Controlling the subcellular localization of DNA polymerases ι and η via interactions with ubiquitin

Plosky, Brian S.; Vidal, Antonio E.; Henestrosa, Antonio R. Fernández de; McLenigan, Mary P.; McDonald, John P.; Mead, Samantha; Woodgate, Roger

doi:10.1038/sj.emboj.7601178

Cited by 192 publications

(203 citation statements)

References 48 publications

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“…To examine the role of UBDs in PCNA binding, we have focused our studies on the UBZ domain of hPolη because evidence from studies with mutations of conserved residues in this domain have been used extensively in support of the model (10,12,17). In a previous study, we showed that hPolη has two PIP motifs in its C terminus, which can substitute for one another and that mutational inactivation of both the PIP motifs renders hPolη nonfunctional in human cells, as the protein can no longer localize onto PCNA in UV irradiated cells and is unable to complement the UV sensitivity of XPV cells (13).…”

Section: Discussionmentioning

confidence: 99%

“…Genetic studies in yeast have shown that PCNA ubiquitylation is a necessary prerequisite for TLS (8,9); however, it still remains unclear how this PCNA modification modulates the access of TLS Pols to PCNA stalled at a lesion site. Because TLS Pols such as η, ι, and κ contain an ubiquitin-binding domain (UBD) in addition to a PCNA-interacting protein (PIP) domain, a view that continues to be generally accepted is that TLS Pols bind to PCNA not only at its interconnector loop domain via their PIP domain, but also to the lys 164-linked ubiquitin moiety via their UBD domain, and that the binding of ubiquitin (Ub) on PCNA is crucial for the TLS Pols to function in lesion bypass (10)(11)(12). To determine the relative significance of the PIP and UBD domains in PCNA binding, in a previous study we examined the roles of these domains in human Polη.…”

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confidence: 99%

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DNA polymerase η lacking the ubiquitin-binding domain promotes replicative lesion bypass in humans cells

Acharya

Yoon

Hurwitz

et al. 2010

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

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The Rad6-Rad18 mediated monoubiquitylation of proliferating cell nuclear antigen (PCNA) at lys 164 plays a crucial role in promoting the access of translesion synthesis (TLS) DNA polymerases (Pols) to PCNA in the replication fork stalled at a lesion site. Although a number of genetic and biochemical observations have provided strong evidence that TLS Pols bind PCNA at its interdomain connector loop (IDCL) via their PCNA-interacting protein (PIP) domain, a more recent proposal formulates that TLS Pols bind PCNA at two sites, to the IDCL via their PIP domain and to lys-164 linked ubiquitin (Ub) via their ubiquitin-binding domain. To ascertain the relative contributions of the PIP and Ub-binding zinc finger (UBZ) domains of human Polη in TLS, we have determined whether the C-terminal truncations of hPolη that contain the PIP1 domain but lack the UBZ and PIP2 domains can still function in TLS in human cells. Our observations that such C-terminally truncated proteins promote efficient TLS opposite a cis-syn TT dimer and confer a high degree of UV resistance to XPV cells provide unambiguous evidence that the binding of PCNA via its PIP domain is essential as well as sufficient for providing hPolη the ability to carry out TLS in human cells.human Pol eta | proliferating cell nuclear antigen ubiquitylation | translesion DNA synthesis | ubiquitin-binding zinc finger domain

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

DNA polymerase η lacking the ubiquitin-binding domain promotes replicative lesion bypass in humans cells

Acharya

Yoon

Hurwitz

et al. 2010

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

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“…One mechanism by which the UBZ domain may allow these proteins to participate in DNA repair is suggested by the observation that for RAD18, Pol, and Pol, the UBZ domain is both sufficient and necessary for correct localization to DNA damage-induced nuclear foci (37,56,57). The UBZ domain may therefore function in recruitment to repair sites upon a DNA damage-specific ubiquitin-mediated signal.…”

Section: Peptidementioning

confidence: 99%

Werner Helicase-interacting Protein 1 Binds Polyubiquitin via Its Zinc Finger Domain

Bish¹,

Myers

2007

Journal of Biological Chemistry

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DNA repair is regulated on many levels by ubiquitination. In order to identify novel connections between DNA repair pathways and ubiquitin signaling, we used mass spectrometry to identify proteins that interact with lysine 6-linked polyubiquitin chains. From this proteomic screen, we identified the DNA repair protein WRNIP1 (Werner helicase-interacting protein 1), along with nucleosome assembly protein 1, as novel ubiquitin-interacting proteins. We found that a small zinc finger domain at the N terminus of WRNIP1 is sufficient and necessary for noncovalent ubiquitin binding. This ubiquitin-binding zinc finger (UBZ) domain binds polyubiquitin but not monoubiquitin and appears to show no specificity for polyubiquitin chain linkage. A homologous zinc finger domain in RAD18 also binds polyubiquitin, suggesting a wider role for the UBZ domain in DNA repair. The WRNIP1 ubiquitin-binding function, along with its previously established ATPase activity, suggests that WRNIP1 plays a role in the metabolism of ubiquitinated proteins. Supporting this model, deletion of MGS1, the yeast homolog of WRNIP1, slows the rate of ubiquitin turnover, rendering yeast resistant to cycloheximide. We also find that WRNIP1 is heavily modified with ubiquitin and SUMO, revealing complex layers in the involvement of ubiquitin pathway proteins in the regulation of DNA repair. The novel ubiquitin-binding ability of WRNIP1 sheds light on the role of UBZ domain-containing proteins in postreplication DNA repair.

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“…The UBZs Are Required for Enhanced Association between Pol and Monoubiquitinated PCNA-Recent studies have demonstrated that monoubiquitination of PCNA in cells exposed to UV radiation promotes a more robust interaction of this accessory replication protein with Pol, Pol, and REV1 protein (10,11,25,30). To determine whether an enhanced association also exists between Pol and monoubiquitinated PCNA, we examined their interaction by GST pulldown experiments (26).…”

Section: Mouse Pol Interacts With Ubiquitin Via Two Ubiquitinbinding mentioning

confidence: 99%

Requirements for the Interaction of Mouse Polκ with Ubiquitin and Its Biological Significance

Guo

Tang

Bienko

et al. 2008

Journal of Biological Chemistry

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Pol protein is a eukaryotic member of the DinB/Pol branch of the Y-family DNA polymerases, which are involved in the tolerance of DNA damage by replicative bypass. Despite universal conservation through evolution, the precise role(s) of Pol in this process has remained unknown. Here we report that mouse Pol can physically interact with ubiquitin by yeast two-hybrid screening, glutathione S-transferase pulldown, and immunoprecipitation methods. The association of Pol with ubiquitin requires the ubiquitin-binding motifs located at the C terminus of Pol. In addition, Pol binds with monoubiquitinated proliferating cell nuclear antigen (PCNA) more robustly than with non-ubiquitinated PCNA. The ubiquitin-binding motifs mediate the enhanced association between monoubiquitinated PCNA and Pol. The ubiquitin-binding motifs are also required for Pol to form nuclear foci after UV radiation. However, the ubiquitin-binding motifs do not affect Pol half-life. Finally, we have examined levels of Pol expression following the exposure of mouse cells to benzo[a]pyrene-dihydrodiol epoxide or UVB radiation. Translesion DNA synthesis (TLS)3 is one of several biochemical mechanisms by which cells can tolerate DNA damage that arrests semiconservative DNA synthesis (1, 2). This process requires the action of specialized DNA polymerases present in bacteria (such as Escherichia coli), lower eukaryotes, and vertebrates. Lower eukaryotes, particularly vertibrates, contain multiple such enzymes, suggesting the ability to bypass many types of DNA damage.Several specialized DNA polymerases are members of a novel polymerase family, the Y-family (3). These enzymes are devoid of 3Ј 3 5Ј proofreading exonuclease activity and replicate undamaged DNA in vitro with low fidelity and weak processivity (4). Members of this family in mammalian cells include Pol, Pol, and Pol, all of which can extend primers for varying distances past various types of template damage (4). A fourth member of the Y-family, REV1 protein, is able to catalyze the incorporation of only one or two dCMP moieties, regardless of the template base composition (5). Pol, Pol, and Pol have been shown to interact with REV1 protein via a highly conserved C-terminal domain in REV1 (6 -8). These polymerases also interact with PCNA (9, 10), and recent observations suggest that PCNA plays a key role in promoting the access of specialized polymerases to arrested replication forks (11-15).Disruption of the PolK gene in mouse and chicken cells results in significant sensitivity to killing by benzo[a]pyrenedihydrodiol epoxide (BPDE) and UV radiation (16 -18). Poldeficient mouse embryonic stem and fibroblast cells also show moderate sensitivity to methyl methanesulfonate (19). Consistent with these results, primer extension assays have shown that human Pol can support TLS across sites of base loss, acetylaminofluorene-G adducts, benzo[a]pyrene-G adducts, and thymine glycol (4). However, the enzyme does not support primer extension past thymine-thymine (TϽϾT) dimers or [6,4]pyrimidine-pyrimidone p...

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Controlling the subcellular localization of DNA polymerases ι and η via interactions with ubiquitin

Cited by 192 publications

References 48 publications

DNA polymerase η lacking the ubiquitin-binding domain promotes replicative lesion bypass in humans cells

DNA polymerase η lacking the ubiquitin-binding domain promotes replicative lesion bypass in humans cells

Werner Helicase-interacting Protein 1 Binds Polyubiquitin via Its Zinc Finger Domain

Requirements for the Interaction of Mouse Polκ with Ubiquitin and Its Biological Significance

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