Human DDK rescues stalled forks and counteracts checkpoint inhibition at unfired origins to complete DNA replication

Ma, Jones; Gelot, Camille; Munk, Stephanie; Koren, Amnon; Kawasoe, Yoshitaka; George, Kelly A.; Santos, Ruth E.; Olsen, Jesper V.; McCarroll, Steven A.; Frattini, Mark G.; Takahashi, Tatsuro; Jallepalli, Prasad V.

doi:10.1016/j.molcel.2021.01.004

Cited by 25 publications

(46 citation statements)

References 101 publications

(131 reference statements)

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“…With respect to post-initiation forms of regulation by the DDK, accumulating evidence, mostly in animal cells, suggests continued DDK activity is necessary for fork advance during replication stress (Dolson et al 2021 ). Roles for the DDK appear to include regulation of fork reversals, nuclease processing, and gap filling as a means to restart stalled forks (Sasi et al 2018 ; Rainey et al 2020 ; Jones et al 2021 ; Cabello-Lobato et al 2021 ). Additionally, continued DDK phosphorylation of Mcm2-7 (Bastia et al 2016 ; Alver et al 2017 ), and potentially phosphorylation of fork pausing factors (Murakami and Keeney 2014 ), may enforce polymerase coupling at stalled forks.…”

Section: Discussionmentioning

confidence: 99%

The yeast Dbf4 Zn2+ finger domain suppresses single-stranded DNA at replication forks initiated from a subset of origins

et al. 2022

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Dbf4 is the cyclin-like subunit for the Dbf4-dependent protein kinase (DDK), required for activating the replicative helicase at DNA replication origin that fire during S phase. Dbf4 also functions as an adaptor, targeting the DDK to different groups of origins and substrates. Here we report a genome-wide analysis of origin firing in a budding yeast mutant, dbf4-zn, lacking the Zn2+ finger domain within the C-terminus of Dbf4. At one group of origins, which we call dromedaries, we observe an unanticipated DNA replication phenotype: accumulation of single-stranded DNA spanning ± 5kbp from the center of the origins. A similar accumulation of single-stranded DNA at origins occurs more globally in pri1-m4 mutants defective for the catalytic subunit of DNA primase and rad53 mutants defective for the S phase checkpoint following DNA replication stress. We propose the Dbf4 Zn2+ finger suppresses single-stranded gaps at replication forks emanating from dromedary origins. Certain origins may impose an elevated requirement for the DDK to fully initiate DNA synthesis following origin activation. Alternatively, dbf4-zn may be defective for stabilizing/restarting replication forks emanating from dromedary origins during replication stress.

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Section: Discussionmentioning

confidence: 99%

The yeast Dbf4 Zn2+ finger domain suppresses single-stranded DNA at replication forks initiated from a subset of origins

et al. 2022

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“…Studies in yeast provide substantial evidence that DDK participates in the S-phase checkpoint activation, however, it is also a target of the S phase checkpoint ( Ogi et al, 2008 ; Snaith et al, 2000 ). There are contrasting reports on the role of DDK under replication stress ( Jones et al, 2021 ; Tsuji et al, 2008 ; Yamada et al, 2014 ). Here, we report that DDK/Hsk1 phosphorylates sirtuin Hst4 in response to replication stress to target it for proteasomal degradation through SCF Pof3 ubiquitin ligase to regulate replication fork recovery.…”

Section: Discussionmentioning

confidence: 99%

“…Studies in mammalian cells have also shown that DDK kinase activity and complex formation is retained and required for protecting replication forks upon replication stress ( Lee et al, 2012 ). Most recent evidence has implicated the role of mammalian DDK in the replication fork restart following stalling of forks ( Jones et al, 2021 ). DDK prefers S/T residues close to acidic amino acids such as aspartate and glutamate for phosphorylation ( Cho et al, 2006 ; Masai and Arai, 2002 ; Masai et al, 2006 ).…”

Section: Discussionmentioning

confidence: 99%

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DDK/Hsk1 phosphorylates and targets fission yeast histone deacetylase Hst4 for degradation to stabilize stalled DNA replication forks

Aricthota

Haldar

2021

eLife

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In eukaryotes, paused replication forks are prone to collapse, which leads to genomic instability, a hallmark of cancer. Dbf4 Dependent Kinase (DDK)/Hsk1Cdc7 is a conserved replication initiator kinase with conflicting roles in replication stress response. Here, we show that fission yeast DDK/Hsk1 phosphorylates sirtuin, Hst4 upon replication stress at C-terminal serine residues. Phosphorylation of Hst4 by DDK marks it for degradation via the ubiquitin ligase SCFpof3. Phosphorylation defective hst4 mutant (4SA-hst4) displays defective recovery from replication stress, faulty fork restart, slow S-phase progression and decreased viability. The highly conserved Fork Protection Complex (FPC) stabilizes stalled replication forks. We found that the recruitment of FPC components, Swi1 and Mcl1 to the chromatin is compromised in the 4SA-hst4 mutant, although whole cell levels increased. These defects are dependent upon H3K56ac and independent of intra S-phase checkpoint activation. Finally, we show conservation of H3K56ac dependent regulation of Timeless, Tipin and And-1 in human cells. We propose that degradation of Hst4 via DDK increases H3K56ac, changing the chromatin state in the vicinity of stalled forks facilitating recruitment and function of FPC. Overall, this study identified a crucial role of DDK and FPC in the regulation of replication stress response with implications in cancer therapeutics.

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“…CHEK1 encodes the cell cycle checkpoint kinase Chk1, which modulates aging through regulating the DNA replication. [ 43 ] Chk1 is an Hsp90 client and requires Hsp90 to acquire its kinase activity. [ 44 ] Thus, targeting Chk1 alone or in combination with Hsp90 inhibitors might attenuate the development of AMD through regulating cell senescence and reducing the dose or duration of Hsp90 inhibition to lower the toxicity on the photoreceptor cells.…”

Section: Discussionmentioning

confidence: 99%

Hsp90-associated DNA replication checkpoint protein and proteasome-subunit components are involved in the age-related macular degeneration

Chen

Liu

Zhang

et al. 2021

Chinese Medical Journal

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Background: Age-related macular degeneration (AMD) is the leading cause of vision loss worldwide. However, the mechanisms involved in the development and progression of AMD are poorly delineated. We aimed to explore the critical genes involved in the progression of AMD. Methods: The differentially expressed genes (DEGs) in AMD retinal pigment epithelial (RPE)/choroid tissues were identified using the microarray datasets GSE99248 and GSE125564, which were downloaded from the gene expression omnibus database. The overlapping DEGs from the two datasets were screened to identify DEG-related biological pathways using gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses. The hub genes were identified from these DEGs through protein-protein interaction network analyses. The expression levels of hub genes were evaluated by quantitative real-time polymerase chain reaction following the induction of senescence in ARPE-19 with FK866. Following the identification of AMD-related key genes, the potential small molecule compounds targeting the key genes were predicted by PharmacoDB. Finally, a microRNA-gene interaction network was constructed. Results: Microarray analyses identified 174 DEGs in the AMD RPE compared to the healthy RPE samples. These DEGs were primarily enriched in the pathways involved in the regulation of DNA replication, cell cycle, and proteasome-mediated protein polyubiquitination. Among the top ten hub genes, HSP90AA1 , CHEK1 , PSMA4 , PSMD4 , and PSMD8 were upregulated in the senescent ARPE-19 cells. Additionally, the drugs targeting HSP90AA1, CHEK1, and PSMA4 were identified. We hypothesize that Hsa-miR-16-5p might target four out of the five key DEGs in the AMD RPE. Conclusions: Based on our findings, HSP90AA1 is likely to be a central gene controlling the DNA replication and proteasome-mediated polyubiquitination during the RPE senescence observed in the progression of AMD. Targeting HSP90AA1 , CHEK1 , PSMA4 , PSMD4 , and/or PSMD8 genes through specific miRNAs or small molecules might potentially alleviate the progression of AMD through attenuating RPE senescence.

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Human DDK rescues stalled forks and counteracts checkpoint inhibition at unfired origins to complete DNA replication

Cited by 25 publications

References 101 publications

The yeast Dbf4 Zn2+ finger domain suppresses single-stranded DNA at replication forks initiated from a subset of origins

The yeast Dbf4 Zn2+ finger domain suppresses single-stranded DNA at replication forks initiated from a subset of origins

DDK/Hsk1 phosphorylates and targets fission yeast histone deacetylase Hst4 for degradation to stabilize stalled DNA replication forks

Hsp90-associated DNA replication checkpoint protein and proteasome-subunit components are involved in the age-related macular degeneration

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