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Insights into the function of FhaA, a cell division-associated protein in mycobacteria
Abstract:
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.
Insights
Mycobacterium smegmatis FhaA protein is crucial for cell wall synthesis and stability. Its absence leads to shorter cells and increased antibiotic sensitivity, revealing its role in maintaining cell envelope integrity.
Area of Science:
- Microbiology
- Molecular Biology
- Cell Biology
Background:
- FhaA is a forkhead-associated domain protein in Mycobacterium smegmatis.
- Its depletion causes peptidoglycan precursor accumulation, but its function remains unclear.
Purpose of the Study:
- To elucidate the function of FhaA in Mycobacterium smegmatis.
- To investigate the mechanism behind FhaA's role in cell wall synthesis.
Main Methods:
- Construction and analysis of an fhaA knockout (ΔfhaA) strain.
- Complementation studies using homologous and heterologous fhaA genes.
- Interaction studies with penicillin-binding protein A (PbpA).
- Assessment of antibiotic sensitivity and cell morphology.
Main Results:
- The ΔfhaA mutant exhibited a short cell length phenotype, indicating a defect in cell elongation or cell wall synthesis.
- FhaA was identified as an interacting partner of PbpA, a peptidoglycan biosynthesis enzyme.
- FhaA appears to stabilize PbpA, as its absence led to reduced PbpA levels.
- The ΔfhaA mutant displayed sensitivity to various antibiotics, suggesting a general permeability defect.
Conclusions:
- FhaA plays a significant role in peptidoglycan biosynthesis and mycobacterial cell wall synthesis.
- FhaA is involved in stabilizing PbpA, contributing to cell envelope integrity.
- The findings highlight FhaA as a potential target for novel antimycobacterial strategies.
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