Mechanism of histone H2B monoubiquitination by Bre1

Zhao, Fan; Hicks, Chad; Wolberger, Cynthia

doi:10.1101/2023.03.27.534461

Cited by 2 publications

References 28 publications

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Structure of the human Bre1 complex bound to the nucleosome

Onishi,

Uchiyama,

Sato

et al. 2024

Nat Commun

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Histone H2B monoubiquitination (at Lys120 in humans) regulates transcription elongation and DNA repair. In humans, H2B monoubiquitination is catalyzed by the heterodimeric Bre1 complex composed of Bre1A/RNF20 and Bre1B/RNF40. The Bre1 proteins generally function as tumor suppressors, while in certain cancers, they facilitate cancer cell proliferation. To obtain structural insights of H2BK120 ubiquitination and its regulation, we report the cryo-electron microscopy structure of the human Bre1 complex bound to the nucleosome. The two RING domains of Bre1A and Bre1B recognize the acidic patch and the nucleosomal DNA phosphates around SHL 6.0–6.5, which are ideally located to recruit the E2 enzyme and ubiquitin for H2BK120-specific ubiquitination. Mutational experiments suggest that the two RING domains bind in two orientations and that ubiquitination occurs when Bre1A binds to the acidic patch. Our results provide insights into the H2BK120-specific ubiquitination by the Bre1 proteins and suggest that H2B monoubiquitination can be regulated by nuclesomal DNA flexibility.

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Structure of the human Bre1 complex bound to the nucleosome

Onishi,

Uchiyama,

Sato

et al. 2024

Nat Commun

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Structural mechanisms of autoinhibition and substrate recognition by the ubiquitin ligase HACE1

Duering,

Wolter,

Toplak

et al. 2023

Preprint

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Ubiquitin ligases (E3s) are pivotal specificity determinants in the ubiquitin system by selecting substrates and decorating them with distinct ubiquitin signals. Structure determination of the underlying, specific E3-substrate complexes, however, has proven challenging due to their transient nature. In particular, it is incompletely understood how members of the catalytic cysteine-driven class of HECT-type ligases position substrate proteins for modification. Here we report a cryo-EM structure of the full-length human HECT-type ligase HACE1, along with solution-based conformational analyses by small-angle X-ray scattering and hydrogen-deuterium exchange mass spectrometry. Structure-based functional analyses in vitro and in cells reveal that the activity of HACE1 is stringently regulated by dimerization-induced autoinhibition. The inhibition occurs at the first step of the catalytic cycle and is thus substrate-independent. We employ mechanism-based chemical crosslinking to reconstitute a complex of activated, monomeric HACE1 with its major substrate, RAC1, visualize its structure by cryo-EM, and validate the binding mode by solution-based analyses. Our findings explain how HACE1 achieves selectivity in ubiquitinating the active, GTP-loaded state of RAC1 and establish a framework for interpreting mutational alterations of the HACE1-RAC1 interplay in disease. More broadly, this work illuminates central unexplored aspects in the architecture, conformational dynamics, regulation, and specificity of full-length HECT-type ligases.

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Mechanism of histone H2B monoubiquitination by Bre1

Cited by 2 publications

References 28 publications

Structure of the human Bre1 complex bound to the nucleosome

Structure of the human Bre1 complex bound to the nucleosome

Structural mechanisms of autoinhibition and substrate recognition by the ubiquitin ligase HACE1

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