Mechanistic Insight into Site-Restricted Monoubiquitination of FANCD2 by Ube2t, FANCL, and FANCI

• Published in: Molecular cell 2008-12, Vol.32 (6), p.767-777
• Main Authors: Alpi, Arno F., Pace, Paul E., Babu, M. Madan, Patel, Ketan J.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Cell Line; Chickens; Conserved Sequence; DNA; Fanconi Anemia Complementation Group D2 Protein - metabolism; Fanconi Anemia Complementation Group L Protein - chemistry; Fanconi Anemia Complementation Group L Protein - metabolism; Fanconi Anemia Complementation Group Proteins - metabolism; Humans; HUMDISEASE; Molecular Sequence Data; Protein Structure, Tertiary; PROTEINS; Substrate Specificity; Ubiquitin-Conjugating Enzymes - metabolism; Ubiquitination
• ISSN: 1097-2765; 1097-4164
• DOI: 10.1016/j.molcel.2008.12.003

A key step in the Fanconi anemia (FA) tumor suppressor pathway is the site-specific monoubiquitination of the FANCD2 protein. Genetic studies indicate that this crucial modification requires eight known FA gene products and the E2-conjugating enzyme Ube2t. Here, we minimally reconstitute this monoubiquitination reaction with Ube2t and the FANCL protein, revealing that monoubiquitination is stimulated by a conserved RWD-like domain in FANCL. Furthermore, addition of the FANCI protein enhances monoubiquitination and also restricts it to the in vivo substrate lysine residue on FANCD2. This work therefore establishes a system that provides mechanistic insight into the functions of FANCL and FANCI in the catalysis of FANCD2 monoubiquitination.