RNF168 ubiquitinates K13-15 on H2A/H2AX to drive DNA damage signaling

Ubiquitin-dependent signaling during the DNA damage response (DDR) to double-strand breaks (DSBs) is initiated by two E3 ligases, RNF8 and RNF168, targeting histone H2A and H2AX. RNF8 is the first ligase recruited to the damage site, and RNF168 follows RNF8-dependent ubiquitination. This suggests th...

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Published in:Cell Vol. 150; no. 6; p. 1182
Main Authors: Mattiroli, Francesca, Vissers, Joseph H A, van Dijk, Willem J, Ikpa, Pauline, Citterio, Elisabetta, Vermeulen, Wim, Marteijn, Jurgen A, Sixma, Titia K
Format: Journal Article
Language:English
Published: United States 14.09.2012
Subjects:
Amino Acid Sequence
DNA-Binding Proteins - chemistry
DNA-Binding Proteins - metabolism
Histones - chemistry
Histones - metabolism
Humans
Lysine - metabolism
Models, Molecular
Molecular Sequence Data
Nucleosomes - chemistry
Nucleosomes - metabolism
Protein Structure, Tertiary
Scattering, Small Angle
Ubiquitin-Protein Ligases - chemistry
Ubiquitin-Protein Ligases - metabolism
X-Ray Diffraction
ISSN:1097-4172, 1097-4172
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Summary:Ubiquitin-dependent signaling during the DNA damage response (DDR) to double-strand breaks (DSBs) is initiated by two E3 ligases, RNF8 and RNF168, targeting histone H2A and H2AX. RNF8 is the first ligase recruited to the damage site, and RNF168 follows RNF8-dependent ubiquitination. This suggests that RNF8 initiates H2A/H2AX ubiquitination with K63-linked ubiquitin chains and RNF168 extends them. Here, we show that RNF8 is inactive toward nucleosomal H2A, whereas RNF168 catalyzes the monoubiquitination of the histones specifically on K13-15. Structure-based mutagenesis of RNF8 and RNF168 RING domains shows that a charged residue determines whether nucleosomal proteins are recognized. We find that K63 ubiquitin chains are conjugated to RNF168-dependent H2A/H2AX monoubiquitination at K13-15 and not on K118-119. Using a mutant of RNF168 unable to target histones but still catalyzing ubiquitin chains at DSBs, we show that ubiquitin chains per se are insufficient for signaling, but RNF168 target ubiquitination is required for DDR.
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ISSN:1097-4172
1097-4172
DOI:10.1016/j.cell.2012.08.005