The Diversity of Ubiquitin Recognition: Hot Spots and Varied Specificity

Winget, Jason M.; Mayor, Thibault

doi:10.1016/j.molcel.2010.05.003

Cited by 144 publications

(162 citation statements)

References 70 publications

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“…Ub12 is a quadruple alanine mutant where the leucinearginine-leucine-arginine motif near the C terminus of ubiquitin has been changed. This region is critical for ubiquitin recognition by the ubiquitylation machinery, displays structural plasticity, and has been implicated in many known ubiquitin interactions (17,18,30,31).…”

Section: A C-terminal Ubiquitin Patch Is Critical For Hdot1l Stimulatmentioning

confidence: 99%

“…Thus, compared with smaller PTMs such as acetylation and methylation, ubiquitin is "information rich" in that it alters the steric and electrostatic properties around its attachment site, as well as presenting a large surface area for the recruitment of binding factors. Structural and biochemical studies of ubiquitin-ligand complexes, including the ubiquitylation of histone H2A at lysine 15, have revealed a canonical binding hotspot on ubiquitin involving a hydrophobic patch centered on Leu8/Ile44 (15, 16), however additional interaction surfaces have been extensively characterized (17,18). It is currently unclear whether the chromatin-associated functions of ubiquitin-for instance, the stimulation of Set1 and Dot1 activities-operate through a single functional epitope or whether discrete surfaces of ubiquitin are involved.…”

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confidence: 99%

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Identification of a functional hotspot on ubiquitin required for stimulation of methyltransferase activity on chromatin

Holt

David

Pollock

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Ubiquitylation of histone H2B at lysine 120 (H2B-Ub) plays a critical role in transcriptional elongation, chromatin conformation, as well as the regulation of specific histone H3 methylations. Herein, we report a strategy for the site-specific chemical attachment of ubiquitin to preassembled nucleosomes. This allowed expedited structure-activity studies into how H2B-Ub regulates H3K79 methylation by the methyltransferase human Dot1. Through an alanine scan of the ubiquitin surface, we identified a functional hotspot on ubiquitin that is required for the stimulation of human Dot1 in vitro. Importantly, this result was validated in chromatin from isolated nuclei by using a synthetic biology strategy that allowed selective incorporation of the hotspot-deficient ubiquitin mutant into H2B. The ubiquitin hotspot additionally impacted the regulation of ySet1-mediated H3K4 methylation but was not required for H2B-Ub-induced impairment of chromatin fiber compaction. These data demonstrate the utility of applying chemical ligation technologies to preassembled chromatin and delineate the multifunctionality of ubiquitin as a histone posttranslational modification.H istone posttranslational modifications (PTMs) modulate chromatin structure and function either by directly altering the intrinsic physical properties of the chromatin fiber or by nucleating the recruitment and activity of a host of transacting nuclear factors (1-3). The chemical diversity, differential dynamics, and sheer number (currently over 100) (4, 5) of these PTMs, along with their combinatorial occurrence at the level of the nucleosome, create a complex and nonstatic molecular architecture in which all chromatin-related processes function. A central challenge in the field of epigenetics is to disentangle how distinct chromatin states control biochemical outputs, which requires the elucidation of the critical determinants governing histone PTM readout.A particularly fascinating histone PTM is the ubiquitylation of H2B at lysine 120 (H2B-Ub). H2B-Ub is enriched near the 5′ end of highly expressed genes and has been implicated in transcriptional elongation, as well as chromatin structure definition (6-9). Moreover, H2B-Ub directly regulates the H3K4 and H3K79 methyltransferases Set1 and Dot1, respectively (10-13). The mechanistic principles underlying these various phenomena remain poorly understood. At 8.5 kDa, ubiquitin is nearly as large as the histone to which it is linked (13.8 kDa in the case of H2B), increasing the nucleosome surface by as much as 4,800 A 2 (14). Thus, compared with smaller PTMs such as acetylation and methylation, ubiquitin is "information rich" in that it alters the steric and electrostatic properties around its attachment site, as well as presenting a large surface area for the recruitment of binding factors. Structural and biochemical studies of ubiquitin-ligand complexes, including the ubiquitylation of histone H2A at lysine 15, have revealed a canonical binding hotspot on ubiquitin involving a hydrophobic patch centered on Leu8/I...

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Section: A C-terminal Ubiquitin Patch Is Critical For Hdot1l Stimulatmentioning

confidence: 99%

mentioning

confidence: 99%

Identification of a functional hotspot on ubiquitin required for stimulation of methyltransferase activity on chromatin

Holt

David

Pollock

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Comparing many different complexes of ␤-grasp fold proteins highlights that the complete surface of the domain can act as a binding site (21)(22)(23). However, there are interaction hot spots like the exposed bottom side, strand ␤2, and the ␣/␤ groove between strand ␤2 and the single helix (24). The SUMO1⅐Daxx20 complex represents one example where the SUMOinteracting motif of Daxx20 binds into the ␣/␤ groove of SUMO-1 in an extended conformation (25).…”

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confidence: 99%

MET-activating Residues in the B-repeat of the Listeria monocytogenes Invasion Protein InlB

Bleymüller

Lämmermann

Ebbes

et al. 2016

Journal of Biological Chemistry

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Edited by Norma AllewellThe facultative intracellular pathogen Listeria monocytogenes causes listeriosis, a rare but life-threatening disease. Host cell entry begins with activation of the human receptor tyrosine kinase MET through the bacterial invasion protein InlB, which contains an internalin domain, a B-repeat, and three GW domains. The internalin domain is known to bind MET, but no interaction partner is known for the B-repeat. Adding the B-repeat to the internalin domain potentiates MET activation and is required to stimulate Madin-Darby canine kidney (MDCK) cell scatter. Therefore, it has been hypothesized that the B-repeat may bind a co-receptor on host cells. To test this hypothesis, we mutated residues that might be important for binding an interaction partner. We identified two adjacent residues in strand ␤2 of the ␤-grasp fold whose mutation abrogated induction of MDCK cell scatter. Biophysical analysis indicated that these mutations do not alter protein structure. We then tested these mutants in human HT-29 cells that, in contrast to the MDCK cells, were responsive to the internalin domain alone. These assays revealed a dominant negative effect, reducing the activity of a construct of the internalin domain and mutated B-repeat below that of the individual internalin domain. Phosphorylation assays of MET and its downstream targets AKT and ERK confirmed the dominant negative effect. Attempts to identify a host cell receptor for the B-repeat were not successful. We conclude that there is limited support for a co-receptor hypothesis and instead suggest that the B-repeat contributes to MET activation through low affinity homodimerization.

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“…Among the PIPs are a group of ubiquitin-like (UBL) domaincontaining proteins, including the substrate shuttle proteins Rad23A/B and Dsk2/PLIC, the E3 ubiquitin ligase Parkin, and the deubiquitinating enzyme Ubp6/USP14 (9,10). Despite low similarity in amino acid sequence, UBL domains and ubiquitin (Ub) share a similar core structure known as the β-grasp fold and are recognized by a variety of ubiquitin/UBL-binding proteins, including Rpn1, 10, and 13 of the 19S RP (11)(12)(13)(14)(15)(16).…”

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confidence: 99%

UBLCP1 is a 26S proteasome phosphatase that regulates nuclear proteasome activity

Guo

Engel

Xiao

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Protein degradation by the 26S proteasome is a fundamental process involved in a broad range of cellular activities, yet how proteasome activity is regulated remains poorly understood. We report here that ubiquitin-like domain-containing C-terminal domain phosphatase 1 (UBLCP1) is a 26S proteasome phosphatase that regulates nuclear proteasome activity. UBLCP1 directly interacts with the proteasome via its UBL domain and is exclusively localized in the nucleus. UBLCP1 dephosphorylates the 26S proteasome and inhibits proteasome activity in vitro. Knockdown of UBLCP1 in cells promotes 26S proteasome assembly and selectively enhances nuclear proteasome activity. Our results describe the first identified proteasome-specific phosphatase and uncover a unique mechanism for phosphoregulation of the proteasome.

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The Diversity of Ubiquitin Recognition: Hot Spots and Varied Specificity

Cited by 144 publications

References 70 publications

Identification of a functional hotspot on ubiquitin required for stimulation of methyltransferase activity on chromatin

Identification of a functional hotspot on ubiquitin required for stimulation of methyltransferase activity on chromatin

MET-activating Residues in the B-repeat of the Listeria monocytogenes Invasion Protein InlB

UBLCP1 is a 26S proteasome phosphatase that regulates nuclear proteasome activity

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