PhD Student, Savitribai Phule Pune University
Drosophila melanogaster, commonly known as fruit fly shares many similarities with higher organisms like humans at the genetic and molecular level. They can be easily maintained in lab conditions, easily manipulated and thus provide an excellent tool to study processes like aging and immunity. My area of work focuses to find out whether losing an enzyme has any impact on the immune system of these flies. After losing the gene (thus making them null) of the enzyme, named methyltransferase, flies do not show any dramatic effect in their life span when grown without any stress. But when these flies are grown at 29 degrees or when they are injected with bacteria show a reduced life span when compared with wild type flies under same conditions. Interestingly when I infect 15 day old null flies the observed effect becomes more dramatic. 15 day old wild type is better in handling stress (infection and temperature) than null fly. This work assigns a novel role to the methyltransferase, which was not discovered in the earlier studies. Fruit fly only bares one gene for methyltransferase enzyme unlike other organisms like mouse or humans thus making it a simple system to unravel the functional aspect of this enzyme. I’ll be presenting these results at the 25th European Drosophila Research Conference, where many eminent scientists and enthusiastic researchers will also showcase their work. People who are working in the field will give inputs and suggestions to my work which might give a new direction and can help me understand the mechanism of aging and immunity in a great detail.
Abstract: Drosophila melanogaster lacks DNMT1/DNMT3 based methylation machinery. Despite recent reports confirming the presence of low DNA methylation in Drosophila; little is known about the methyltransferase. Therefore, in this study, we have aimed to investigate the possible functioning of DNA methyltransferase in Drosophila. The 14 K oligo microarray slide was incubated with native cell extract from adult Drosophila to check the presence of the methyltransferase activity. After incubation under appropriate conditions, the methylated oligo sequences were identified by the binding of anti 5-methylcytosine monoclonal antibody. The antibody bound to the methylated oligos was detected using Cy3 labeled secondary antibody. Methylation sensitive restriction enzyme mediated PCR was used to assess the methylation at a few selected loci identified on the array. It could be seen that a few of the total oligos got methylated under the assay conditions. Analysis of methylated oligo sequences provides evidence for the presence of de novo methyltransferase activity and allows identification of its sequence specificity in adult Drosophila. With the help of methylation sensitive enzymes we could detect presence of CpC methylation in the selected genomic regions. This study reports presence of an active DNA methyltransferase in adult Drosophila, which exhibits sequence specificity confirmed by presence of asymmetric methylation at corresponding sites in the genomic DNA. It also provides an innovative approach to investigate methylation specificity of a native methyltransferase.
Pub.: 09 Nov '15, Pinned: 18 Aug '17
Abstract: Drosophila use small-interfering RNA mechanisms to limit the amplification of viral genomes. However, it is unclear how small RNA interference components recognize and separate viral from cellular RNA. Dnmt2 enzymes are highly conserved RNA methyltransferases with substrate specificity towards cellular tRNAs. We report here that Dnmt2 is required for efficient innate immune responses in Drosophila. Dnmt2 mutant flies accumulate increasing levels of Drosophila C virus and show activated innate immune responses. Binding of Dnmt2 to DCV RNA suggests that Dnmt2 contributes to virus control directly, possibly by RNA methylation. These observations demonstrate a role for Dnmt2 in antiviral defence.
Pub.: 02 Feb '13, Pinned: 18 Aug '17