Mechanism and regulation of V(D)J recombination mediated by Recombination Activation Genes RAG1 and RAG2

Wednesday, October 10, 2012 — Poster Session II
Noon – 2:00 p.m	Natcher Conference Center, Building 45	NIDDK	MOLBIO-9

Authors

• SK Singh
• W Yang
• M Gellert

Abstract

V(D)J recombination catalyzed by Recombination Activation Gene proteins RAG1-RAG2 is the process which give rise to variations of B and T cell receptors. The regulatory functions of the N-terminal end of RAG1 and the C-terminal end of RAG2 have not been fully studied because until now the full length proteins were difficult to express and purify. We succeeded in expressing and purifying the mouse full length RAG1-RAG2. Simultaneously, for comparative studies we also expressed and purified various deletion mutants of these complexes. The complexes were completely active in catalyzing RSS cleavage. We also discovered the conditions for ubiquitination of full length RAG1. The ubiquitination of FLRAG1 results in enhancement of cleavage activity. Previous studies show that RAG2 ubiquitination leads to degradation of the protein, hence from this study it can be suggested that RAG1 ubiquitination leads to positive regulation while RAG2 ubiquitination leads to negative regulation. We also found that full length complex can transpose signal ends into other DNA. This study would be the first to address cleavage activity mediated by the purified full length RAG complexes and their regulation by protein modification, which may open a new window for investigating the role of these biologically essential proteins in-vivo.

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