reminiscent of the behavior of PimA, the mannosyltransferase that initiates the PIM pathway, also found to be essential for M. tuberculosis growth in vitro and in vivo. Altogether, the experimental data highlights the significance of the early steps of PIM biosynthetic pathway for M. tuberculosis physiology and uncover that PatA is a novel target for drug discovery programs against this major human pathogen. IMPORTANCE TB is the leading cause of death from a single infectious agent. The emergence of drug resistance in strains of M. tuberculosis, the etiologic agent of TB, emphasizes the need of identifying new targets and antimicrobial agents. The mycobacterial cell envelope is a major factor in this intrinsic drug resistance. Here, we have focused on the biosynthesis of PIMs, key virulence factors and important components of the cell envelope. Specifically, we have determined PatA, the acyltransferase responsible for the first acylation step of the PIM synthesis pathway, is essential in M. tuberculosis. These results highlight the importance of early steps of PIM biosynthetic pathway for mycobacterial physiology and the suitability of PatA as a potential new drug target.

Protective Effect of Rifampicin Loaded by HPMA-PLA Nanopolymer on Macrophages Infected with Mycobacterium Tuberculosis

“…Although it plays a certain role in reducing the mortality of TB, its low targeting of macrophages leads to a low killing rate, resulting in a longer treatment cycle for patients as well as some inevitable toxic and side effects [ 4 , 5 ]. Therefore, more efficient and active killing of MTB in macrophages and reducing the emergence of drug-resistant MTB are the most pressing challenges facing TB treatment [ 6 ]. This study will also contribute to the treatment of TB, hoping to provide some reference for the effective treatment of TB patients from the breakthrough point of the nanotargeted drug delivery system.…”

Understanding the Mannose Transfer Mechanism of Mycobacterial Phosphatidyl-myo-inositol Mannosyltransferase A from Molecular Dynamics Simulations

“…7 Phosphatidylinositol mannoside acyltransferase A (PatA) can add an acyl group to the 6-position of the mannose ring attached at the 2-position of the myo-inositol of PIM 2 to produce Ac 1 PIM 2 . 8 An unknown acyltransferase can add another acyl group at the 3-position of the inositol group of Ac 1 PIM 2 to synthesize Ac 2 PIM 2 . 9 In one of the strains of Mycobacterium tuberculosis , PimC had been identified that can add another mannose to the Ac 1 PIM 2 to produce Ac 1 PIM 3 .…”

Molecular ruler mechanism and interfacial catalysis of the integral membrane acyltransferase PatA

“…Specifically, they regulate the selective transport of molecules across the bacterial membranes and, as enzymes, they catalyze a myriad of biochemical reactions. The bacterial membrane constituents are represented by a substantial number of species, the amount and distribution of which also varies in space and time (6)(7)(8)(9)(10)(11)(12). Therefore, the structural diversity of membranes provides identity to bacterial cells, which strongly influence the development and maintenance of a large number of bacteria life forms.…”

“…1B ) ( 20 , 23 ). PatA was found important for the optimal growth of M. smegmatis mc 2 155 and an essential enzyme for the growth of M. tuberculosis in vitro and in vivo ( 11 , 23 ). Two models were originally proposed for the biosynthesis of Ac 1 PIM 2 in mycobacteria.…”

“…PatA is an essential acyltransferase involved in the biosynthesis of phosphatidyl-myo-inositol mannosides (PIMs) and the metabolically related lipomannan (LM) and lipoarabinomannan (LAM). These species are glycolipids/lipoglycans noncovalently anchored through their phosphatidyl-myo-inositol (PI) moiety to the inner and outer membranes of the mycobacterial cell envelope (2,(7)(8)(9)(10)(11)(12)(13)(14). PIMs, LM, and LAM are thought to be important virulence factors during of Mycobacterium tuberculosis infection (15).…”