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Characterising resuscitation promoting factor fluorescent-fusions in mycobacteria
Iria Uhía1, Nitya Krishnan1, Brian D Robertson2
1MRC Centre for Molecular Bacteriology and Infection, Department of Medicine, Imperial College London, London, SW7 2AZ, UK.
BMC Microbiology
|April 14, 2018
Summary
Resuscitation promoting factor proteins (Rpfs) from Mycobacterium tuberculosis, when fused to mCherry, can revive dormant bacteria. These functional mCherry fusions aid in studying Rpf proteins, crucial in tuberculosis pathogenesis.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Resuscitation promoting factor proteins (Rpfs) are essential for reviving dormant mycobacteria.
- Their specific roles and localization in Mycobacterium tuberculosis pathogenesis are largely unknown.
- Understanding Rpf function is critical for tuberculosis research.
Purpose of the Study:
- To construct fluorescent fusions of M. tuberculosis Rpf proteins.
- To investigate the function and localization of Rpfs using these tools.
Main Methods:
- Construction of fluorescent protein fusions (mCherry and EGFP) of five M. tuberculosis Rpfs.
- Functional assays to assess resuscitation promotion.
- Time-lapse video microscopy for cellular localization studies.
Main Results:
- mCherry-Rpf fusions were functional, promoting cell growth.
- EGFP-Rpf fusions were non-functional and not secreted.
- No specific cellular localization was observed for either fusion type.
Conclusions:
- Functional mCherry-Rpf fusions were developed as tools for studying Rpf proteins.
- Rpfs are likely secreted via the Sec pathway.
- Further studies using these tools can elucidate Rpf localization, expression, and function in M. tuberculosis.
