A unique E1-E2 interaction required for optimal conjugation of the ubiquitin-like protein NEDD8

Huang, Danny T.; Miller, David W.; Mathew, Rose; Cassell, Robert J.; Holton, James M.; Roussel, Martine F.; Schulman, Brenda A.

doi:10.1038/nsmb826

Cited by 132 publications

(120 citation statements)

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“…In plants, RUB is activated by an E1-like heterodimer of E1-CONJUGATING ENZYME-RELATED1 (ECR1; orthologous to human UBA3) and AUXIN RESISTANT1 (AXR1; orthologous to human APPBP1; del Pozo et al, 1998), then transferred to the E2-like RUB1-CONJUGATING ENZYME1 (RCE1; orthologous to human UBC12; del Pozo and Gray et al, 2002;Huang et al, 2004). RUB transfer to the CUL1 target protein is facilitated by the RING-H2 protein RBX1/ROC .…”

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confidence: 99%

Mutation ofE1-CONJUGATING ENZYME-RELATED1Decreases RELATED TO UBIQUITIN Conjugation and Alters Auxin Response and Development

et al. 2007

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The ubiquitin-like protein RELATED TO UBIQUITIN (RUB) is conjugated to CULLIN (CUL) proteins to modulate the activity of Skp1-Cullin-F-box (SCF) ubiquitylation complexes. RUB conjugation to specific target proteins is necessary for the development of many organisms, including Arabidopsis (Arabidopsis thaliana). Here, we report the isolation and characterization of e1-conjugating enzyme-related1-1 (ecr1-1), an Arabidopsis mutant compromised in RUB conjugation. The ecr1-1 mutation causes a missense change located two amino acid residues from the catalytic site cysteine, which normally functions to form a thioester bond with activated RUB. A higher ratio of unmodified CUL1 relative to CUL1-RUB is present in ecr1-1 compared to wild type, suggesting that the mutation reduces ECR1 function. The ecr1-1 mutant is resistant to the auxin-like compound indole-3-propionic acid, produces fewer lateral roots than wild type, displays reduced adult height, and stabilizes a reporter fusion protein that is degraded in response to auxin, suggesting reduced auxin signaling in the mutant. In addition, ecr1-1 hypocotyls fail to elongate normally when seedlings are grown in darkness, a phenotype shared with certain other RUB conjugation mutants that is not general to auxin-response mutants. The suite of ecr1-1 molecular and morphological phenotypes reflects roles for RUB conjugation in many aspects of plant growth and development. Certain ecr1-1 elongation defects are restored by treatment with the ethylene-response inhibitor silver nitrate, suggesting that the short ecr1-1 root and hypocotyl result from aberrant ethylene accumulation. Further, silver nitrate supplementation in combination with various auxins and auxin-like compounds reveals that members of this growth regulator family may differentially rely on ethylene signaling to inhibit root growth.

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confidence: 99%

Mutation ofE1-CONJUGATING ENZYME-RELATED1Decreases RELATED TO UBIQUITIN Conjugation and Alters Auxin Response and Development

et al. 2007

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“…Despite these striking similarities, Rub1/Nedd8 and Ub employ their own cognate E1 and E2 to post-translationally modify their specific substrates (22,23). The recognition of the cognate E2s by their respective E1 paralogues is critical in avoiding cross-talk between these two parallel regulatory pathways (24,25). The Ub pathway involves the attachment of monomeric Ub or polyUb chains to substrates and has been implicated in degradative and regulatory or protein sorting/trafficking functions.…”

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confidence: 99%

Recognition and Cleavage of Related to Ubiquitin 1 (Rub1) and Rub1-Ubiquitin Chains by Components of the Ubiquitin-Proteasome System

Singh

Zerath

Kleifeld

et al. 2012

Molecular & Cellular Proteomics

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Of all ubiquitin-like proteins, Rub1 (Nedd8 in mammals) is the closest kin of ubiquitin. We show via NMR that structurally, Rub1 and ubiquitin are fundamentally similar as well. Despite these profound similarities, the prevalence of Rub1/Nedd8 and of ubiquitin as modifiers of the proteome is starkly different, and their attachments to specific substrates perform different functions. Recently, some proteins, including p53, p73, EGFR, caspase-7, and Parkin, have been shown to be modified by both Rub1/ Nedd8 and ubiquitin within cells. To understand whether and how it might be possible to distinguish among the same target protein modified by Rub1 or ubiquitin or both, we examined whether ubiquitin receptors can differentiate between Rub1 and ubiquitin. Surprisingly, Rub1 interacts with proteasome ubiquitin-shuttle proteins comparably to ubiquitin but binds more weakly to a proteasomal ubiquitin receptor Rpn10. We identified Rub1-ubiquitin heteromers in yeast and Nedd8-Ub heteromers in human cells. We validate that in human cells and in vitro, human Rub1 (Nedd8) forms chains with ubiquitin where it acts as a chain terminator. Interestingly, enzymatically assembled K48-linked Rub1-ubiquitin heterodimers are recognized by various proteasomal ubiquitin shuttles and receptors comparably to K48-linked ubiquitin homodimers. Furthermore, these heterologous chains are cleaved by COP9 signalosome or 26S proteasome. A derubylation function of the proteasome expands the repertoire of its enzymatic activities. In contrast, Rub1 conjugates may be somewhat resilient to the actions of other canonical deubiquitinating enzymes. Taken together, these findings suggest that once Rub1/Nedd8 is channeled into ubiquitin pathways, it is recognized essentially like The selective degradation of many proteins in eukaryotic cells is a highly specific and irreversible process required to perform vital cellular functions such as cell cycle progression (1, 2), differentiation and development (3, 4), and transcriptional control (5). One of the major pathways involved in this selective degradation is the ubiquitin-proteasome pathway. In this pathway, ubiquitin (Ub), a 76-amino-acid protein, is attached to the substrate, and this is followed by the recognition and degradation of the substrate by a protein complex collectively known as proteasome (6 -8).The attachment of Ub (ubiquitination) to a substrate protein is achieved via a cascade of enzymatic reactions (involving E1, E2, and E3 enzymes) that results in the formation of an isopeptide bond between the C-terminal glycine of Ub and a lysine residue of the substrate (9 -11). Sequential repetition of this cascade can result in the attachment of a chain of Ub molecules (polyubiquitin) to the substrate protein. The length and topology of the polyubiquitin (polyUb) tag decide the fate of the substrate protein. For example, we and others have shown that a K48-linked tetraubiquitin (tetraUb) chain that acts as a proteasomal degradation signal forms a "closed" structure (12, 13), whereas a K63-linked Ub...

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“…Along with other E1 structures, [85][86][87] our studies of the E1 for NEDD8, alone and in complexes with NEDD8, MgATP, and with NEDD8 E2s provided a structural framework for understanding many E1 functions. 61,[88][89][90][91][92][93] The structures revealed three independently-folded globular domains, each specifying distinct E1 activities: (1) an adenylation domain binds the UBL and MgATP and contains the adenylation reaction active site; (2) a distinct domain houses the catalytic cysteine that becomes covalently linked to the UBL C-terminus; and (3) a ''ubiquitin-fold domain'' (UFD) unexpectedly adopts a ubiquitin-like fold and recruits an E2. How does the E1 architecture mediate so many binding and catalytic activities?…”

Section: Dynamic Assembly-line-like Ubl Activation By E1smentioning

confidence: 99%

Twists and turns in ubiquitin‐like protein conjugation cascades

Schulman

2011

Protein Science

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Post-translational modification by ubiquitin-like proteins (UBLs) is a predominant eukaryotic regulatory mechanism. The vast reach of this form of regulation extends to virtually all eukaryotic processes that involve proteins. UBL modifications play critical roles in controlling the cell cycle, transcription, DNA repair, stress responses, signaling, immunity, plant growth, embryogenesis, circadian rhythms, and a plethora of other pathways. UBLs dynamically modulate target protein properties including enzymatic activity, conformation, half-life, subcellular localization, and intermolecular interactions. Moreover, the enzymatic process of UBL ligation to proteins is itself dynamic, with the UBL moving between multiple enzyme active sites and ultimately to a target. This review highlights our work on how the dynamic conformations of selected enzymes catalyzing UBL ligation help mediate this fascinating form of protein regulation.

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A unique E1-E2 interaction required for optimal conjugation of the ubiquitin-like protein NEDD8

Cited by 132 publications

References 51 publications

Mutation ofE1-CONJUGATING ENZYME-RELATED1Decreases RELATED TO UBIQUITIN Conjugation and Alters Auxin Response and Development

Mutation ofE1-CONJUGATING ENZYME-RELATED1Decreases RELATED TO UBIQUITIN Conjugation and Alters Auxin Response and Development

Recognition and Cleavage of Related to Ubiquitin 1 (Rub1) and Rub1-Ubiquitin Chains by Components of the Ubiquitin-Proteasome System

Twists and turns in ubiquitin‐like protein conjugation cascades

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