Structural basis for the Rad6 activation by the Bre1 N-terminal domain
Curation statements for this article:-
Curated by eLife
eLife assessment
This fundamental study presents a crystal structure of two proteins catalyzing histone H2B ubiquitination. Findings from the structural study are further validated by mutagenesis and functional assays. This is a well-executed study providing useful information to the field.
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Biochemistry and Chemical Biology (eLife)
Abstract
The mono-ubiquitination of the histone protein H2B (H2Bub1) is a highly conserved histone post-translational modification that plays critical roles in many fundamental processes. In yeast, this modification is catalyzed by the conserved Bre1–Rad6 complex. Bre1 contains a unique N-terminal Rad6-binding domain (RBD), how it interacts with Rad6 and contributes to the H2Bub1 catalysis is unclear. Here, we present crystal structure of the Bre1 RBD–Rad6 complex and structure-guided functional studies. Our structure provides a detailed picture of the interaction between the dimeric Bre1 RBD and a single Rad6 molecule. We further found that the interaction stimulates Rad6’s enzymatic activity by allosterically increasing its active site accessibility and likely contribute to the H2Bub1 catalysis through additional mechanisms. In line with these important functions, we found that the interaction is crucial for multiple H2Bub1-regulated processes. Our study provides molecular insights into the H2Bub1 catalysis.
Article activity feed
-
-
eLife assessment
This fundamental study presents a crystal structure of two proteins catalyzing histone H2B ubiquitination. Findings from the structural study are further validated by mutagenesis and functional assays. This is a well-executed study providing useful information to the field.
-
Reviewer #1 (Public Review):
The manuscript by Shi et al reports a crystal structure of partial Rad6 from K lactis in complex with Bre1 RBD domain. The structure provides detailed interactions between these two proteins, which are validated by mutagenesis and functional studies. Overall, this is a well-executed study with information useful for the histone ubiquitin field.
-
Reviewer #2 (Public Review):
The X-ray crystal structure of the K. lactis Rad6-Bre1 interaction solved by Shi et al. adds an important piece to the puzzle of how H2B mono-ubiquitination is deposited. The primary strength of this work is the new structural information on the Rad6-Bre1 interaction, which reveals contacts of the E3 (Bre1) to the backside of the E2 (Rad6). Through mutagenesis and biochemical experiments, Shi et al. probe the importance of these contacts for the Rad6-Bre1 interaction, active site accessibility, and Rad6 catalytic activity. In general, the functional data support the structural model and confirm the importance of Bre1 in stimulating Rad6 catalytic activity in vitro and H2Bub1 in yeast. In comparison to the structural data, some of the functional data are not as robust and are at times over-interpreted. …
Reviewer #2 (Public Review):
The X-ray crystal structure of the K. lactis Rad6-Bre1 interaction solved by Shi et al. adds an important piece to the puzzle of how H2B mono-ubiquitination is deposited. The primary strength of this work is the new structural information on the Rad6-Bre1 interaction, which reveals contacts of the E3 (Bre1) to the backside of the E2 (Rad6). Through mutagenesis and biochemical experiments, Shi et al. probe the importance of these contacts for the Rad6-Bre1 interaction, active site accessibility, and Rad6 catalytic activity. In general, the functional data support the structural model and confirm the importance of Bre1 in stimulating Rad6 catalytic activity in vitro and H2Bub1 in yeast. In comparison to the structural data, some of the functional data are not as robust and are at times over-interpreted. However, in general, the conclusions drawn by the authors about the importance of the newly revealed Rad6-Bre1 interface are appropriate and substantiated by the data.
-
Reviewer #3 (Public Review):
Meng, Shi et al determined the crystal structure of the Bre1 RBD-Rad6 complex from Kluyveromyces lactis and found that RBD forms an asymmetric dimer binding to a single Rad6 molecule. Subsequently, the author confirmed the binding mode of RBD-Rad6 complex by structure-based mutagenesis. They show that the binding of Bre1-RBD to Rad6 is important for both Rad6-mediated ubiquitin chain production and ubiquitin discharging of the E2~ubiquitin conjugate. In addition, they show that the interaction between Bre1 RBD and Rad6 is crucial for Bre1-mediated H2B mono-ubiquitination or homologous recombination repair inside the cell.
This study presents a useful finding on the mechanism of Bre1/Rad6-mediated ubiquitination and the conclusions of this paper are mostly well supported by data, but some aspects of claims …
Reviewer #3 (Public Review):
Meng, Shi et al determined the crystal structure of the Bre1 RBD-Rad6 complex from Kluyveromyces lactis and found that RBD forms an asymmetric dimer binding to a single Rad6 molecule. Subsequently, the author confirmed the binding mode of RBD-Rad6 complex by structure-based mutagenesis. They show that the binding of Bre1-RBD to Rad6 is important for both Rad6-mediated ubiquitin chain production and ubiquitin discharging of the E2~ubiquitin conjugate. In addition, they show that the interaction between Bre1 RBD and Rad6 is crucial for Bre1-mediated H2B mono-ubiquitination or homologous recombination repair inside the cell.
This study presents a useful finding on the mechanism of Bre1/Rad6-mediated ubiquitination and the conclusions of this paper are mostly well supported by data, but some aspects of claims need to be clarified and extended.
-
