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Moraes, Trevor F.; Edwards, Ross A.; McKenna, Sean Andrew; Pastushok, Landon; Xiao, Wei; Glover, J. N. Mark; Ellison, M.

doi:10.1038/90373

Cited by 139 publications

(95 citation statements)

References 28 publications

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“…Recently, McKenna et al (33) showed that in human cells, hMms2 forms a complex with hUbc13, and this interaction is required for hUbc13-mediated polyubiquitination through Lys-63. The crystal structure of this complex also was determined recently (34). Our data support the hypothesis that the hMms2͞hUbc13 complex is required to recruit or activate proteins needed by human cells to be able to make use of an undamaged homologous copy of DNA as an alternative template to avoid fork-blocking lesions and continue DNA replication.…”

Section: Indirect Evidence That Human Cells Use the Newly Synthesizedsupporting

confidence: 70%

Identification of a protein essential for a major pathway used by human cells to avoid UV- induced DNA damage

Xiao

McCormick

et al. 2002

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

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When DNA replication stalls at a fork-blocking lesion, cells use damage tolerance pathways to continue replication. One pathway, ''translesion synthesis,'' involves specialized DNA polymerases that can use damaged DNA as a template. Translesion synthesis can result in mutations (i.e., can be error-prone), but it can also be error-free. An alternative pathway has been hypothesized (sometimes called ''damage avoidance''), by which cells make temporary use of an undamaged copy of the blocked sequence as a template, i.e., the newly synthesized daughter strand of the sister duplex or the allelic copy. This pathway is error-free. Evidence of the use of the daughter strand of the sister duplex as a template in intact mammalian cells has not been available heretofore. To determine whether hMms2, a ubiquitin-conjugating enzyme-like protein, plays a critical role in such damage avoidance, a human fibroblast cell strain in which both error-prone translesion synthesis and error-free damage avoidance can be detected and quantified simultaneously, and several derivative strains in which expression of hMms2 protein had been eliminated or greatly decreased, were compared for their ability to avoid translesion synthesis past UV 254nm-induced DNA photoproducts. Loss of hMms2 protein eliminated the ability of the latter strains to use an allelic copy of a target gene for damage avoidance, i.e., to produce a wild-type gene from two nonfunctional allelic copies of that gene. Molecular analysis of the wild-type gene showed that this process involves gene conversion unassociated with crossing-over. That the loss of hMms2 also eliminated use of the daughter strand of the sister duplex as a template for damage avoidance could be inferred from the fact that the frequency of mutations induced by UV in the single copy HPRT gene of the derivative strains was significantly higher than that observed in the parental strain. These data indicate that hMMS2 is essential for human cells to carry out damage avoidance by using either type of homolog, and that damage avoidance and translesion synthesis are alternative pathways for tolerating fork-blocking photoproducts. D NA is constantly exposed to damaging agents. If the damage is not removed, e.g., by excision repair, before the onset of S-phase, certain kinds of lesions can block replication by the major DNA polymerase complex (1). The damage tolerance mechanisms developed by prokaryotic and eukaryotic cells to overcome such replication blocks fall into two categories: translesion synthesis and damage avoidance. Evidence suggests that translesion synthesis is a process in which specialized, distributive DNA polymerases take over for the major DNA polymerase complex to carry out DNA replication by using the damaged DNA as a template (2-5). After distributive incorporation of nucleotides past the damage, the major DNA replication complex resumes its replication activity. Translesion synthesis can be either ''error-prone'' or ''error-free,'' depending on such factors as the type of damage, its seque...

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Section: Indirect Evidence That Human Cells Use the Newly Synthesizedsupporting

confidence: 70%

Identification of a protein essential for a major pathway used by human cells to avoid UV- induced DNA damage

Xiao

McCormick

et al. 2002

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

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“…Recently, the high-resolution x-ray crystal structures of both the human (45) and S. cerevisiae (38) Ubc13/Mms2 heterodimer have been solved, and both structures propose models that could accommodate the active site tethered and noncovalently bound Ub molecules. In combination with the data presented in this manuscript, these structures establish a foundation for the assembly of Lys-63 chains by the Ubc13/Mms2 heterodimer.…”

Section: Discussionmentioning

confidence: 99%

Noncovalent Interaction between Ubiquitin and the Human DNA Repair Protein Mms2 Is Required for Ubc13-mediated Polyubiquitination

McKenna¹,

Spyracopoulos²,

Moraes³

et al. 2001

Journal of Biological Chemistry

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127

160

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Ubiquitin-conjugating enzyme variants share significant sequence similarity with typical E2 (ubiquitin-conjugating) enzymes of the protein ubiquitination pathway but lack their characteristic active site cysteine residue. The MMS2 gene of Saccharomyces cerevisiae encodes one such ubiquitin-conjugating enzyme variant that is involved in the error-free DNA postreplicative repair pathway through its association with Ubc13, an E2. The Mms2-Ubc13 heterodimer is capable of linking ubiquitin molecules to one another through an isopeptide bond between the C terminus and Lys-63. Using highly purified components, we show here that the human forms of Mms2 and Ubc13 associate into a heterodimer that is stable over a range of conditions. The ubiquitin-thiol ester form of the heterodimer can be produced by the direct activation of its Ubc13 subunit with E1 (ubiquitin-activating enzyme) or by the association of Mms2 with the Ubc13-ubiquitin thiol ester. The activated heterodimer is capable of transferring its covalently bound ubiquitin to Lys-63 of an untethered ubiquitin molecule, resulting in diubiquitin as the predominant species. In 1 H 15 N HSQC ( 1 H 15 N heteronuclear single quantum coherence) NMR experiments, we have mapped the surface determinants of tethered and untethered ubiquitin that interact with Mms2 and Ubc13 in both their monomeric and dimeric forms. These results have identified a surface of untethered ubiquitin that interacts with Mms2 in the monomeric and heterodimeric form. Furthermore, the C-terminal tail of ubiquitin does not participate in this interaction. These results suggest that the role of Mms2 is to correctly orient either a target-bound or untethered ubiquitin molecule such that its Lys-63 is placed proximally to the C terminus of the ubiquitin molecule that is linked to the active site of Ubc13.

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“…The UEV Domains Share a Common Fold but Use This Fold to Make Diverse Interactions-The structure of the Vps23 UEV domain has an overall fold that is similar to the two other structurally characterized UEV domains, human Tsg101 (54) and Mms2 (56,57) (Fig. 5, A and B).…”

Section: Both Unique and Commonly Exploited Ub Surfaces Interact Withmentioning

confidence: 99%

“…5B; Refs. 56,57). Similarly, the S1-S2 loop of Ubc13 interacts with Mms2, and the bound Mms2 covers a surface of Ubc13 analogous to the Vps23 UEV surface interacting with Ub (Fig.…”

Section: Both Unique and Commonly Exploited Ub Surfaces Interact Withmentioning

confidence: 99%

Structural Insights into Endosomal Sorting Complex Required for Transport (ESCRT-I) Recognition of Ubiquitinated Proteins

Teo

Veprintsev

Williams

2004

Journal of Biological Chemistry

121

101

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The endosomal sorting complex required for transport (ESCRT-I) is a 350-kDa complex of three proteins, Vps23, Vps28, and Vps37. The N-terminal ubiquitin-conjugating enzyme E2 variant (UEV) domain of Vps23 is required for sorting ubiquitinated proteins into the internal vesicles of multivesicular bodies. UEVs are homologous to E2 ubiquitin ligases but lack the conserved cysteine residue required for catalytic activity. The crystal structure of the yeast Vps23 UEV in a complex with ubiquitin (Ub) shows the detailed interactions made with the bound Ub. Compared with the solution structure of the Tsg101 UEV (the human homologue of Vps23) in the absence of Ub, two loops that are conserved among the ESCRT-I UEVs move toward each other to grip the Ub in a pincer-like grasp. The contacts with the UEV encompass two adjacent patches on the surface of the Ub, one containing several hydrophobic residues, including Ile-8 Ub , Ile-44 Ub , and Val-70 Ub , and the second containing a hydrophilic patch including residues Asn-60 Ub , Gln-62 Ub , Glu-64 Ub . The hydrophobic Ub patch interacting with the Vps23 UEV overlaps the surface of Ub interacting with the Vps27 ubiquitin-interacting motif, suggesting a sequential model for ubiquitinated cargo binding by these proteins. In contrast, the hydrophilic patch encompasses residues uniquely interacting with the ESCRT-I UEV. The structure provides a detailed framework for design of mutants that can specifically affect ESCRT-I-dependent sorting of ubiquitinated cargo without affecting Vps27-mediated delivery of cargo to endosomes.Ubiquitination serves as a general mechanism for regulating protein localization and destiny in eukaryotic cells. This mechanism involves conjugation of the 76-residue polypeptide ubiquitin (Ub) 1 to the ⑀-amino group of lysine residues on target proteins. The conjugation occurs by means of a sequential series of thiolester-bound intermediates involving the E1, E2, and E3 enzymes and results in proteasomal, nuclear, and endosomal targeting, depending upon the protein and the nature of the ubiquitination. Whereas proteasomal targeting requires attachment of a chain of at least four ubiquitins linked by isopeptide bonds between the C terminus of one Ub and Lys-48 of the next (1, 2), mono-ubiquitination is sufficient for endosomal targeting. Ubiquitination regulates internalization and/or

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Untitled

Cited by 139 publications

References 28 publications

Identification of a protein essential for a major pathway used by human cells to avoid UV- induced DNA damage

Identification of a protein essential for a major pathway used by human cells to avoid UV- induced DNA damage

Noncovalent Interaction between Ubiquitin and the Human DNA Repair Protein Mms2 Is Required for Ubc13-mediated Polyubiquitination

Structural Insights into Endosomal Sorting Complex Required for Transport (ESCRT-I) Recognition of Ubiquitinated Proteins

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