Functional characterization of Leishmania amazonensis overexpressing RPA-1 (LaRPA-1)

• Grant number: 14/19622-4
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: November 01, 2014
• End date: October 31, 2015
• Field of knowledge: Biological Sciences - Genetics - Molecular Genetics and Genetics of Microorganisms
• Principal Investigator: Maria Isabel Nogueira Cano
• Grantee: Gabriel Arantes Galvão Dias dos Santos
• Host Institution: Instituto de Biociências (IBB). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil

Abstract

Leishmaniasis is a neglected disease affecting millions of people around the world, including Brazil. This zoonosis is caused by protozoan of the genus Leishmania to which there is no treatment and control. Therefore, efforts for the establishment of intensive research to better understand the molecular and cellular biology of these parasites may provide new strategies to treat and to eradicate the disease. A possible strategy is the elucidation of the mechanisms involved in DNA metabolism and telomere maintenance by studying for example, the roles played by Replication Protein A subunit 1 (RPA-1), a single-stranded DNA-binding protein involved in many eukaryotic DNA processing pathways, including telomere maintenance and DNA damage signaling. RPA-1 is present in most eukaryotes including Leishmania spp., although protozoa RPA-1 (LaRPA-1) presents special structural features that are associated with cell viability, regulation of the DNA repair machinery, telomerase recruitment and telomeres maintenance. This study aims to functionally characterize parasites overexpressing LaRPA-1. We intend to clone the population by serial dilution and test its proliferation in growth curves, protein expression and subcellular localization using western blot and indirect immunofluorescence, and telomere length by southern blot. All these assays will be performed in parallel with wild type populations and with a cloned population transfected only with the empty vector. Our goal is to understand the roles played by Leishmania amazonensis RPA-1 (LaRPA-1) in telomere maintenance.