Protein RuvB-like 1: the biological relevance of putative arginine methylation in Leishmania braziliensis

• Grant number: 20/14059-0
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: January 01, 2021
• End date: December 31, 2022
• Field of knowledge: Biological Sciences - Parasitology - Protozoology of Parasites
• Principal Investigator: Angela Kaysel Cruz
• Grantee: Mariana Loterio Silva
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated research grant: 18/14398-0 - UK:Brazil Joint Centre Partnership in Leishmaniasis (JCPiL), AP.TEM

Abstract

The protozoan parasite Leishmania, besides its medical relevance, has peculiar genetic features including the polycistronic nature of the transcriptional process and the implicated differences on regulation of gene expression, that happens mainly at the posttranscriptional level. Thus, RNA Binding Proteins play a central role in modulating gene expression in these organisms; the interaction of these proteins with an mRNA may affect its stability, fate into the cell or the translation initiation rates. The protein interactions with a given mRNA may change depending on their abundance and other factors, such as post-translational modifications. Arginine methylation, one of these modifications, is catalyzed by Protein Arginine Methyltransferases (PRMTs). The PRMTs are subject of study in our laboratory and in previous work a protein annotated as RuvB-like 1 has been identified as a potential target of PRMT5 in Leishmania braziliensis (LbrPRMT5). Our goals are to investigate whether the arginine residues in the typical RG domains are methylated in RuvB-like and to characterize the effect of the arginine methylation on protein stability and intracellular localization. Levels and intracellular localization will be evaluated across developmental stages; procyclic promastigotes, metacyclic promastigotes and amastigotes. The study will be conducted using parasites with tagged versions of the endogenous RuvB-like; the protein will be tagged in the wild type and PRMT5 knockout L. braziliensis, already available. Modified versions of the protein, in which the arginines (in the RG domain) will be substituted by lysine or glutamate (mimetics of hypo and hypermethylated RuvB-like), will be generated by site-direct mutagenesis. Finally, the tagged proteins in different genetic background (wild type and PRMT5 knockout cells) will be used to rescue interacting proteins.