Viswanathan Gopinath, Yadav Sangya, Joshi Shrilaxmi V, Raghunand Tirumalai R
FEMS Microbiol Lett. 2017 Jan 1;364(2). doi: 10.1093/femsle/fnw294.
FhaA is a forkhead-associated domain-containing protein, the depletion of which leads to accumulation of peptidoglycan (PG) precursors at the septum and poles in Mycobacterium smegmatis (M. smegmatis), by a mechanism undefined thus far. To elucidate its function, we constructed an fhaA (MSMEG_0035) knockout (ΔfhaA) strain in M. smegmatis and demonstrated that this gene is dispensable for in vitro growth. The mutant showed a short cell length phenotype due to a probable defect in cell elongation/cell wall synthesis, which was reversed by complementation with both M. smegmatis and Mycobacterium tuberculosis (M. tb) fhaA (Rv0020c), confirming their association with the observed phenotype. The identification of penicillin binding protein A (PbpA), a PG biosynthesis enzyme as an interacting partner for mycobacterial FhaA, provided a hint into the functioning of FhaA. A drastic reduction in the levels of ectopically expressed PbpA in the ΔfhaA mutant vs wild-type M. smegmatis suggested that FhaA interacts with and stabilises PbpA. In addition, the fhaA deletion mutant was sensitive to multiple classes of antibiotics pointing to a general permeability defect. Our findings uncover a role for FhaA in PG biosynthesis and suggest its involvement in the maintenance of mycobacterial cell envelope integrity.
FhaA是一种含有叉头相关结构域的蛋白质，在耻垢分枝杆菌中，其缺失会导致肽聚糖（PG）前体在隔膜和两极积累，其机制迄今尚未明确。为了阐明其功能，我们构建了耻垢分枝杆菌fhaA（MSMEG_0035）基因敲除（ΔfhaA）菌株，并证明该基因对于体外生长并非必需。该突变体表现出细胞长度较短的表型，这可能是由于细胞伸长/细胞壁合成存在缺陷所致，用耻垢分枝杆菌和结核分枝杆菌的fhaA（Rv0020c）进行互补可逆转此表型，证实了它们与观察到的表型相关。青霉素结合蛋白A（PbpA）作为分枝杆菌FhaA的相互作用伴侣被鉴定出来，这为FhaA的功能提供了线索。与野生型耻垢分枝杆菌相比，ΔfhaA突变体中外源表达的PbpA水平大幅降低，这表明FhaA与PbpA相互作用并使其稳定。此外，fhaA缺失突变体对多种抗生素敏感，表明存在一般通透性缺陷。我们的研究结果揭示了FhaA在PG生物合成中的作用，并表明其参与维持分枝杆菌细胞壁的完整性。
