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A Loss of Function in LprG-Rv1410c Homologues Attenuates Growth during Biofilm Formation in Mycobacterium smegmatis
Lisa-Marie Nisbett1, Mary L Previti1, Jessica C Seeliger1
1Department of Pharmacological Sciences, Stony Brook University, 100 Nicolls Road, Stony Brook, NY 11794, USA.
Abstract:
MmpL (mycobacterial membrane protein large) proteins are integral membrane proteins that have been implicated in the biosynthesis and/or transport of mycobacterial cell wall lipids. Given the cellular location of these proteins, however, it is unclear how cell wall lipids are transported beyond the inner membrane. Moreover, given that mycobacteria grow at the poles, we also do not understand how new cell wall is added in a highly localized and presumably coordinated manner. Here, we examine the relationship between two lipid transport pathways associated with the proteins MmpL11 and LprG-Rv1410c. The lipoprotein LprG has been shown to interact with proteins involved in cell wall processes including MmpL11, which is required in biofilms for the surface localization of certain lipids. Here we report that deletion of mmpL11 (MSMEG_0241) or the lprG-rv1410c operon homologues MSMEG_3070-3069 in Mycobacterium smegmatis produced similar biofilm defects that were distinct from that of the previously reported mmpL11 transposon insertion mutant. Analysis of pellicle biofilms, bacterial growth, lipid profiles, and gene expression revealed that the biofilm phenotypes could not be directly explained by changes in the synthesis or localization of biofilm-related lipids or the expression of biofilm-related genes. Instead, the shared biofilm phenotype between ΔMSMEG_3070-3069 and ΔmmpL11 may be related to their modest growth defect, while the origins of the distinct mmpL11::Tn biofilm defect remain unclear. Our findings suggest that the mechanisms that drive pellicle biofilm formation in M. smegmatis are not connected to crosstalk between the LprG-Rv1410c and MmpL11 pathways and that any functional interaction between these proteins does not relate directly to their lipid transport function.
Insights
Investigating mycobacterial membrane protein large (MmpL) and LprG-Rv1410c pathways in Mycobacterium smegmatis reveals shared biofilm defects. These defects are linked to growth, not direct lipid transport, suggesting independent functions in pellicle biofilm formation.
Area of Science:
- Microbiology and Molecular Biology
- Cellular Biology
- Biochemistry
Background:
- Mycobacterial membrane protein large (MmpL) proteins are integral membrane proteins involved in cell wall lipid transport.
- The precise mechanisms of cell wall lipid transport beyond the inner membrane and localized cell wall synthesis in mycobacteria remain unclear.
- LprG-Rv1410c is a lipoprotein known to interact with proteins in cell wall processes, including MmpL11, which influences lipid localization in biofilms.
Purpose of the Study:
- To investigate the relationship between the MmpL11 and LprG-Rv1410c lipid transport pathways in *Mycobacterium smegmatis*.
- To understand the role of these pathways in pellicle biofilm formation and cell wall integrity.
Main Methods:
- Deletion mutants of *mmpL11* (MSMEG_0241) and the *lprG-rv1410c* operon homologues (MSMEG_3070-3069) were created in *Mycobacterium smegmatis*.
- Analysis included pellicle biofilm formation assays, bacterial growth rate measurements, lipid profiling, and gene expression analysis.
- Comparison with a previously characterized *mmpL11* transposon insertion mutant.
Main Results:
- Deletion of *mmpL11* or the *lprG-rv1410c* operon homologues resulted in similar biofilm defects, distinct from the *mmpL11* transposon mutant.
- The observed biofilm phenotypes were not directly explained by alterations in the synthesis or localization of biofilm-related lipids or changes in gene expression.
- Both deletion mutants exhibited a modest growth defect, which may explain their shared biofilm phenotype.
Conclusions:
- The LprG-Rv1410c and MmpL11 pathways in *M. smegmatis* do not appear to interact or crosstalk in a manner directly related to pellicle biofilm formation.
- The distinct biofilm defect of the *mmpL11* transposon mutant suggests complex regulation or alternative functions not captured by simple deletion.
- The findings indicate that the mechanisms driving pellicle biofilm formation are not solely dependent on the direct functional interaction between these two lipid transport pathways.
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