title: Protein kinase A (PknA) of Mycobacterium tuberculosis is independently activated and is critical for growth in vitro and survival of the pathogen in the host.
abstract: The essential mycobacterial protein kinases PknA and PknB play crucial roles in modulating cell shape and division. However, the precise in vivo functional aspects of PknA have not been investigated. This study aims to dissect the role of PknA in mediating cell survival in vitro as well as in vivo. We observed aberrant cell shape and severe growth defects when PknA was depleted. Using the mouse infection model, we observe that PknA is essential for survival of the pathogen in the host. Complementation studies affirm the importance of the kinase, juxtamembrane, and transmembrane domains of PknA. Surprisingly, the extracytoplasmic domain is dispensable for cell growth and survival in vitro. We find that phosphorylation of the activation loop at Thr(172) of PknA is critical for bacterial growth. PknB has been previously suggested to be the receptor kinase, which activates multiple kinases, including PknA, by trans-phosphorylating their activation loop residues. Using phospho-specific PknA antibodies and conditional pknB mutant, we find that PknA autophosphorylates its activation loop independent of PknB. Fluorescently tagged PknA and PknB show distinctive distribution patterns within the cell, suggesting that although both kinases are known to modulate cell shape and division, their modes of action are likely to be different. This is supported by our findings that expression of kinase-dead PknA versus kinase-dead PknB in mycobacterial cells leads to different cellular phenotypes. Data indicate that although PknA and PknB are expressed as part of the same operon, they appear to be regulating cellular processes through divergent signaling pathways.
authors.affiliation: From the National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India, the Department of Biotechnology, Nehru Arts and Science College, Coimbatore 641105, India, and.
authors.forename: Sathya Narayanan
authors.initials: SN
authors.lastname: Nagarajan
authors.affiliation: From the National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Sandeep
authors.initials: S
authors.lastname: Upadhyay
authors.affiliation: From the National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Yogesh
authors.initials: Y
authors.lastname: Chawla
authors.affiliation: From the National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Shazia
authors.initials: S
authors.lastname: Khan
authors.affiliation: From the National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Saba
authors.initials: S
authors.lastname: Naz
authors.affiliation: the Department of Biotechnology, Nehru Arts and Science College, Coimbatore 641105, India, and.
authors.forename: Jayashree
authors.initials: J
authors.lastname: Subramanian
authors.affiliation: CSIR-Institute of Genomics and Integrative Biology, Mathura Road, New Delhi 110020, India.
authors.forename: Sheetal
authors.initials: S
authors.lastname: Gandotra
authors.affiliation: From the National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India, vinaykn@nii.ac.in.
authors.forename: Vinay Kumar
authors.initials: VK
authors.lastname: Nandicoori
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D015153:Blotting, Western; D004789:Enzyme Activation; D005260:Female; D054884:Host-Pathogen Interactions; D008297:Male; D008810:Mice, Inbred C57BL; D050296:Microbial Viability; D008855:Microscopy, Electron, Scanning; D008856:Microscopy, Fluorescence; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D014376:Tuberculosis / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; C451020:PknA protein, Mycobacterium tuberculosis; C475060:PknB protein, Mycobacterium tuberculosis; D017346:Protein Serine-Threonine Kinases
keywords: Bacterial Protein Kinase; Cell Signaling; Conditional Mutant; Mutant; Mycobacterium tuberculosis; Phosphorylation; PknA