Nov 9, 2005

Methods for the functional genomic analysis of ubiquitin ligases

Methods in Enzymology
Wei LiClaudio A P Joazeiro

Abstract

Ubiquitin ligases (E3s) are critical components of the ubiquitin-proteasome system as they are the major determinants of specificity in ubiquitin conjugation. The number of predicted E3s in the mammalian genome is exceeding 400 and is represented by two major subfamilies: HECT domain-containing E3s and RING finger-type E3s. Given the size of this protein family and lack of knowledge on the functions of most of these 400 proteins, their functional annotation should benefit from modern genomic tools. This article presents a methodology consisting of the use of a cDNA expression library to identify suppressors of polyglutamine (polyQ)-mediated protein aggregate formation in cells, as an example of a genomic approach to assign functions to E3s. In this screen, we identified novel RING finger-type E3s exhibiting suppressor activity among >50% of all the potential E3s in the mouse and human genomes. This method could be adapted easily to identify E3s that function in other processes and signaling pathways.

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Mentioned in this Paper

Ring Finger Domain
Biochemical Pathway
Proteasome Pathway
Conjugation
Transfection
Protein Family
Genome
Enzymes, antithrombotic
RNA, Small Interfering
Polyglutamine

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