Functional studies of SECIS elements in Trypanossomatideos

• Grant number: 11/20863-8
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: May 01, 2012
• End date: July 31, 2013
• Field of knowledge: Biological Sciences - Biochemistry - Biochemistry of Microorganisms
• Principal Investigator: Otavio Henrique Thiemann
• Grantee: Júlia Assirati
• Host Institution: Instituto de Física de São Carlos (IFSC). Universidade de São Paulo (USP). São Carlos , SP, Brazil
• Associated research grant: 98/14138-2 - Center for Structural Molecular Biotechnology, AP.CEPID

Abstract

The traditional genetic code consisting of 20 amino acids now includes selenocysteine and pirrolisina, resulting in its expansion to a total of 22 amino acids. The 21st amino acid selenocysteine (Sec - U), discovered as amino acid incorporated into selenoproteins in 1989, represents the major biological form of selenium and has a specific synthesis pathway in eukaryotes / archaebacteria and other bacteria. Their incorporation is dependent on the presence of an in phase UGA stop codon and a messenger RNA tertiary structure known as SECIS element (Sec Incorporation Sequence). Our group has identified the existence of the pathway and its physiological role in Kinetoplastida. In Kinetoplastida protozoa three selenoproteins were identified, SelK, and SelT SelTryp, the last has no homology with mammalian selenoproteins. Sec is incorporated as one of the last amino acids in SelK and SelTryp respectively, and at position 108 to 259 amino acids in SelT. Comparative analysis of the location dispersion of Sec and SECIS sites among variants in other organisms we were motivated to further investigate this trend in Kinetoplastida. In this project we will study the insertion of Sec in vivo using the reporter gene GFP (Green Fluorescent Protein) containing mutations Cys-Sec and T. brucei SECIS elements These constructs should allow to verify the influence of the position of UGA-Sec codon and SECIS element and test variants of SECIS T. brucei by identifying the minimal SECIS structural determinants for insertion of Sec. We hope that the data generated here will contribute to the characterization of the process of Sec insertion and the development of a tool for future analysis and validation of SECIS elements and selenoproteins in vivo. (AU)