Functions In Genome Stability Research Articles

Disordered regions and folded modules in CAF-1 promote histone deposition in Schizosaccharomyces pombe

Genome and epigenome integrity in eukaryotes depends on the proper coupling of histone deposition with DNA synthesis. This process relies on the evolutionary conserved histone chaperone CAF-1 for which the links between structure and functions are still a puzzle. While studies of the Saccharomyces cerevisiae CAF-1 complex enabled to propose a model for the histone deposition mechanism, we still lack a framework to demonstrate its generality and in particular, how its interaction with the polymerase accessory factor PCNA is operating. Here, we reconstituted a complete SpCAF-1 from fission yeast. We characterized its dynamic structure using NMR, SAXS and molecular modeling together with in vitro and in vivo functional studies on rationally designed interaction mutants. Importantly, we identify the unfolded nature of the acidic domain which folds up when binding to histones. We also show how the long KER helix mediates DNA binding and stimulates SpCAF-1 association with PCNA. Our study highlights how the organization of CAF-1 comprising both disordered regions and folded modules enables the dynamics of multiple interactions to promote synthesis-coupled histone deposition essential for its DNA replication, heterochromatin maintenance, and genome stability functions.

Newborn ribosomal DNA content as a potential early marker of genomic instability and longevity in association with prenatal air pollution exposure

BACKGROUND AND AIM: Repetitive regions in the genome, including ribosomal DNA repeats, have essential functions in genome stability. Alterations in ribosomal DNA copy numbers (rDNAcn) associates with genome-wide methylation and gene-expression, and links with longevity, as proposed in the rDNA theory of aging. Prenatal air pollution exposures affect molecular markers of aging that may underly later life health effects. We studied the association between prenatal air pollution exposure and this novel potential aging marker (rDNAc) in newborns. METHODS: Cord blood rDNAcn (45S rDNA) was measured in 177 newborns from the ENVIRONAGE birth cohort using ddPCR. Maternal residential exposure to PM10, PM2.5, NO2, and BC during pregnancy was estimated using a high-resolution spatial-temporal interpolation method. Using multivariable-adjusted distributed lag models, newborn rDNAcn was associated with average weekly air pollution exposures. RESULTS: The mean cord blood rDNAcn was 245 (SD: 64). Higher prenatal exposure to PM10, PM2.5, and NO2 in early to mid-pregnancy (weeks 9-18) was associated with an increase in rDNAcn. Each SD increment in air pollutants was associated with an average increase of 22.5 (95%CI: 5.9 to 35.0) copy numbers. Higher exposure to PM10, PM2.5, NO2, and BC late in pregnancy (weeks 25-35) was strongly associated with a decrease in rDNAcn. Each SD increment in air pollutants was associated with an average decrease of –29.6 (95%CI: –49.9 to –9.4) copy numbers. CONCLUSIONS: We studied for the first time rDNAcn in newborns and show that prenatal air pollution exposure is associated with rDNA repeats at birth. These results add evidence to the impact of air pollution on genome stability and early aging mechanisms. Evaluating the rDNA concept of aging in relation to diseases may further show the link between prenatal air, molecular alterations, and health later in life. KEYWORDS: genome instability, aging, ribosomal DNA copy number, prenatal air pollution

The role of histone H3K36me3 writers, readers and erasers in maintaining genome stability

Histone Post-Translational Modifications (PTMs) play fundamental roles in mediating DNA-related processes such as transcription, replication and repair. The histone mark H3K36me3 and its associated methyltransferase SETD2 (Set2 in yeast) are archetypical in this regard, performing critical roles in each of these DNA transactions. Here, we present an overview of H3K36me3 regulation and the roles of its writers, readers and erasers in maintaining genome stability through facilitating DNA double-strand break (DSB) repair, checkpoint signalling and replication stress responses. Further, we consider how loss of SETD2 and H3K36me3, frequently observed in a number of different cancer types, can be specifically targeted in the clinic through exploiting loss of particular genome stability functions.

SUMOylated Senataxin functions in genome stability, RNA degradation, and stress granule disassembly, and is linked with inherited ataxia and motor neuron disease.

BackgroundSenataxin (SETX) is a DNA/RNA helicase critical for neuron survival. SETX mutations underlie two inherited neurodegenerative diseases: Ataxia with Oculomotor Apraxia type 2 (AOA2) and Amyotrophic Lateral Sclerosis type 4 (ALS4).MethodsThis review examines SETX key cellular processes and we hypothesize that SETX requires SUMO posttranslational modification to function properly.ResultsSETX is localized to distinct foci during S‐phase of the cell cycle, and these foci represent sites of DNA polymerase/RNA polymerase II (RNAP) collision, as they co‐localize with DNA damage markers 53BP1 and H2AX. At such sites, SETX directs incomplete RNA transcripts to the nuclear exosome for degradation via interaction with exosome component 9 (Exosc9), a key component of the nuclear exosome. These processes require SETX SUMOylation. SETX was also recently localized within stress granules (SGs), and found to regulate SG disassembly, a process that similarly requires SUMOylation.ConclusionSETX undergoes SUMO modification to function at S‐phase foci in cycling cells to facilitate RNA degradation. SETX may regulate similar processes in non‐dividing neurons at sites of RNAP II bidirectional self‐collision. Finally, SUMOylation of SETX appears to be required for SG disassembly. This SETX function may be crucial for neuron survival, as altered SG dynamics are linked to ALS disease pathogenesis. In addition, AOA2 point mutations have been shown to block SETX SUMOylation. Such mutations induce an ataxia phenotype indistinguishable from those with SETX null mutation, underscoring the importance of this modification.

DNA Polymerase Eta Prevents Tumor Cell-Cycle Arrest and Cell Death during Recovery from Replication Stress.

: Neoplastic transformation and genome instability are enhanced by replication stress, conditions that slow or stall DNA replication forks. Consequently, cancer cells require multiple enzymes and checkpoint signaling pathways to mitigate replication stress for their viability and proliferation. Targeting proteins that enhance cancer cell survival during replication stress is a recent approach in clinical strategies, especially when targets produce synthetic lethality. DNA polymerase eta (Pol η) has many key functions in genome stability, particularly for translesion synthesis. Here we demonstrate that endogenous Pol η displays significant protein induction and forms intense foci throughout the nucleus in response to replication stress induced by drugs that do not directly form DNA adducts. During replication stress, Pol η-deficient cells displayed hyperactivation of the ATR replication checkpoint and arrested late in the cell cycle. During recovery from replication stress, Pol η-deficient cells continue to display aberrant phenotypes, including delayed cell-cycle progression, apoptosis, and cell survival. Depletion or inhibition of ATR was synthetically lethal with Pol η deficiency, particularly when tumor cells were treated with replication stress-inducing drugs. Together our data expand knowledge of the cellular environments that increase endogenous Pol η expression beyond DNA damaging agents and demonstrate that Pol η regulation is central to the replication stress response. Because Pol η is aberrantly expressed in several tumor types, our results are critical for developing more effective chemotherapy approaches and identify coinhibition of Pol η and ATR as a potential therapeutic strategy. SIGNIFICANCE: This study demonstrates that replication stress upregulates Pol η (POLH) in tumor cells and reveals a role for Pol η in tumor cell recovery following replication stress.

Cullin family proteins and tumorigenesis: genetic association and molecular mechanisms.

Cullin family proteins function as scaffolds to form numerous E3 ubiquitin ligases with RING proteins, adaptor proteins and substrate recognition receptors. These E3 ligases further recognize numerous substrates to participate in a variety of cellular processes, such as DNA damage and repair, cell death and cell cycle progression. Clinically, cullin-associated E3 ligases have been identified to involve numerous human diseases, especially with regard to multiple cancer types. Over the past few years, our understanding of cullin proteins and their functions in genome stability and tumorigenesis has expanded enormously. Herein, this review briefly provides current perspectives on cullin protein functions, and mainly summarizes and discusses molecular mechanisms of cullin proteins in tumorigenesis.

Phosphoinositide 3-kinase beta controls replication factor C assembly and function

Genomic integrity is preserved by the action of protein complexes that control DNA homeostasis. These include the sliding clamps, trimeric protein rings that are arranged around DNA by clamp loaders. Replication factor C (RFC) is the clamp loader for proliferating cell nuclear antigen, which acts on DNA replication. Other processes that require mobile contact of proteins with DNA use alternative RFC complexes that exchange RFC1 for CTF18 or RAD17. Phosphoinositide 3-kinases (PI3K) are lipid kinases that generate 3-poly-phosphorylated-phosphoinositides at the plasma membrane following receptor stimulation. The two ubiquitous isoforms, PI3Kalpha and PI3Kbeta, have been extensively studied due to their involvement in cancer and nuclear PI3Kbeta has been found to regulate DNA replication and repair, processes controlled by molecular clamps. We studied here whether PI3Kbeta directly controls the process of molecular clamps loading. We show that PI3Kbeta associated with RFC1 and RFC1-like subunits. Only when in complex with PI3Kbeta, RFC1 bound to Ran GTPase and localized to the nucleus, suggesting that PI3Kbeta regulates RFC1 nuclear import. PI3Kbeta controlled not only RFC1– and RFC–RAD17 complexes, but also RFC–CTF18, in turn affecting CTF18-mediated chromatid cohesion. PI3Kbeta thus has a general function in genomic stability by controlling the localization and function of RFC complexes.