Parental Template Research Articles

Improving the label-free rapid semi-quantification of E.coli by AgNPs-decorated hydrogel inverse opal photonic crystals

Using photonic crystal materials for colorimetric sensing, the influences of structure and composition on the photonic bandgap (PBG) of materials are always vital to detection. In this work, the effects of pores in the structure of polyethylene diacrylate-based inverse opal photonic crystal (PEGDA-based IOPC) and the concentration of silver nanoparticles (AgNPs) used for decoration of the IOPC by chemical reaction on the shift of PBG center (λrp) have been studied for rapid semi-quantification of Escherichia coli (E. coli). We found that pores in the structure of PEGDA-based IOPC significantly increased the amount of AgNPs attached to this material compared with the structure created by the ordered self-assembly of spherical SiO2 particles of the parent template, leading to a pronounced red-shift of λrp. At the concentration of 20 µg/L AgNPs (CAgNPs) used for decoration, the shift of λrp reached 80 nm due to the increase in the average refractive index (naver.) of the material. It thus enabled us to observe changes in the color of the reflected light with the naked eye. In addition, since the increase in the concentration of E.coli (CE.coli) also caused a red-shift of the λrp, the CAgNPs, thus, could be reduced but still give a sufficiently strong shift of λrp to detect E.coli at a high level. The spectral position of the reflectance peak changed from green to red with increasing the CE.coli from 50 cfu/mL to 106 cfu/mL at the CAgNPs = 10 µg/L. These results indicated the potential application of AgNPs decorated PEGDA-based IOPC in rapid semi-quantification of E.coli by the naked eye.

One-ended and two-ended breaks at nickase-broken replication forks

Replisome collision with a nicked parental DNA template can lead to the formation of a replication-associated double strand break (DSB). How this break is repaired has implications for cancer initiation, cancer therapy and therapeutic gene editing. Recent work shows that collision of a replisome with a nicked DNA template can give rise to either a single-ended (se) or a double-ended (de)DSB, with potentially divergent effects on repair pathway choice and genomic instability. Emerging evidence suggests that the biochemical environment of the broken mammalian replication fork may be specialized in such a way as to skew repair in favor of homologous recombination at the expense of non-homologous end joining.

Strand dependent bypass of DNA lesions during fork reversal by ATP-dependent translocases SMARCAL1, ZRANB3, and HLTF.

During DNA replication, the replisome encounters obstacles including DNA lesions, transcription-replication conflicts, and other sources of replication stress. These obstacles must be efficiently overcome to complete DNA synthesis and minimize genome instability. One pathway to tolerate replication stress is replication fork reversal, in which parental template DNA strands are reannealed and a nascent-nascent DNA duplex is formed. Several enzymes promote replication fork reversal, including the ATP-dependent translocases SMARCAL1, ZRANB3, and HLTF. How these enzymes translocate on DNA that contains fork-stalling lesions is unknown. Here, we examined the abilities of SMARCAL1, ZRANB3, and HLTF to tolerate various lesions on leading or lagging template strands. We demonstrate that SMARCAL1 and ZRANB3 are selectively inhibited by lesions on the leading template strand, whereas HLTF is insensitive to bulky lesions on either strand. These results suggest that SMARCAL1 and ZRANB3 contact the leading strand during fork reversal and therefore are more sensitive to inhibition by bulky lesions on this strand. In contrast, HLTF DNA translocation is inherently insensitive to DNA lesions. These biochemical differences between the fork reversal enzymes provide insights into their mechanism of DNA remodeling and suggest they may act in lesion-specific contexts.

Mechanisms and regulation of replication fork reversal

DNA replication is remarkably accurate with estimates of only a handful of mutations per human genome per cell division cycle. Replication stress caused by DNA lesions, transcription-replication conflicts, and other obstacles to the replication machinery must be efficiently overcome in ways that minimize errors and maximize completion of DNA synthesis. Replication fork reversal is one mechanism that helps cells tolerate replication stress. This process involves reannealing of parental template DNA strands and generation of a nascent-nascent DNA duplex. While fork reversal may be beneficial by facilitating DNA repair or template switching, it must be confined to the appropriate contexts to preserve genome stability. Many enzymes have been implicated in this process including ATP-dependent DNA translocases like SMARCAL1, ZRANB3, HLTF, and the helicase FBH1. In addition, the RAD51 recombinase is required. Many additional factors and regulatory activities also act to ensure reversal is beneficial instead of yielding undesirable outcomes. Finally, reversed forks must also be stabilized and often need to be restarted to complete DNA synthesis. Disruption or deregulation of fork reversal causes a variety of human diseases. In this review we will describe the latest models for reversal and key mechanisms of regulation.

Stimulus-Responsive Four-Stranded DNA Nanoring Assembly to Host Multiple Nanosilver Clusters for Cooperatively Enhanced Fluorescence Biosensing.

Exploring the ability of four-stranded DNA nanorings (fsDNRs) to host multiple nanosilver clusters (NAgCs) for cooperatively amplifiable fluorescence biosensing to a specific initiator (tI*) is fascinating. By designing three DNA single strands and three analogous stem-loop hairpins, we developed a functional fsDNR through sequential cross-opening and overlapped hybridization. Note that a substrate strand (SS) was programmed with six modules: two severed splits (sT and sT') of NAgCs template, two sequestered segments by a middle unpaired spacer, and a partition for tI*-recognizable displacement, while sT and sT' were also tethered in two ends of three hairpins. At first, a triple dsDNA complex with stimulus-responsiveness was formed to guide the specific binding to tI*, while the exposed toehold of the SS activated the forward cascade hybridization of three hairpins, until the ring closure in the tailored self-assembly pathway for forming the fsDNR. The resulting four duplexes forced each pair of sT/sT' to be merged as the parent template in four nicks, guiding the preferential synthesis of four clusters in the shared fsDNR, thereby cooperatively amplifying the green fluorescence signal for sensitive assay of tI*. Meanwhile, the topological conformation of fsDNR can be stabilized by the as-formed cluster adducts to rivet the pair of two splits in the nicks. Benefitting from the self-enhanced effect of multiple emitters, this label-free fluorescent sensing strategy features simplicity, rapidity, and high on-off contrast, without involving complicated nucleic acid amplifiers.

Template assisted sol-gel synthesis of BiFeO3 hollow tubes: Introducing kapok fiber as a bio-template

Bismuth ferrite (BiFeO3) is a perovskite material well known for its multifunctional properties and related applications. The recent research on this material includes the synthesis of hollow-structured BiFeO3 using various methods and exploring their applications in photocatalysis and sensing. Here, we introduce a facile method for the synthesis of BiFeO3 hollow tubes using the sol-gel method, where kapok fiber collected from Ceiba pentandra is used as a biotemplate for the first time. In the procedure, the annealing path was modified by introducing intermediate holding steps besides varying the final annealing temperature. The structural analysis was carried out using X-ray diffraction (XRD) and Raman analysis, whereas the formation of hollow tube morphology was confirmed with the help of FESEM analysis. The complete decomposition of the kapok fiber template during the annealing process was confirmed with the help of Fourier transform infrared spectroscopy. The XRD analysis indicated that monitoring both the annealing pathway and the final annealing temperature is pivotal in attaining the formation of phase pure BiFeO3. The proposed method eliminates the requirement for additional procedures for extracting the synthesized hollow tubes from the parent template, in addition to providing a less expensive strategy for the synthesis of BiFeO3 hollow tubes.

Identification, heterologous expression and characterization of a new unspecific peroxygenase from Marasmius fiardii PR-910

Unspecific peroxygenases (UPOs) are glycosylated enzymes that provide an efficient method for oxyfunctionalizing a variety of substrates using only hydrogen peroxide (H2O2) as the oxygen donor. However, their poor heterologous expression has hindered their practical application. Here, a novel UPO from Marasmius fiardii PR910 (MfiUPO) was identified and heterologously expressed in Pichia pastoris. By employing a two-copy expression cassette, the protein titer reached 1.18 g L−1 in a 5 L bioreactor, marking the highest record. The glycoprotein rMfiUPO exhibited a smeared band in the 40 to 55 kDa range and demonstrated hydroxylation, epoxidation and alcohol oxidation. Moreover, the peroxidative activity was enhanced by 150% after exposure to 50% (v/v) acetone for 40 h. A semi-preparative production of 4-OH-β-ionone on a 100 mL scale resulted in a 54.2% isolated yield with 95% purity. With its high expression level, rMfiUPO is a promising candidate as an excellent parental template for enhancing desirable traits such as increased stability and selectivity through directed evolution, thereby meeting the necessary criteria for practical application.Graphical

Fluorescence Biosensing Based on Bifurcated DNA Scaffold-Aggregated Ag Nanocluster via Responsive Conformation Switch of Quasi-Molecular Beacon.

To address the limitations of typical hairpin-structural molecular beacons, exploring the ability of a quasi-molecular beacon (qMB) to create label-free fluorescence biosensors is intriguing and remains a challenge. Herein, we propose the first example of modular qMB with the feature of a stimulation-responsive conformation switch to develop an aggregated Ag nanocluster (aAgNC) in a bifurcated DNA scaffold for fluorescently sensing a specific initiator (I*). This qMB was well designed to program four functional modules: I*-recognizable element adopting metastable stem-loop bihairpin structure and two DNA splits (exposed C3GT4 and locked C4AC4T) of aAgNC template that is separated by a tunable hairpin spacer for the customized combination of selective recognition and signaling readout. When presenting I* in an assay route, the specific hybridization induces the directional disassembly of the bihairpin unit, on which the qMB is configurationally switched to liberate the locked split. Thus, the bifurcated parent template pair of C3GT4/C4AC4T is proximal, affording in situ nucleation and clustering of emissive aAgNC. By collecting the fluorescence signal, the quantitative detection of I* is achieved. Benefiting from the ingenious programming of qMB, the recognizing and signaling integration actuates the construction of a facile and convenient fluorescent biosensor featuring rapid reaction kinetics, a wide linear range, high sensitivity, and specificity. This would provide a new paradigm to exploit versatile qMB-based biosensing platforms via stimulation-responsive conformation switches for developing various DNA-scaffolded Ag clusters.

Specialized replication of heterochromatin domains ensures self-templated chromatin assembly and epigenetic inheritance

Heterochromatin, defined by histone H3 lysine 9 methylation (H3K9me), spreads across large domains and can be epigenetically inherited in a self-propagating manner. Heterochromatin propagation depends upon a read-write mechanism, where the Clr4/Suv39h methyltransferase binds to preexisting trimethylated H3K9 (H3K9me3) and further deposits H3K9me. How the parental methylated histone template is preserved during DNA replication is not well understood. Here, we demonstrate using Schizosaccharomyces pombe that heterochromatic regions are specialized replication domains demarcated by their surrounding boundary elements. DNA replication throughout these domains is distinguished by an abundance of replisome components and is coordinated by Swi6/HP1. Although mutations in the replicative helicase subunit Mcm2 that affect histone binding impede the maintenance of a heterochromatin domain at an artificially targeted ectopic site, they have only a modest impact on heterochromatin propagation via the read-write mechanism at an endogenous site. Instead, our findings suggest a crucial role for the replication factor Mcl1 in retaining parental histones and promoting heterochromatin propagation via a mechanism involving the histone chaperone FACT. Engagement of FACT with heterochromatin requires boundary elements, which position the heterochromatic domain at the nuclear peripheral subdomain enriched for heterochromatin factors. Our findings highlight the importance of replisome components and boundary elements in creating a specialized environment for the retention of parental methylated histones, which facilitates epigenetic inheritance of heterochromatin.

Silicon Radical-Induced CH4 Dissociation for Uniform Graphene Coating on Silica Surface.

Due to the manufacturability of highly well-defined structures and wide-range versatility in its microstructure, SiO2 is an attractive template for synthesizing graphene frameworks with the desired pore structure. However, its intrinsic inertness constrains the graphene formation via methane chemical vapor deposition. This work overcomes this challenge by successfully achieving uniform graphene coating on a trimethylsilyl-modified SiO2 (denote TMS-MPS). Remarkably, the onset temperature for graphene growth dropped to 720°C for the TMS-MPS, as compared to the 885°C of the pristine SiO2. This is found to be mainly from the Si radicals formed from the decomposition of the surface TMS groups. Both experimental and computational results suggest a strong catalytic effect of the Si radicals on the CH4 dissociation. The surface engineering of SiO2 templates facilitates the synthesis of high-quality graphene sheets. As a result, the graphene-coated SiO2 composite exhibits a high electrical conductivity of 0.25Scm-1. Moreover, the removal of the TMP-MPS template has released a graphene framework that replicates the parental TMS-MPS template on both micro- and nano- scales. This study provides tremendous insights into graphene growth chemistries as well as establishes a promising methodology for synthesizing graphene-based materials with pre-designed microstructures and porosity.

Atomic Substitution to Tune ScO6 Distortion in Ba2MSc2(BO3)4 (M = Na, K, Ba) to Acquire a Large Birefringence.

Replacing alkali metals (K, Na atoms) by an alkaline-earth metal (Ba atom), α-Ba3Sc2(BO3)4 (high-temperature phase) is successfully obtained by a molten salt method, taking Ba2K1.6Na0.4Sc2(BO3)4 as the parent template. Although both of them exhibit similar layered structures composed of ScO6 and BO3 units, α-Ba3Sc2(BO3)4 shows largely distorted ScO6 octahedra (Δd = 0.56) forced by the uniform tension of a larger space effect from BaO12 polyhedrons, rather than regular ScO6 octahedra like in Ba2K1.6Na0.4Sc2(BO3)4. Experimental measurements and calculated analyses elucidate that distorted ScO6 octahedra in α-Ba3Sc2(BO3)4, displaying a second-order Jahn-Teller (SOJT) effect, enlarge the experimental birefringence up to 0.14@550 nm, while Ba2K1.6Na0.4Sc2(BO3)4 with regular ScO6 octahedra only shows Δn = 0.11 under the same condition. In addition, other optical and thermal properties of the two title compounds were characterized. The experimental results indicate that Ba2K1.6Na0.4Sc2(BO3)4 and α-Ba3Sc2(BO3)4 are promising birefringent materials.

DNA Mismatch Repair System Imbalances in Breast Adenocarcinoma.

DNA mismatch repair system (MMR) is considered a leading genetic mechanism in stabilizing DNA structure and maintaining its function. DNA MMR is a highly conserved system in bacteria, prokaryotic, and eukaryotic cells, and provides the highest protection to DNA by repairing micro-structural alterations. DNA MMR proteins are involved in the detection and repair of intra-nucleotide base-to-base errors inside the complementary DNA strand recognizing the recently synthesized strand from the parental template. During DNA replication, a spectrum of errors including base insertion, deletion, and miss-incorporation negatively affect the molecule's structure and its functional stability. A broad spectrum of genomic alterations such as promoter hyper methylation, mutation, and loss of heterozygosity (LOH) in MMR genes including predominantly hMLH1, hMSH2, hMSH3, hMSH6, hPMS1, and hPMS2 lead to their loss of base-to-base error repairing procedure. Microsatellite instability (MSI) refers to the DNA MMR gene alterations that are observed in a variety of malignancies of different histological origins. In the current review, we present the role of DNA MMR deficiency in breast adenocarcinoma, a leading cancer-based cause of death in females worldwide.

Non-birthing mothers’ experiences of perinatal anxiety and depression: Understanding the perspectives of the non-birthing mothers in female same-sex parented families

ObjectivePartners of birthing mothers can themselves experience perinatal mental health (PMH) difficulties. Despite birth rates increasing amongst LGBTQIA+ communities and the significant impact of PMH difficulties, this area is under-researched. This study aimed to examine the experiences of perinatal depression and anxiety of non-birthing mothers in female same-sex parented families. DesignInterpretative Phenomenological Analysis (IPA) was used to explore the experiences of non-birthing mothers who self-identified as having experienced perinatal anxiety and/or depression. Setting and participantsSevenparticipants were recruited from online and local voluntary and support networks for LGBTQIA+ communities and for PMH. Interviews were in-person, online or via telephone. Measurements and findingsSix themes were generated. Distress was characterised by feelings of “Failure and Inadequacy in Role” (i.e., parent, partner and individual) and “Powerlessness and Intolerable Uncertainty” in their parenting journey. These feelings were reciprocally influenced by perceptions of the “Legitimacy of (Di)stress as a Non-birthing Parent”, which impacted help-seeking. Stressors that contributed to these experiences were: “Parenting Without” a parental role template, social recognition and safety, and parental connectedness; and “Changed Relationship Dynamics” with their partner. Finally,participants spoke about “Moving Forward” in their lives. Key conclusionsSome findings are consistent with the literature on paternal mental health, including parents’ emphasis on protecting their family and experiencing services as focusing on the birthing parent. Others appeared distinct or amplified for LGBTQIA+ parents, including the lack of a defined and socially recognised role; stigma concerning both mental health and homophobia; exclusion from heteronormative healthcare systems; and the importance placed on biological connectedness. Implications for practiceCulturally competent care is needed to tackle minority stress and recognise diverse family forms.

New Lactobacillus plantarum membrane proteins (LpMPs) towards oral anti-inflammatory agents against dextran sulfate sodium-induced colitis.

Inflammatory bowel disease (IBD), a progressive and unpredictable colorectal inflammatory disease, is a global health problem. Currently, therapeutic strategies for the management of the disease are limited. Results from our previous studies indicated that probiotic Lactobacillus plantarum exhibits therapeutic effects against IBD, and through screening, we obtained an active 61-amino-acid long protein, L. plantarum membrane protein 1 (LpMP-1). Based on druggability-guided strategies, the search for LpMPs with lower molecular weights and better bioactivities contributes to the development of new anti-inflammatory agents to overcome the limitations of existing therapies against IBD. We used amino-acid-truncation strategies to obtain modified LpMPs (LpMP-2 - LpMP-9) using LpMP-1 as the parent template. Furthermore, we systematically evaluated the anti-colitis pharmacodynamics of these LpMPs in terms of symptomatology, histopathology, and cytokine levels in DSS-induced ulcerative colitis mice. Their possible targets of action against IBD was investigated under an iTRAQ-based pharmacoproteomic system and a docking-guided receptor-ligand relationship frame. We found a new active protein, LpMP-8, which had a lower molecular weight than LpMP-1. LpMP-8 was found to exhibit anti-colitis activity following oral administration in vivo (50μg/kg) by improving symptoms of colitis, colonic ulcerations, and cytokine disorders. TLRs and TGF-β were found to be involved in the action of LpMP-8 against colitis; LpMP-8 was to compete with TLR4-MD2-bound LPS and reverse TGF-β and Smad2/7 disorders. Our probiotic-derived LpMP-8 was shown to elicit oral anti-colitis activity, and its significant efficacy is probably associated with TLR4 and TGF-β.

Structure-Activity Relationship Study of Subtype-Selective Positive Modulators of KCa2 Channels.

A series of modified N-cyclohexyl-2-(3,5-dimethyl-1H-pyrazol-1-yl)-6-methylpyrimidin-4-amine (CyPPA) analogues were synthesized by replacing the cyclohexane moiety with different 4-substituted cyclohexane rings, tyrosine analogues, or mono- and dihalophenyl rings and were subsequently studied for their potentiation of KCa2 channel activity. Among the N-benzene-N-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-4-pyrimidinamine derivatives, halogen decoration at positions 2 and 5 of benzene-substituted 4-pyrimidineamine in compound 2q conferred a ∼10-fold higher potency, while halogen substitution at positions 3 and 4 of benzene-substituted 4-pyrimidineamine in compound 2o conferred a ∼7-fold higher potency on potentiating KCa2.2a channels, compared to that of the parent template CyPPA. Both compounds retained the KCa2.2a/KCa2.3 subtype selectivity. Based on the initial evaluation, compounds 2o and 2q were selected for testing in an electrophysiological model of spinocerebellar ataxia type 2 (SCA2). Both compounds were able to normalize the abnormal firing of Purkinje cells in cerebellar slices from SCA2 mice, suggesting the potential therapeutic usefulness of these compounds for treating symptoms of ataxia.

Deposition Bias of Chromatin Proteins Inverts under DNA Replication Stress Conditions.

Following DNA replication, equal amounts of chromatin proteins are distributed over sister chromatids by re-deposition of parental chromatin proteins and deposition of newly synthesized chromatin proteins. Molecular mechanisms balancing the allocation of new and old chromatin proteins remain largely unknown. Here, we studied the genome-wide distribution of new chromatin proteins relative to parental DNA template strands and replication initiation zones using the double-click-seq. Under control conditions, new chromatin proteins were preferentially found on DNA replicated by the lagging strand machinery. Strikingly, replication stress induced by hydroxyurea or curaxin treatment and inhibition of ataxia telangiectasia and Rad3-related protein (ATR) or p53 inactivation inverted the observed chromatin protein deposition bias to the strand replicated by the leading strand polymerase in line with previously reported effects on replication protein A occupancy. We propose that asymmetric deposition of newly synthesized chromatin proteins onto sister chromatids reflects differences in the processivity of leading and lagging strand synthesis.

Mechanisms for Maintaining Eukaryotic Replisome Progression in the Presence of DNA Damage.

The eukaryotic replisome coordinates template unwinding and nascent-strand synthesis to drive DNA replication fork progression and complete efficient genome duplication. During its advancement along the parental template, each replisome may encounter an array of obstacles including damaged and structured DNA that impede its progression and threaten genome stability. A number of mechanisms exist to permit replisomes to overcome such obstacles, maintain their progression, and prevent fork collapse. A combination of recent advances in structural, biochemical, and single-molecule approaches have illuminated the architecture of the replisome during unperturbed replication, rationalised the impact of impediments to fork progression, and enhanced our understanding of DNA damage tolerance mechanisms and their regulation. This review focusses on these studies to provide an updated overview of the mechanisms that support replisomes to maintain their progression on an imperfect template.

Replication of Sequence Information in Synthetic Oligomers.

ConspectusThe holy grail identified by Orgel in his 1995 Account was the development of novel chemical systems that evolve using reactions in which replication and information transfer occur together. There has been some success in the adaption of nucleic acids to make artificial analogues and in templating oligomerization reactions to form synthetic homopolymers, but replication of sequence information in synthetic polymers remains a major unsolved problem. In this Account, we describe our efforts in this direction based on a covalent base-pairing strategy to transfer sequence information between a parent template and a daughter copy. Oligotriazoles, which carry information as a sequence of phenol and benzoic acid side chains, have been prepared from bifunctional monomers equipped with an azide and an alkyne. Formation of esters between phenols and benzoic acids is used as the equivalent of nucleic base pairing to covalently attach monomer building blocks to a template oligomer. Sequential protection of the phenol side chains on the template, ester coupling of the benzoic acid side chains, and deprotection and ester coupling of the phenol side chains allow quantitative selective base-pair formation on a mixed sequence template. Copper catalyzed azide alkyne cycloaddition (CuAAC) is then used to oligomerize the monomers on the template. Finally, cleavage of the ester base pairs in the product duplex by hydrolysis releases the copy strand. This covalent template-directed synthesis strategy has been successfully used to copy the information encoded in a trimer template into a sequence-complementary oligomer in high yield.The use of covalent base pairing provides opportunities to manipulate the nature of the information transferred in the replication process. By using traceless linkers to connect the phenol and benzoic acid units, it is possible to carry out direct replication, reciprocal replication, and mutation. These preliminary results are promising, and methods have been developed to eliminate some of the side reactions that compete with the CuAAC process that zips up the duplex. In situ end-capping of the copy strand was found to be an effective general method for blocking intermolecular reactions between product duplexes. By selecting an appropriate concentration of an external capping agent, it is also possible to intercept macrocyclization of the reactive chain ends in the product duplex. The other side reaction observed is miscoupling of monomer units that are not attached to adjacent sites on the template, and optimization is required to eliminate these reactions. We are still some way from an evolvable synthetic polymer, but the chemical approach to molecular replication outlined here has some promise.

Efficacy of cobalt‐incorporated mesoporous silica toward photodegradation of Alizarin Red S and its kinetic study

AbstractCobalt‐incorporated mesoporous silica (Co‐MCM‐41) has been synthesized along with MCM‐41. The materials were characterized using powder X‐ray diffraction, scanning electron microscope, and nitrogen adsorption–desorption study techniques. The surface area (SBET, m2/g), pore size (Å), and pore volume (cc/g) have been found to be reduced in the Co‐MCM‐41 compared with MCM‐41. Furthermore, the Scanning electron microscopy coupled with energy dispersive X‐Ray spectroscopy (SEM‐EDAX) analysis has clearly shown the presence of the respective elements in the materials. The bandgap (eV) was also significantly reduced in the Co‐MCM‐41 compared with its parent template, observed by applying the Kubelka‐Munk function to the results of UV–Vis DRS. The materials were successfully used as photocatalysts in the photodegradation studies of the Alizarin Red S dye, and pseudo‐first‐order kinetics was performed using the Langmuir‐Hinshelwood kinetic model. All the required experimental conditions were optimized.

Molecular mechanisms of eukaryotic origin initiation, replication fork progression, and chromatin maintenance.

Eukaryotic DNA replication is a highly dynamic and tightly regulated process. Replication involves several dozens of replication proteins, including the initiators ORC and Cdc6, replicative CMG helicase, DNA polymerase α-primase, leading-strand DNA polymerase ε, and lagging-strand DNA polymerase δ. These proteins work together in a spatially and temporally controlled manner to synthesize new DNA from the parental DNA templates. During DNA replication, epigenetic information imprinted on DNA and histone proteins is also copied to the daughter DNA to maintain the chromatin status. DNA methyltransferase 1 is primarily responsible for copying the parental DNA methylation pattern into the nascent DNA. Epigenetic information encoded in histones is transferred via a more complex and less well-understood process termed replication-couple nucleosome assembly. Here, we summarize the most recent structural and biochemical insights into DNA replication initiation, replication fork elongation, chromatin assembly and maintenance, and related regulatory mechanisms.