Open AccessMycobacterium smegmatis PrrAB two-component system influences triacylglycerol accumulation during ammonium stress
Author(s) -
Jason D. Maarsingh,
Shelley E. Haydel
Publication year - 2018
Publication title -
microbiology
Language(s) - Uncategorized
Resource type - Journals
SCImago Journal Rank - 0.352
H-Index - 35
eISSN - 1465-2080
pISSN - 1350-0872
DOI - 10.1099/mic.0.000705
Subject(s) - mycobacterium smegmatis , mutant , operon , lipidomics , biochemistry , ammonium , mycobacterium tuberculosis , biology , strain (injury) , biosynthesis , mycobacterium , chemistry , wild type , microbiology and biotechnology , bacteria , gene , genetics , tuberculosis , medicine , organic chemistry , pathology , anatomy
The PrrAB two-component system is conserved across all sequenced mycobacterial species and is essential for viability in Mycobacterium tuberculosis, thus making it a promising drug target. The prrAB operon was successfully deleted in nonpathogenic Mycobacterium smegmatis, and the ∆prrAB mutant strain exhibited clumping in ammonium-limited medium and significantly reduced growth during ammonium and hypoxic stress. To assess the influence of M. tuberculosis PrrA overexpression, we constructed a recombinant M. smegmatis ∆prrAB mutant strain which overexpresses M. tuberculosis prrA. M. smegmatis prrAB and M. tuberculosis prrA complemented the M. smegmatis ∆prrAB deletion mutant in Middlebrook M7H9 and ammonium-limited media and during hypoxic and ammonium stress. Based on quantitative untargeted mass spectrometry-based lipidomics, triacylglycerol lipid species were significantly upregulated in the ∆prrAB mutant strain compared to the wild-type when cultured in ammonium-limited medium, revealing that M. smegmatis PrrAB influences triacylglycerol levels during ammonium stress. These results were qualitatively corroborated by thin-layer chromatography. Furthermore, the ∆prrAB mutant significantly upregulated expression of several genes (glpK, GPAT, WS/DGAT, accA3, accD4, accD6 and Ag85C) that participate in triacylglycerol and lipid biosynthetic pathways, thus corroborating the lipidomics analyses.