Ratnaparkhi Laboratory

             Animals as different as humans, worms and flies use remarkably similar molecular mechanisms to control their development. Discoveries of developmental paradigms in one organism have provided insights into the development processes of other organisms, including Homo sapiens. My research group utilizes the fruit fly Drosophila melanogaster as a model organism to study common molecular principles underlying animal development and disease, with a focus on Signalling. Regulation of signalling, within a cell, between two cell types and between organs is the theme of interest in my laboratory’s research.

Signalling in the innate immune response. In Drosophila, the Toll/NFkappaB and IMD/NFkappaB signalling cascades are the major facilitators of the host-defence response to pathogens. We are interested in uncovering new mechanisms and players that fine-tune the immune response. 

                   We hypothesize that the post-translational modifier SUMO is an important regulator of the host immune response (Hegde et. al., 2020). We find, using a proteomics screen, that ~650 proteins undergo SUMO conjugation upon immune challenge (Handu et. al., 2015).  Utilizing CRISPR/Cas9 genome editing, we have generated SUMO conjugation resistant (SCR) mutants of a number of SUMOylated proteins, by replacing target Lysines with Arginine. We have systematically subjected each of these mutants to an immune challenge and compared their response to wild-type animals. The broad findings emerging from these studies suggest that SUMO conjugation fine-tunes immune signalling by increasing or decreasing the strength of signalling in a context specific manner. We find that Jun SUMOylation attenuates IMD/NFkappaB signalling in the adult gut (Soory & Ratnaparkhi, 2022); Dorsal/ NFkappaB SUMOylation restrains Toll signalling in both immune and developmental contexts (Hegde et. al., 2022); SUMOylation of Caspar (Kaduskar et. al., 2020) dampens its ability as a negative regulator of IMD/NFB signalling and that SUMOylation of Arginyl tRNA synthetase, a member of the Muti-acetyl tRNA synthetase complex can modulate the immune response (Nayak et. al., 2021). Our collaborators, working on mammalian Fos/Jun signaling find that Fos/Jun transcriptionally regulate the gut SUMO cycle.

                  Serine hydrolases (SH), especially proteases, are important regulators of Toll/NFkappaB signalling. We have used Activity based protein profiling to develop a SH activity atlas (Kumar et. al., 2021). Further, six novel metabolic SH’s have been identified to respond to infection and we are currently generating reagents that will allow us to identify mechanistic roles for these in host defence. Earlier, we found that DmMt2, a DNA/RNA methyl transferase regulates sphingolipid metabolism and contributes to the host-defense response (Abhyankar et. al., 2018).

Signalling mechanisms regulate disease progression in a Drosophila model for Amyotrophic Lateral Sclerosis 8 (ALS8).  ALS8 is caused by a missense mutation in VAPB, an ER resident protein that maintains membrane contact sites.  A fly model for ALS involves a mutant form of Drosophila VAP (VAPALS8). The disease model shows robust VAPALS8 inclusions in the brain, has a shortened lifespan and show progressive motor deterioration with age. A reverse genetic screen identified a Gene Regulatory Network (GRN) for VAP (Deivasigamani et al., 2014) that included the TOR pathway and orthologs of other ALS genes as genetic interactors. A detailed study of one such interaction between SOD1ALS1 and VAPALS8 has led to the discovery that increase in ROS levels and TOR signalling triggers the clearance of VAPALS8 inclusions in the brain, via the proteasome (Chaplot et. al., 2019). A recent study exploring the interaction between VAPALS8 and TER94ALS14 shows that Caspar, a negative regulator of IMD/NFkB signalling acts as an adaptor, forming the VAPB:Caspar:TER94 complex. This complex, possibly acting in glial PM:ER contact sites modulate progression of the disease (Tendulkar et. al., 2021), by regulating age-dependent inflammation. Recently, we characterised age-dependent modulation of aggregate dynamics in the adult brain and found that aggregate density/size does not change dramatically with age (Thulasidharan et al., 2024), further, TER94/VCP – an ALS/FTD locus (ALS14) can modulate the extent of aggregation (Thulasidharan et al., 2024); and finally, there appears to be a switch between proteasomal clearance to autophagy between the larval and adult brain (Thulasidharan et al., 2024). 

Mon1 is a modulator of cell:cell signalling. In collaboration with Dr. Anuradha Ratnaparkhi, ARI, Pune, we have been involved in understanding Mon1-regulated signalling in the nervous system. Mon1 is a component of the Rab7 GEF complex. At the NMJ, Mon1 and Rab7 interact pre-synaptically to regulate post-synaptic receptor levels underscoring the role of transsynaptic anterograde signalling mechanisms in receptor regulation (Deivasigamani et. al., 2015; Basargekar et. al., 2020). In the context of the adult brain, Mon1 in octopaminergic-tyraminergic neurons (OPNs), which synapse with Insulin-producing cells (IPCs), regulate the release of insulin, which in turn controls the vitellogenic checkpoint in ovary maturation (Dhiman et. al., 2019). Finally, in multidendritic neurons, Mon1 regulates the complexity of dendritic arbors and tiling via a process involving the recycling pathway (Krishnan et. al., 2019).

         Future work in the laboratory is directed towards organ:organ signalling in the context of neurodegenerative disease. Specifically, we will be exploring roles of inflammation in the progression of disease.