Nandicoori Vinay Kumar, Ph.D
Signal Transduction Lab 1
National Institute of Immunology
Aruna Asaf Ali Marg
New Delhi 110 067
Tel: (91)-11-26703789,
Fax: (91)-11-26742125
e.mail: vinaykn@nii.res.in

1. Cell signaling & host-pathogen interactions

            Cell-signaling is a consortium of cellular events wherein the transmission of environmental signals to cells sets off a cascade of reactions that ultimately culminate in changes in gene expression and activity.  One of the major mechanisms through which cell signaling pathways operate is the reversible phosphorylation of cellular proteins. The current research in our laboratory has two main focuses. The first is the characterization of M. tuberculosis serine/threonine protein kinases (STPKs) with special emphasis on examining the role they play in modulating the pathogen’s survival in the host. The second is the investigation of signalling events that are triggered in host macrophages upon M. tuberculosis infection, with particular interest in the impact of M. tuberculosis infection on the MAP kinase pathways of host macrophages.

Signaling events in Mycobacterium tuberculosis and their host macrophages.

Analysis of the M. tuberculosis genome sequence suggests the presence of 11 putative eukaryotic-like serine/threonine protein kinases. Overexpressing of PknA in M. tuberculosis results in a deviation from normal cell morphology with the cells forming an elongated and branched structure, while overexpression of PknB in M. tuberculosis results in the formation of widened and bulging cells.  Protein kinase G (PknG) has been shown to play a role in the survival of the pathogen in host macrophages by modulating phagosome-lysosome fusion (lysosomal transfer) after macrophages phagocytose mycobacterium, and PknG kinase activity is essential for mediating its effect on lysosomal transfer. Our laboratory is presently focusing on four protein kinases - PknA, PknB, PknK and PknG. We have cloned the genes encoding these kinases and the proteins have been overexpressed and purified. We are now engaged in biochemically characterizing them, with special emphasis on the identification and characterization of direct targets of these kinases both in M. tuberculosis and in host macrophages. The role of kinase - mediated phosphorylation events in M. tuberculosis growth and its survival in the host are being examined.

Signalling events in the MAP kinase pathways of host macrophages after M. tuberculosis infection

            Mouse macrophages infected by a pathogenic strain of mycobacteria respond with a decreased activation of the ERK and p38 MAP kinase pathways. MAP kinases activate a number of transcription factors, which upon phosphorylation stimulate the synthesis of various inflammatory proteins.  Not much is known about substrates of MAP kinases that are differentially phosphorylated upon mycobacterial infection. The technology developed during my years of postdoctoral research will be utilized to identify ERK2 substrates that are differentially phosphorylated in macrophages infected with Mycobacterium tuberculosis as compared to uninfected macrophages.

2. The role of cell signaling events in modulating the functions and dynamics of nucleoporins.

            Nucleocytoplasmic transport between the nucleus and the cytoplasm occurs through nuclear pore complexes (NPC), ~ 60 MDa macromolecular structures that span from cytoplasm to nucleus across the lipid bilayers of the nuclear envelope. Various molecules (cargo), including proteins and RNA, are transported through the NPC in both, energy dependent and energy-independent manner. The nuclear pore complex is composed of proteins named nucleoporins (Nups) which are approximately 30 in number in vertebrates. NPCs are suggested to be dynamic structures which undergo conformational changes during transport and cell cycle progression.  Phosphorylation of nuclear transport proteins was shown by various groups to either up regulate or down regulate transport machinery. However, the precise role of these phosphorylations in the nuclear pore function have not been elucidated.  We are interested in addressing the role of phosphorylation in the function of nucleoporins and their dynamics. 

SELECTED PUBLICATIONS

1. Vomastek, T, Iwanicki, M. P, Burack, W. R, Tiwari, D., Kumar, D., Parsons, J.T., Weber, M. J.^§  & Nandicoori, V. K.^§ (2008) ERK2 phosphorylation sites and docking domain on the Nuclear Pore Complex protein Tpr cooperatively regulate ERK2-Tpr interaction.     Mol. Cell Biol. Sep 15. [Epub ahead of print]

2. Samantaray, S., Marathe, U., Dasgupta, S., Nandicoori, V. K.  & Roy, R. P. (2008) Peptide-sugar ligation catalyzed by transpeptidase sortase: a facile approach to neoglycoconjugate synthesis. J Am Chem Soc. 130, 2132-3.

3. Sajish, M., Tiwari, D., Rananaware, D., Nandicoori, V. K.^§ & Prakash, B.^§ (2007) A Charge Reversal Differentiates (p)ppGpp Synthesis by  Monofunctional and Bifunctional Rel Proteins. J. Biol. Chem. 282, 34977-83.

          §  Corresponding authors

4. Kumar, N. V.*, Eblen, S.T.* & Weber, M. J. (2004) Identifying specific kinase substrates through engineered kinases and ATP analogs. Methods 32, 389-397.

5. Eblen, S.T.*, Kumar, N. V.*, Shah, K., Henderson, M. J., Watts, C. K. W., Shokat, K. M. & Weber, M. J. (2003) Identification of novel ERK2 substrates through use of an engineered kinase and ATP analogs. J. Biol. Chem. 278, 14926-14935.

          * Co-first authors

6. Kumar, N.V. & Bernstein, L. R. (2001) A new analytical scale DNA affinity binding assay for analyses of specific protein DNA interactions. Anal. Biochem. 299, 203-210.       

7.  Kumar, N. V. & Bernstein, L. R.  (2001) Ten ERK-related proteins in three distinct classes associate with AP-1 proteins and/or AP-1 DNA. J. Biol. Chem. 276, 32362-32372.

8.  Kumar, N. V.  & Bernstein, L. R. (2000) Screening of a cDNA protein expression library by enhanced chemiluminescence detection. Biotechniques 29, 418-424.

9.  Thanedar, S. S., Kumar, N. V. & Varshney, U. (2000) Fate of the initiator tRNAs is sensitive to the critical balance between interacting proteins. J. Biol. Chem. 275, 20361-20367.

10. Kumar, N.V. & Varshney, U. (1997) Contrasting effects of single stranded DNA binding protein on the activity of uracil DNA glycosylase from Escherichia coli towards different DNA substrates. Nucleic Acids Res. 25, 2336-2343. 

11. Li, S., Kumar, N.V.,  Varshney, U. & RajBhandary, U.L. (1996)  Role of amino acids  attached to tRNA in formylation  and in initiation of protein synthesis.   J. Biol. Chem.  271, 1022-1028.

12.  Kumar, N.V. & Varshney, U. (1994) Inefficient  excision of uracil from loop regions of DNA oligomers by E. coli  uracil DNA glycosylase.  Nucl. Acids Res.  22, 3737-3741.