My research work has gained an international reputation over the last 44 years for innovations and original procedures in widely disparate areas, involving enzymology and signal transduction, in the areas of the cardiovascular system and colorectal cancer. Of note, the discovery of the role of lipid modification in my laboratory is likely to lead to early diagnosis and treatment of colorectal cancer.
Dr. Rajendra Sharma’s research has spanned more than 40 years. In four manuscripts, he is summarizing his areas of interest in research, including enzymology and signal transduction in the cardiovascular system and colorectal cancer.
The first manuscript “Calmodulin-binding proteins: A journey of 40 years,” looks at the crucial roles calcium-binding proteins play in multiple cell processes; how these proteins could help tissues protect and repair the heart in ischemic conditions; and how calcineurin, a protein enzyme present in neuron cell death after epilepsy, could potentially be used as a marker to diagnose colorectal cancer.
I have had a longstanding research interest in the area of biochemical mechanisms of the signal transduction processes, with special emphasis on Ca2+ calmodulin (CaM)-regulated enzymes and their involvement in the regulation of both cAMP and Ca2+ second messenger systems. One of the central questions concerning signal transduction processes is how cells use the limited number of signalling cascades to achieve diverse responses to each of the multitude of cell stimuli. The existence of multiple regulatory activities at each step of signalling cascades provides the potential mechanisms for achieving such diverse and specific cell responses. Multiple signalling cascades undergo complex and rigorously - regulated interactions to achieve a unique and integrated cell response. Biochemical studies have revealed numerous interactions in the signal transduction pathways and the number continues to increase. However, the contributions made by these regulatory reactions under any specific physiological condition are not yet fully understood.
I have focused my effort mainly on the regulation of calmodulin-dependent cyclic nucleotide phosphodiesterase (PDE1). My research has contributed significantly to the discovery of PDE1 isozymes, the delineation of the molecular, catalytic and regulatory properties of the isozymes and the elucidation of the physiological and regulatory significance of these isozymes. These isozymes are regulated by multiple second messenger-dependent regulatory systems. For each of the isozymes, we have proposed a working hypothesis describing how the multiple regulatory activities interact with each other to achieve regulatory advantages in controlling cAMP metabolism in the cell during cell activation. One of the main features of the hypothesis is that during cell activation, the various regulatory activities are temporally separated. The result suggests that a temporal separation of second messenger-dependent reactions can be conceived as a natural consequence of dynamic fluxes of the messengers.
I have also focused my research interest of the regulation of CaM-dependent protein kinases, CaM-stimulated protein phosphatase (calcineurin) and high molecular weight CaM-binding protein (HMWCaMBP) which was discovered in my laboratory. Subsequently it was established that HMWCaMBP is homologous to calpastatin, an inhibitor of the Ca2+-activated cysteine proteases, calpains.
Our laboratory is currently working towards elucidating the role and expression of CaM-regulated proteins in cardiac ischemia and perfusion. Besides this endeavour, the significance of the non-cardiomyocyte population in heart muscle is being investigated.
In addition, my interest in the role of spatial separation of regulatory activities during the process of signal transduction has led me to initiate a study of the lipid modification by myristic acid (a 14 carbon fatty acid). Myristoylation of proteins is catalyzed by the ubiquitously distributed eukaryotic enzyme N-myristoyltransferase (NMT). This lipid modification is important for the proper functions of several proteins including many of those involved in cancer progression. Originally we have established that NMT is more active in human colonic epithelial neoplasms than in normal human colonic tissue. A several fold increase in NMT activity was observed in polyps and stage B1 tumors located in descending colon close to the rectum as compared to adjacent normal appearing mucosa. Since NMT activity is highest level in the colon cancer tissue, a limitation of assessing the activity and protein expression of NMT for prognostic/ diagnostic purposes is difficult because endoscopic biopsy must be performed to obtain tumor tissue. Towards this direction, we demonstrated that altered expression and localization of NMT is found in the peripheral blood and bone marrow of colon cancer patients. Furthermore, immunohistochemical analysis revealed weak to negative staining for NMT in peripheral blood mononuclear cells (PBMC) chosen as controls, whereas strong positivity was observed in the PBMC of colon cancer patients. In addition, we observed that NMT was localized mostly in the nuclei of the bone marrow mononuclear cells of the colon cancer patients, whereas NMT remained cytoplasmic in the control bone marrow specimens. The different NMT expression offers the basis of a potential adjunct investigative tool for screening or diagnosis of patients at risk for, or suspected of, having colon cancer.
We are working on understanding the regulation of this protein in vivo to develop therapies directed to NMT to inhibit the development and progression of cancer. We have identified that HSC70, a protein generally involved in cellular protection from stress, inhibits NMT functions. We aim to understand the mechanisms of NMT inhibition by HSC70. Further we are also exploiting the roles of phosphorylation on the regulation of NMT function.
- Sreejit Nair, Sharma RK (2019) Effect of ischemia and reperfusion on Ca2+ and calmodulin-regulated proteins in primary murine cardiac non-myocytes. J Mol Biol Ther 1, 72-79
- Sreejit Nair, Sharma RK (2019) The relevance of cardiac fibroblasts in ischemic heart disease. J Mol Biol Ther 1, 56-64
- Sharma RK (2019) Ca2+ /Calmodulin-dependent Cyclic Nucleotide Phosphodiesterase (PDE1): 40 Years Journey. J Mol Biol Ther 1, 24-44
- Sharma RK (2019) Protein N-Myristoyltransferase: A potential biomarker for colon cancer. J Mol Biol Ther 1, 4-5
- Sharma RK, Das U, Kumar S, Dimmock JR (2018) Protein N-myristoyltransferase: 30 Years Journey. J Drug Discov Dev Res 1, 7-38
- Sharma RK, Parameswaran S (2018) Calmodulin-Binding Proteins: A Journey of 40 Years. Cell Calcium 75, 89-100
- Das U, Howlett J, Kumar S, Parameswaran S, Sharma A, Dimmock JR, Sharma RK (2018) NMT (N-myristoyltransferase). Encyclopaedia of Signalling Molecules 2, 3522-3533
- Parameswaran S, Sharma RK (2016) Insulin cannot induce adipogenic differentiation in primary cardiac cultures. Int J Angiol 25, 181-185
- Kumar S, Parameswaran S, Sharma RK (2015) Novel myristoylation of the sperm-specific hexokinase 1 isoform regulates its atypical localization. Biolopen. 4, 1679-1687
- Parameswaran S, Sharma RK (2015) Expression of calcineurin, calpastatin and heat shock proteins during ischemia and reperfusion. Biochem Biophys Reports 4, 207-214
- Kumar S, Sharma RK (2015) N-Terminal region of the catalytic domain of human N-myristoyltransferase 1 acts as an inhibitory module. PLoS One DOI: 10.1371/journal. pone. 0127661
- Parameswaran S, Sharma RK (2014) Ischemia and reperfusion induce differential expression of calpastatin and its homologue high molecular weight calmodulin-binding protein in murine cardiomyocytes. PLoS One 9:e114653.Doi:10.1371/journal. pone. 0114653
- Parameswaran S, Sharma RK (2014) Altered expression of calcineurin, calpain, calpastatin and HMWCaMBP in cardiac cells following ischemia and reperfusion. Biochem Biophys Res Commun 443, 604-609
- Parameswaran S, Kumar S, Sharma RK (2013) Cardiomyocyte culture – an update on the in vitro cardiovascular model and future challenges. Can J Physiol Pharm 91, 985-998
- Das U, Kumar S, Dimmock JR, Sharma RK (2012) Inhibition of protein myristoylation : A therapeutic protocol in developing anticancer agents. Curr Cancer Drug Targ 12, 667-692
- Sreejit P, Sharma RK (2012) High molecular weight calmodulin-binding protein: 20 years onward – A potential therapeutic calpain inhibitor. Cardiovase Drug Ther 26, 321-330
- Selvakumar P, Lakshmikuttyamma A, Das U, Pati HN, Dimmock JR, Sharma RK (2009) NC2213: a novel methionine aminopeptidase 2 inhibitor in human colon cancer HT29 cells. Mol Cancer 8.65 doi 10.1186/1476-4598-8-65
- Shrivastav A, Varma S, Lawman Z, Yang SH, Ritchie SA, Bonham K, Singh, SM, Saxena A, Sharma RK (2008) Requirement of N-myristoyltransferase – 1 in the development of monocytic lineage . J Immunol 180, 1019-1028
- Shrivastav A, Varma S, Saxena A, DeCoteau J, Sharma RK (2007) N-myristoyltransferase: A novel potential diagnostic tool for colon cancer. J Transl Med 5, 58-63
- Lakshmikuttyamma A, Selvakumar P, Charavaryamath, Singh B, Tuchek J, Sharma RK (2006) Expression of calcineurin and its interacting proteins in epileptic fowl. J Neurochem 96, 366-373
- Yang SH, Shrivastav A, Kosinsk C, Sharma RK, Gavino B, Chen MH, Peters L, Chuang PT, Young SG, Bergo M (2005) N-myristoyltransferase 1 is essential for early mouse development. J Biol Chem 280, 18990-18995
- Lakshmikuttyamma A, Selvakumar P, Anderson DH, Datla RS, Sharma RK (2004) Molecular cloning of bovine cardiac muscle heat shock protein 70 kDa and its phosphorylation by cAMP-dependent protein kinase in vitro. Biochemistry 43, 13340-13347
- Shrivastav A, Pasha MK, Selvakumar P, Olson DJH, Ross ARS, Dimmock JR, Sharma RK (2003) Potent inhibitor of N-myristoylation: A novel molecular target for cancer. Cancer Research 63, 7975-7978
- Seitz DP, Pasha MK, Singh B, Chu A, Sharma RK (2002) Localization and characterization of Calcineurin in Bovine Eye. Invest Ophth Vis Sci 43, 15-21
- Kakkar R, Wang X, Radhi JM, Rajala RVS, Wang R, Sharma RK (2001) Decreased expression of high molecular weight calmodulin-binding protein and its correlation in the apoptosis in ischemia-reperfused rat heart. Cell Calcium 29, 59-71
- Rajala RVS, Radhi JM, Kakkar R, Datla RSS, Sharma RK (2000) Increased expression of N-myristoyltransferase in human gallbladder carcinomas. Cancer 88, 1992-1999
- Raju RVS, Kakkar R, Datla RSS, Radhi JM and Sharma RK (1998) Myristoyl CoA: protein N-myristoyl transferase from bovine cardiac muscle: Molecular cloning, kinetic analysis and in vitro proteolytic cleavage by m-calpain. Exp Cell Res 241, 23-35
- Kakkar R, Raju RVS, Mellgren RL, Radhi JM and Sharma RK (1997) High molecular weight calmodulin-binding protein contains calpastatin activity. Biochemistry 36, 11550-11555
- Magnuson B, Raju RVS, Moyana TN, Sharma RK (1995) Increased N-myristoyl transferase activity observed in rat and human colonic tumors. J Natl Cancer Inst 87, 1630-1635
- Raju RVS, Kalra J, Sharma RK (1994) Purification and properties of bovine spleen N-myristoyl CoA protein: N-myristoyl transferase. J Biol Chem 269, 12080-12083
- King MJ, Sharma RK (1993) Identification, purification and characterization of a membrane-associated N-myristoyl transferase inhibitor protein from bovine brain. Biochem J 291, 635-639
- Sharma RK (1991) Phosphorylation and characterization of bovine heart calmodulin‑dependent phosphodiesterase. Biochemistry 30, 5963‑5968
- Sharma RK (1990) Purification and characterization of novel calmodulin‑binding protein from cardiac muscle. J Biol Chem 265, 1152‑1157
- Sharma RK, Wang JH (1986) Calmodulin and Ca2+-dependent phosphorylation and dephosphorylation of 63-kDa subunit-containing bovine brain calmodulin‑stimulated cyclic nucleotide phosphodiesterase isozyme. J Biol Chem 261, 1322‑1328
- Sharma RK, Wang JH (1985) Differential regulation of bovine brain calmodulin‑dependent cyclic nucleotide phosphodiesterase isozyme by cyclic AMP-dependent protein kinase and calmodulin-dependent phosphatase. Proc Natl Acad Sci USA 82, 2603‑2607
- Sharma RK, Adachi AM, Adachi K, Wang JH (1984) Demonstration of bovine brain calmodulin‑dependent cyclic nucleotide phosphodiesterase isozymes by monoclonal antibodies. J Biol Chem 259, 9248‑9254
- Huang CY, Chau V, Chock PB, Wang JH, Sharma RK (1982) Mechanism of activation of cyclic nucleotide phosphodiesterase: Requirement of the binding of four Ca2+ to calmodulin for activation. Proc Natl Acad Sci USA 78, 874‑876
- Sharma RK, Wang TH, Wirch E, Wang JH (1980) Purification and properties of bovine brain calmodulin‑dependent cyclic nucleotide phosphodiesterase. J Biol Chem 255, 5916‑5923
- Sharma RK, Wirch E (1979) Cyclic nucleotide phosphodiesterase from rabbit lung. Biochem Biophys Res Commun 91, 338‑344
- Sharma RK, Desai R, Waisman DM, Wang JH (1979) Purification and subunits structure of bovine brain modulator binding protein. J Biol Chem 254, 4276-4282