Human SHPRH is a ubiquitin ligase for Mms2–Ubc13-dependent polyubiquitylation of proliferating cell nuclear antigen

Unk, Ildikó; Fátyol, Károly; Szakál, Barnabás; Blastyák, András; Bermudez, Vladimir P.; Hurwitz, Jerard; Prakash, Louise; Prakash, Satya; Haracska, Lajos

doi:10.1073/pnas.0608595103

Cited by 212 publications

(196 citation statements)

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“…On the other hand, SHPRH has an H1 and H5 linker sequence and a PHD domain not present in either Rad5 or HLTF. Because SHPRH can also promote the polyubiquitination of PCNA (12,13), it will be important to examine the overlaps and differences between the functions of HLTF and SHPRH, suggested by their domain structures.…”

Section: Discussionmentioning

confidence: 99%

“…cDNAs of human PCNA and human Rad18 were cloned into pRK2H and pDSREDC1, resulting in pIL1460 and pIL1263, respectively. Human Rad6 -Rad18, Mms2-Ubc13, RFC, and ubiquitin-activating enzymes were purified as described (13). GST-HLTF was expressed in yeast and purified on glutathione-Sepharose beads.…”

Section: Methodsmentioning

confidence: 99%

“…Next, we investigated whether HLTF can catalyze the polyubiquitination of PCNA in vitro by using highly purified proteins. We have already shown that for PCNA monoubiquitination to occur by Rad6-Rad18, PCNA first has to be loaded onto DNA by replication factor C (RFC) (13,15). Therefore, first we preincubated PCNA with RFC and DNA followed by the addition of ubiquitin, Uba1, and various combinations of HLTF, Rad6-Rad18, and Mms2-Ubc13 (Fig.…”

Section: Hltf Functions As a Ubiquitin Ligase For K63-linked Polyubiqmentioning

confidence: 99%

“…Previously, we and others have shown that human SHPRH, which resembles Rad5 in having a SWI/SNF helicase domain and a C 3 HC 4 RING motif, exhibits a ubiquitin ligase activity able to promote PCNA polyubiquitination in a Rad6-Rad18-and Mms2-Ubc13-dependent manner (12,13).…”

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Human HLTF functions as a ubiquitin ligase for proliferating cell nuclear antigen polyubiquitination

Unk

Hajdú

Fátyol

et al. 2008

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

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221

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Human helicase-like transcription factor (HLTF) is frequently inactivated in colorectal and gastric cancers. Here, we show that HLTF is a functional homologue of yeast Rad5 that promotes error-free replication through DNA lesions. HLTF and Rad5 share the same unique structural features, including a RING domain embedded within a SWI/SNF helicase domain and an HIRAN domain. We find that inactivation of HLTF renders human cells sensitive to UV and other DNA-damaging agents and that HLTF complements the UV sensitivity of a rad5⌬ yeast strain. Also, similar to Rad5, HLTF physically interacts with the Rad6 -Rad18 and Mms2-Ubc13 ubiquitin-conjugating enzyme complexes and promotes the Lys-63-linked polyubiquitination of proliferating cell nuclear antigen at its Lys-164 residue. A requirement of HLTF for error-free postreplication repair of damaged DNA is in keeping with its cancersuppression role.yeast Rad5 ͉ damage bypass ͉ K63 polyubiquitination ͉ tumor suppressor L esions in DNA impose a block to synthesis by the replicative polymerases (Pols), and unless replication is rescued by the timely action of lesion bypass processes, stalled replication forks can collapse, leading to genomic instability. In eukaryotes the Rad6-Rad18 enzyme complex regulates lesion bypass processes that ensure the completion of replication. Rad6, a ubiquitinconjugating enzyme, forms a tight complex with Rad18, a RING-finger type ubiquitin ligase that binds DNA (1, 2), and in cells treated with DNA-damaging agents, Rad6-Rad18 monoubiquitinates proliferating cell nuclear antigen (PCNA), a DNA Pol sliding clamp that is a key component of the replication machinery (3). Ubiquitination at the Lys-164 residue of PCNA and its subsequent polyubiquitination serves as a molecular switch between various DNA damage bypass processes (3-5).In the yeast Saccharomyces cerevisiae, Rad6-Rad18 governs at least three alternative pathways for promoting replication through DNA lesions (6). Two pathways activated by PCNA monoubiquitination are carried out by specialized translesion synthesis (TLS) DNA Pols, such as Pol and Pol , which are able to copy DNA directly from the damaged template on an error-free or error-prone way (6). The third pathway called postreplication repair (PRR), however, is activated by PCNA polyubiquitination and operates by template switching using the information of the undamaged newly synthesized nascent strand on the sister duplex for DNA synthesis across damaged DNA (7-10). The PRR pathway depends on the Rad5, Mms2, and Ubc13 proteins and promotes error-free replication through DNA lesions. Recently, we have shown that yeast Rad5 has a DNA helicase activity that is specialized for replication fork regression, as Rad5 can concertedly unwind and anneal the nascent and the parental strands of the fork without exposing any single-stranded regions (7). This Rad5 activity would ensure damage bypass by promoting replication fork regression where the newly synthesized DNA strand of the sister duplex can be used as a template. In addition to its r...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Hltf Functions As a Ubiquitin Ligase For K63-linked Polyubiqmentioning

confidence: 99%

mentioning

confidence: 99%

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Human HLTF functions as a ubiquitin ligase for proliferating cell nuclear antigen polyubiquitination

Unk

Hajdú

Fátyol

et al. 2008

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

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“…Regardless of the mode of reaction, it is abundantly clear that the error-free DDT process is highly conserved in the entire eukaryotic kingdom, from yeast to human. Sequence and functional homologs of all proteins involved in error-free DDT, including Mms2 [140], Ubc13 [141] and Rad5 [142,143], have been found in mammals, plants and other higher eukaryotes [144,145]. For a few limited examples, suppression of the above genes resulted in phenotypes reminiscent of the corresponding yeast mutants [143,146,147].…”

Section: Error-free Ddtmentioning

confidence: 99%

Eukaryotic DNA damage tolerance and translesion synthesis through covalent modifications of PCNA

Andersen¹,

Xiao³

2007

Cell Res

185

187

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npg In addition to well-defined DNA repair pathways, all living organisms have evolved mechanisms to avoid cell death caused by replication fork collapse at a site where replication is blocked due to disruptive covalent modifications of DNA. The term DNA damage tolerance (DDT) has been employed loosely to include a collection of mechanisms by which cells survive replication-blocking lesions with or without associated genomic instability. Recent genetic analyses indicate that DDT in eukaryotes, from yeast to human, consists of two parallel pathways with one being error-free and another highly mutagenic. Interestingly, in budding yeast, these two pathways are mediated by sequential modifications of the proliferating cell nuclear antigen (PCNA) by two ubiquitination complexes Rad6-Rad18 and Mms2-Ubc13-Rad5. Damage-induced monoubiquitination of PCNA by Rad6-Rad18 promotes translesion synthesis (TLS) with increased mutagenesis, while subsequent polyubiquitination of PCNA at the same K164 residue by Mms2-Ubc13-Rad5 promotes error-free lesion bypass. Data obtained from recent studies suggest that the above mechanisms are conserved in higher eukaryotes. In particular, mammals contain multiple specialized TLS polymerases. Defects in one of the TLS polymerases have been linked to genomic instability and cancer. DNA damage toleranceIn the presence of spontaneous or carcinogen-induced DNA damage, living cells have to maintain and complete DNA synthesis or risk replication fork collapse. Since the process of DNA licensing is to ensure the genome is duplicated once and only once during each cell cycle, stalled or collapsed replication forks may not be able to restart, which often results in double-strand breaks (DSBs) and causes compromised genome integrity or cell death. In addition to highly conserved DNA repair pathways, all living organisms have evolved schemes to ensure continuation of DNA synthesis in the presence of damage. These schemes were originally termed DNA postreplication repair (PRR) due to observations of transient shortened nascent DNA structures following S phase in response to DNA damage. In bacteria and unicellular yeast, these shortened DNA segments can be measured by an alkaline sedimentation assay [1] or directly observed in electron micrographs [2]. In wild-type cells, these truncated DNA segments were restored to full length following a short recovery time. One typical experiment [1] involved the restoration of the nascent strand following UV exposure in nucleotide excision repair (NER)-deficient cells and was originally assumed to represent a mechanism of DNA repair. However, further investigation revealed that, although the nascent fragments were re-annealed, the original UV-induced pyrimidine dimers, which were responsible for the generation of single-strand gaps, often persisted in the genome [3,4]. It was argued that the replication-blocking lesion was not necessarily corrected, but rather transiently bypassed and carried over to the next generation. Perhaps it is more beneficial for the organi...

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The Small Open Reading Frame‐Encoded Peptides: Advances in Methodologies and Functional Studies

et al. 2021

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Small open reading frames (sORFs) are an important class of genes with less than 100 codons. They were historically annotated as noncoding or even junk sequences. In recent years, accumulating evidence suggests that sORFs could encode a considerable number of polypeptides, many of which play important roles in both physiology and disease pathology. However, it has been technically challenging to directly detect sORF‐encoded peptides (SEPs). Here, we discuss the latest advances in methodologies for identifying SEPs with mass spectrometry, as well as the progress on functional studies of SEPs.

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Human SHPRH is a ubiquitin ligase for Mms2–Ubc13-dependent polyubiquitylation of proliferating cell nuclear antigen

Cited by 212 publications

References 36 publications

Human HLTF functions as a ubiquitin ligase for proliferating cell nuclear antigen polyubiquitination

Human HLTF functions as a ubiquitin ligase for proliferating cell nuclear antigen polyubiquitination

Eukaryotic DNA damage tolerance and translesion synthesis through covalent modifications of PCNA

The Small Open Reading Frame‐Encoded Peptides: Advances in Methodologies and Functional Studies

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