INSULIN RESISTANT DIABETES
During last two decades our research interests have been understanding the molecular mechanism of insulin resistant diabetes. In one hand my laboratory has contributed in deciphering the complex signal transduction cascade to elucidate the mechanism of regulation of insulin resistance in skeletal muscle and neuronal cells. On the other hand the laboratory has developed novel models of insulin resistance in skeletal muscle cells as well as in neuron, first of its kind, based on insulin signal transduction. Through in vitro and in vivo studies my laboratory has identified FAK, PPARγ, p38 MAP kinase, PTEN, SHIP, Sirt2, Pak 2, Ncor as important regulators of insulin resistance in skeletal muscle and/or neuronal cells.My laboratory has also made attempts to demonstrate the potential of the insulin resistant cell models for molecular target-based screening, althoughlow-throughput, of prospective anti-diabetic compounds. We have also made attempt to understand the effect of some drugs associated with signal transduction cascades those regulate insulin resistant diabetes. Currently we are looking deeper into the interactions and functions of very critical insulin signalling proteins Akt, AS160 in regulating insulin resistant diabetes.
Regulation of flagellar motility of the protozoan parasite Leishmania
Leishmaniasis represents a group of geographically widespread diseases caused by different species of kinetoplastid parasites of genus Leishmania. These parasites lead a digenetic life in two specific hosts, the sandfly, where they proliferate as motile, flagellated promastigotes and in mammals (including humans) where they invade utilizing their flagellum and then grow intracellularly. In the sandfly host, the flagellum performs several attachment mechanisms that allow the passage of the promastigotes to anterior parts of the gut. Once the promastigotes are transferred to the mammalian host, the vigorous and unusual oscillations of flagellar tip invade the macrophages, reorienting the parasite and damaging the macrophage plasma membrane. The Leishmania flagellum thus, is a highly versatile organelle that exhibits intricate environment triggered responses far beyond simple fluid swimming behaviour. However, till date flagellar motility and its regulation in Leishmania remains poorly understood despite the importance in its survival and infectivity.
Studies of the flagellar ultrastructure have been possible in the related trypanosome Trypanosoma brucei using RNAi techniques which is not possible in Leishmania spp. except only in L. braziliensis. In humans, defects in cilia cause a group of severe diseases called ciliopathies. These defects constitute both structural defects as well as defects in the motility of the cilia. Eukaryotic parasites like trypanosomes have served as attractive models for the study of such genetic defects in humans with extensive research on structure and assembly of the cilia. However, there is no suitable model till date for the study of the signalling and regulatory mechanisms of ciliary motility in ciliopathies.
In our laboratory, we have developed a demembranated ATP-reactivated model (LRP) for studying molecular mechanisms that regulate flagellar motility in L. donovani promastigotes. Such a model allows the researcher complete control to manipulate the cell chemically and mechanically under controlled standardized conditions. With our model we investigated the role of one of the popular second messenger molecules, cyclic-AMP and discovered a novel role. We observed that cAMP, via protein kinase A mediates “wave reversal” of the flagellar waveform to a ciliary waveform. The ciliary waveform has been believed to have a role in tactic responses such as obstacle-avoidance, chemotaxis, osmotaxis etc. We thus, proposed a cAMP-dependent signalling pathway that regulates tactic responses in Leishmania parasites, necessary for its survival. We believe that reactivation of flagellar motility in demembranated Leishmania presents it as an attractive model for flagellar motility studies. With this model we are also interested in elucidating the effects of other second messenger molecules, role of dynein ATPases in regulating the flagellar beat and waveform.
Sequential stills from fast-capture videomicroscopy of live (beat frequency = 22.22 Hz) and LRP reactivated (beat frequency 20 Hz) cells. Both cells generate waves proximally (tip-to-base). The cells complete one beat cycle in the times shown. Time intervals in milliseconds (ms) are shown on bottom right. Bar, 10 μm.
Ciliary (base-to-tip) waveform in LRP reactivated L. donovani in presence of 0.5 μM cAMP. (a) A montage of stills showing one complete ciliary beat of a LRP reactivated cell (Beat frequency = 6.89 Hz) with 0.5 μ M cAMP. Images displayed after every 20 ms. Time intervals (ms) are shown on bottom right. Bar, 10 μm. (b) Ciliary waveforms of ‘a’ were tracked and superimposed using BohBohSoft. Red and blue lines correspond to first track and last track of the beat cycle respectively. Bar, 5 μ m. (c) Stills of the cell orientation at the beginning of 4 consecutive beat cycles. The degree of change in direction per beat are shown on top right (white). Bar, 10 μm.