USP51 deubiquitylates H2AK13,15ub and regulates DNA damage response

Wang, Zhi Quan; Zhang, Honglian; Liu, Ji; Cheruiyot, Abigael; Lee, Jeong Heon; Ördög, Tamás; Lou, Zhenkun; You, Zhipeng; Zhang, Zhiguo

doi:10.1101/gad.271841.115

Cited by 83 publications

(77 citation statements)

References 46 publications

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“…The three USP51 constructs were expressed in sf9 insect cells, as previously reported, and purified by His‐tagged Ni 2+ ‐affinity chromatography, the final enzymes were characterized by SDS‐PAGE (Figure S11). Consistent with previous work, the N‐terminal His‐EGFP tag could be readily cleaved by protease to afford the native full‐length USP51.…”

Section: Resultsmentioning

confidence: 97%

“…However, protease treatment of the other two constructs, His‐EGFP‐USP51(191–711) and His‐EGFP‐USP51(361–711), yielded mixtures of truncated USP51 and EGFP that were difficult to separate by size‐exclusion chromatography (SEC). In some previous studies on DUBs, it was shown that the EGFP tag did not affect their deubiquitylating activities . Therefore, we used purified USP51(FL), EFGP‐USP51(191–711), and EGFP‐USP51(361–711) in the deubiquitylation assays against the nucleosomes containing different synthetic ubiquitylated histones.…”

Section: Resultsmentioning

confidence: 97%

“…This observation is consistent with the conclusion of Zhang et al. that USP51(FL) can cleave H2AK13 and H2AK15 ubiquitylated nucleosomes . Next, if the N‐terminal Pro‐rich disordered domain was truncated, a similar amount of H2A formation was observed to that of USP51(FL).…”

Section: Resultsmentioning

confidence: 97%

“…The structural basis for this selective DUB process was recently revealed by reference to the crystal structure of this DUB module bound to H2BK120Ub‐containing nucleosomes . Moreover, another histone‐associated DUB, USP51, was recently found to be able to cleave H2A with ubiquitylation at Lys13/15 (H2AK13/15Ub) in vivo, but not at Lys119 . This selectivity is interesting because USP51 is highly homologous with the catalytic subunit (i.e., USP22) of the SAGA DUB module and can compete with USP22 for two other subunits (ATXN7L3 and ENY2) of the SAGA DUB module to hydrolyze H2BK120Ub …”

Section: Introductionmentioning

confidence: 99%

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Examination of the Deubiquitylation Site Selectivity of USP51 by Using Chemically Synthesized Ubiquitylated Histones

Guo

Sun

et al. 2018

ChemBioChem

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Histone ubiquitylation and deubiquitylation processes and the mechanisms of their regulation are closely relevant to the field of epigenetics. Recently, the deubiquitylating enzyme USP51 was reported to selectively cleave ubiquitylation on histone H2A at K13 or K15 (i.e., H2AK13Ub and H2AK15Ub), but not at K119 (i.e., H2AK119Ub), in nucleosomes in vivo. To elucidate the mechanism for the selectivity of USP51, we constructed structurally well-defined in vitro protein systems with a ubiquitin modification at precise sites. A total chemical protein synthesis procedure was developed, wherein hydrazide-based native chemical ligation was used to efficiently generate five ubiquitylated histones (H2AK13Ub, H2AK15Ub, H2AK119Ub, H2BK34Ub, and H2BK120Ub). These synthetic ubiquitylated histones were assembled into nucleosomes and subjected to in vitro USP51 deubiquitylation assays. Surprisingly, USP51 did not show preference between H2AK13/15Ub and H2AK119Ub, in contrast to previous in vivo observations. Accordingly, an understanding of the selectivity of USP51 may require consideration of other factors, such as alternative pre-existing histone modifications, competitive reader proteins, or different nucleosome quality among the in vivo extraction nucleosome and the in vitro reconstitution one. Further experiments established that USP51 in vitro could deubiquitylate a nucleosome carrying H2BK120Ub, but not H2BK34Ub. Molecular dynamics simulations suggested that USP51-catalyzed hydrolysis of ubiquitylated nucleosomes was affected by steric hindrance of the isopeptide bond.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Examination of the Deubiquitylation Site Selectivity of USP51 by Using Chemically Synthesized Ubiquitylated Histones

Guo

Sun

et al. 2018

ChemBioChem

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“…Antibodies used for Western blot analysis were anti-ubiquitin (Cell Signaling P4D1, 1:1000 dilution), anti-H2A (Millipore 07146, 1:1000 dilution), anti-K15 ubiquitylated H2A (a gift from Dr. Zhiguo Zhang, 1:500 dilution) (Wang et al, 2016), anti-GST (Santa Cruz sc-138, 1:1000 dilution), anti-mouse HRP-conjugated (Cell Signaling 7076, 1:10,000 dilution) and anti-rabbit HRP-conjugated (BioRad 172-1019, 1:10,000 dilution).…”

Section: Methods Detailsmentioning

confidence: 99%

Mechanisms of Ubiquitin-Nucleosome Recognition and Regulation of 53BP1 Chromatin Recruitment by RNF168/169 and RAD18

et al. 2017

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Summary The protein 53BP1 plays a central regulatory role in DNA double-strand break repair. 53BP1 relocates to chromatin by recognizing RNF168-mediated mono-ubiquitylation of histone H2A Lys15 in the nucleosome core particle dimethylated at histone H4 Lys20 (NCP-ubme). 53BP1 relocation is terminated by ubiquitin ligases RNF169 and RAD18 via unknown mechanisms. Using NMR spectroscopy and biochemistry, we show that RNF169 bridges ubiquitin and histone surfaces, stabilizing a pre-existing ubiquitin orientation in NCP-ubme to form a high-affinity complex. This conformational selection mechanism contrasts with the low-affinity binding mode of 53BP1 and ensures 53BP1 displacement by RNF169 from NCP-ubme. We also show that RAD18 binds tightly to NCP-ubme through a ubiquitin-binding domain that contacts ubiquitin and nucleosome surfaces accessed by 53BP1. Our work uncovers diverse ubiquitin recognition mechanisms in the nucleosome, explaining how RNF168, RNF169 and RAD18 regulate 53BP1 chromatin recruitment and how specificity can be achieved in the recognition of a ubiquitin-modified substrate.

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USP51/PD‐L1/ITGB1‐deployed juxtacrine interaction plays a cell‐intrinsic role in promoting chemoresistant phenotypes in non‐small cell lung cancer

Xiao

et al. 2023

Cancer Communications

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Background Programmed death ligand 1 (PD‐L1) has been demonstrated to facilitate tumor progression and therapeutic resistance in an immune‐independent manner. Nevertheless, the function and underlying signaling network(s) of cancer cell‐intrinsic PD‐L1 action remain largely unknown. Herein, we sought to better understand how ubiquitin‐specific peptidase 51 (USP51)/PD‐L1/integrin beta‐1 (ITGB1) signaling performs a cell‐intrinsic role in mediating chemotherapeutic resistance in non‐small cell lung cancer (NSCLC). Methods Western blotting and flow cytometry were employed for PD‐L1 detection in NSCLC cell lines. Coimmunoprecipitation and pulldown analyses, protein deubiquitination assay, tissue microarray, bioinformatic analysis and molecular biology methods were then used to determine the significance of PD‐L1 in NSCLC chemoresistance and associated signaling pathways in several different cell lines, mouse models and patient tissue samples. Ubiquitin‐7‐amido‐4‐methylcoumarin (Ub‐AMC)‐based deubiquitinase activity, cellular thermal shift and surface plasmon resonance (SPR) analyses were performed to investigate the activity of USP51 inhibitors. Results We provided evidence that cancer cell‐intrinsic PD‐L1 conferred the development of chemoresistance by directly binding to its membrane‐bound receptor ITGB1 in NSCLC. At the molecular level, PD‐L1/ITGB1 interaction subsequently activated the nuclear factor‐kappa B (NF‐κB) axis to elicit poor response to chemotherapy. We further determined USP51 as a bona fide deubiquitinase that targeted the deubiquitination and stabilization of the PD‐L1 protein in chemoresistant NSCLC cells. Clinically, we found a significant direct relationship between the USP51, PD‐L1 and ITGB1 contents in NSCLC patients with chemoresistant potency. The elevated USP51, PD‐L1 and ITGB1 levels were strongly associated with worse patient prognosis. Of note, we identified that a flavonoid compound dihydromyricetin (DHM) acted as a potential USP51 inhibitor and rendered NSCLC cells more sensitive to chemotherapy by targeting USP51‐dependent PD‐L1 ubiquitination and degradation in vitro and in vivo. Conclusions Together, our results demonstrated that the USP51/PD‐L1/ITGB1 network potentially contributes to the malignant progression and therapeutic resistance in NSCLC. This knowledge is beneficial to the future design of advanced cancer therapy.

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USP51 deubiquitylates H2AK13,15ub and regulates DNA damage response

Cited by 83 publications

References 46 publications

Examination of the Deubiquitylation Site Selectivity of USP51 by Using Chemically Synthesized Ubiquitylated Histones

Examination of the Deubiquitylation Site Selectivity of USP51 by Using Chemically Synthesized Ubiquitylated Histones

Mechanisms of Ubiquitin-Nucleosome Recognition and Regulation of 53BP1 Chromatin Recruitment by RNF168/169 and RAD18

USP51/PD‐L1/ITGB1‐deployed juxtacrine interaction plays a cell‐intrinsic role in promoting chemoresistant phenotypes in non‐small cell lung cancer

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