Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Replication initiation protein”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 55 records · Page 3Linked to original sources
PubMed1975

Reconstruction of bacteriophage T4 DNA replication apparatus from purified components: rolling circle replication following de novo chain initiation on a single-stranded circular DNA template.

The protein products of T4 bacteriophage genes 41, 43, 45, 44, and 62 have been purified to near homogeneity using an assay which measures their stimulation of DNA synthesis in a crude lysate of Escherichia coli cells in fected by an appropriate mutant phage. When all of these proteins and T4 gene 32 protein are incubated in the presence of deoxyribonucleoside and ribonucleoside triphosphates, extensive DNA synthesis occurs on both single and double-stranded DNA templates. Analysis of this in vitro system reveals most of the features attributed to in vivo DNA replication: (1) De novo DNA chain initiation is found on a single-stranded DNA template only if ribonucleoside triphosphates are present (as expected for RNA priming of Okazaki pieces on the "lagging" strand of a replication fork). (2) With single-stranded circular DNA as template, synthesis continues for many doublings. The products after extensive synthesis resemble a rolling circle as visualized in the electron microscope, with discontinuous "lagging" strand synthesis generating a long, unbranched double-stranded tail. The fact that all six mutationally identified T4 replication gene products are required for these syntheses suggests the existence of a large multienzyme complex, constituting the T4 replication apparatus.

Chromosome Mapping↗
PubMed1975

Nature of R-factor replication in the presence of chloramphenicol.

Covalently closed circular deoxyribonucleic acid molecules of RSF1030, a nonconjugative R-factor, initiate and complete rounds of semiconservative replication in the absence of protein synthesis long after the bacterial chromosome has ceased its replication. RSF1030 replication under these conditions is sensitive to the inhibitor of ribonucleic acid synthesis, rifampicin. The product of this replication, a covalently closed DNA molecule, shows, in contrast to those molecules produced during the replication in a logarithmically growing culture of Escherichia coli, a transition to the open circular form upon treatment with ribonucleases of alkali. Analysis of the product resulting from alkali treatment indicates that there is a single break in one strand of the original circular duplex. This alkali-sensitive site occurs with equal probability in either of the complementary strands. These results are interpreted as a requirement for an RNA primer for the initiation of RSF1030 DNA synthesis and as showing that its removal from the covalently closed molecule is inhibited in the absence of protein synthesis.

Bacterial Proteins↗
PubMed2026

MCM8 promotes colorectal cancer progression by competitively inhibiting HRD1-mediated CDC42 ubiquitination and degradation.

BACKGROUND: Colorectal cancer (CRC) ranks among the top three in both incidence and mortality rates of malignant tumors worldwide. For patients with advanced colon cancer, radical surgery is challenging, and chemotherapy drugs are prone to inducing drug resistance, resulting in a five-year survival rate of only 13.1%. Therefore, in-depth analysis of the occurrence, development, and drug resistance mechanisms of colon cancer is of great clinical significance for optimizing treatment strategies and improving patient prognosis. As one of the homologous recombination repair proteins, minichromosomal maintenance protein 8 (MCM8) not only participates in DNA replication initiation, homologous recombination repair, and genome stability maintenance in normal cells, but also has been reported to be abnormally highly expressed in multiple tumors (e.g. glioblastoma, cholangiocarcinoma, bladder cancer) to promote malignant progression. METHODS: This study focused on the expression and function of MCM8 in colon cancer. The expression level of MCM8 in colon cancer tissues and cells was detected, and its correlation with patients’ clinicopathological features and prognosis was analyzed. Combined with cell function experiments, protein-protein interaction verification assays, and in vivo tumorigenesis experiments, the effects of MCM8 on the biological behaviors of colon cancer cells and the underlying molecular mechanisms were explored. Meanwhile, rescue experiments were conducted to identify the key downstream molecules and pathways mediated by MCM8. Additionally, the relationship between MCM8 and chemoresistance of colon cancer cells was investigated. RESULTS: Our study indicated that MCM8 promotes the transition of the cell cycle from the G1 phase to the S phase in CRC cell lines(SW620, HCT116, CX-1). Moreover, our study showed that MCM8 interacted with Cdc42(Cell Division Cycle 42) and promoted its protein stability by competitively inhibiting the ubiquitination modification of Cdc42‘s E3 ubiquitin ligase HRD1(Hydroxymethylglutaryl Reductase Degradation Protein 1). The rescue experiment showed that MCM8 promoted the proliferation, cell cycle progression, invasion, tumor-forming ability in vivo and resistance to 5-FU of CRC cell lines (SW620FR, HCT15FR) through Cdc42, while inhibiting cell apoptosis. CONCLUSIONS: MCM8 is abnormally highly expressed in CRC and stabilizes Cdc42 protein by competitively inhibiting HRD1, thereby promoting the occurrence and development of CRC and the formation of 5-FU resistance.

Humans↗
PubMed1979

DNA sequences and structural homologies of the replication origins of lambdoid bacteriophages.

The DNA sequences for the origins of replication of the lambdoid bacteriophages phi80, 434, phi21, and lambdaimm21 (identical to phi21) have been determined and compared to the lambda structure. Two presumptive elaborate binding sites for two initiator proteins have been identified in their outer sections, while a replicational primer start site seems to be located in their centres.

Base Sequence↗
PubMed2026

EIF4H and YBX1 are essential host factors for hepatitis E virus replication and pathogenesis.

Hepatitis E virus (HEV) is a leading cause of acute viral hepatitis worldwide, responsible for approximately 20 million infections annually. Despite the availability of a vaccine in China, no direct-acting antivirals are approved, and host factors required for HEV replication remain poorly defined. Here, using a genome-wide CRISPR/Cas9 knockout screen in a replicon system, we identified Eukaryotic Translation Initiation Factor 4H (EIF4H) and Y-Box Binding Protein 1 (YBX1) as essential host factors for HEV replication and pathogenesis. Knockout of either factor markedly impaired replication of HEV genotypes 1, 3, and 4, as well as HEV infection and production in hepatocellular carcinoma cells and human induced pluripotent stem cell-derived hepatocyte-like cells, while leaving SARS-CoV-2, hepatitis B virus, hepatitis C virus, and Zika virus unaffected, underscoring their HEV-specific roles. Mechanistically, EIF4H interacts with ORF1 via its methyltransferase-Y-papain-like protease region, and EIF4H deficiency alters the composition of the ORF1-associated replication complex. By contrast, YBX1 is dispensable for ORF1 translation and RNA binding but is specifically required for ORF1 proteolytic processing, a prerequisite for assembling a functional replication machinery. EIF4H knockout rats and liver-specific YBX1 knockout rats were largely resistant to rat HEV-C1 infection, showing profound reductions in viral shedding, suppressed hepatic and intestinal viral loads, and protection from liver pathology. Together, our findings establish EIF4H and YBX1 as essential host factors for HEV infection and pathogenesis and reveal potential targets for antiviral intervention.

Virus Replication↗
PubMed1978

The RNA primer synthesized by primase to initiate phage G4 DNA replication.

With phage G4 DNA as template, primase (the dnaG protein) synthesizes a 26- to 29-residue RNA transcript at the origin of replication. The sequence starts with ATP and contains a hairpin region of one A-U and seven G-C base pairs. Covalent linkage between the RNA and the newly synthesized complementary DNA chain indicates that the RNA transcript serves as a primer.

Base Sequence↗
PubMed1979

Inhibition of 80S initiation complex formation by infection with poliovirus.

Anisomycin has been shown to stabilize ribosome initiation complexes containing messenger RNA and met-tRNAf met to high salt conditions. Extracts from HeLa cells treated with 5 X 10(-7) M-anisomycin for 15 min accumulate 80S initiation complexes which can be detected by their absorbance in sucrose gradients. Poliovirus-infected cells fail to form the 80S initiation complex early after infection, when inhibition of host cell protein synthesis occurs. These complexes re-form later in infection after virus RNA is synthesized. No re-formation occurs in the absence of virus replication. Thus, the step in protein synthesis inhibited by poliovirus precedes the entry of components into the 80S initiation complex.

Anisomycin↗
PubMed1975

DNA synthesis in polyoma virus infection. III. Mechanism of inhibition of viral DNA replication by cycloheximide.

The formation of viral DNA was inhibited in polyoma virus-infected cells in which protein synthesis had been blocked by cycloheximide. The present studies show the following. (i) The pool of replicating viral DNA molecules was reduced in cycloheximide-treated cells by an amount consistent with inhibition of [3-H]thymidine incorporation into viral DNA, whereas the rate of turnover of the replicating population was not affected. (ii) The rate of conversion of replicating molecules into closed-circular DNA was not affected by cycloheximide. (iii) The rate of elongation of nascent viral DNA fragments into strands of unit genome length was unaffected by cycloheximide. It is concluded that viral DNA synthesis is inhibited in the absence of protein synthesis exclusively at the level of initiation of new rounds of genome replication. Replicating molecules already initiated at the time of addition of cycloheximide matured into progeny closed-circular DNA at a normal rate.

Animals↗
PubMed2023

H4S47 O-GlcNAcylation regulates the activation of mammalian replication origins.

The transmission and maintenance of genetic information in eukaryotic cells relies on the faithful duplication of the entire genome. In each round of division, excessive replication origins are licensed, with only a fraction activated to give rise to bi-directional replication forks in the context of chromatin. However, it remains elusive how eukaryotic replication origins are selectively activated. Here we demonstrate that O-GlcNAc transferase (OGT) enhances replication initiation by catalyzing H4S47 O-GlcNAcylation. Mutation of H4S47 impairs DBF4-dependent protein kinase (DDK) recruitment on chromatin, causing reduced phosphorylation of the replicative helicase mini-chromosome maintenance (MCM) complex and compromised DNA unwinding. Our short nascent-strand sequencing results further confirm the importance of H4S47 O-GlcNAcylation in origin activation. We propose that H4S47 O-GlcNAcylation directs origin activation through facilitating MCM phosphorylation, and this may shed light on the control of replication efficiency by chromatin environment.

Animals↗
PubMed2026

Crown ethers as artificial decoys: A supramolecular strategy to block SARS-CoV-2 entry via host-guest interactions.

Coronavirus disease (COVID-19) remains a major global health challenge, highlighting the need for antiviral strategies that act at the earliest stages of infection. Given that viral entry and spike-receptor interaction are critical steps in the coronavirus life cycle, targeting these processes represents a powerful strategy to block infection at its earliest stage. Inspired by the glycan-recognition and extracellular viral-trapping functions of pulmonary surfactant collectins (SP-A and SP-D), this work integrates supramolecular chemistry, pulmonary surfactant biology, and antiviral research to establish a biomimetic supramolecular molecular-decoy framework based on crown ethers, cyclodextrins, and related macrocyclic architectures. Through host-guest molecular recognition, these macrocyclic scaffolds can be engineered to mimic sialylated host receptors and multivalent glycan motifs, enabling competitive binding to viral spike proteins, virion capture, and sequestration away from epithelial surfaces. By redirecting viruses toward artificial host-mimetic structures, supramolecular decoys could intercept SARS-CoV-2 and other enveloped respiratory viruses before host-cell attachment, membrane fusion, or genome release. Acting upstream of intracellular replication, this strategy may prevent initiation of the viral replication cycle and subsequent hijacking of the host protein synthesis machinery, while potentially minimizing interference with host metabolic pathways and reducing the likelihood of resistance development. Furthermore, it can be translated into inhalation nanoformulations for pulmonary delivery and localized formulations targeting the upper respiratory tract. Overall, by integrating the biological principles of pulmonary surfactant immunity with supramolecular host-guest chemistry, this work provides a conceptual foundation for biomimetic molecular-decoy antivirals and highlights a promising direction for next-generation broad-spectrum antiviral design against emerging respiratory viruses.

Antiviral Agents↗
PubMed1975

Rifampin inhibition of bacteriophage phiX174 parental replicative-form DNA synthesis in an Escherichia coli dnaC mutant.

The Escherichia coli dnaC protein is not absolutely required in vivo for bacteriophage phiX174 parental replicative-form synthesis (Kranias and Dumas, 1974). However, when rifampin is present at a concentration that inhibits DNA-dependent RNA polymerase, phiX174 parental replicative-form synthesis is dependent on the dnaC protein activity. We conclude that E. coli DNA-dependent RNA polymerase can substitute for the dnaC protein in phiX174 parental replicative-form DNA synthesis, presumably in its initiation. The implications of this result with respect to the in vitro synthesis of the complementary strand of phiX174 DNA are discussed.

Bacterial Proteins↗
PubMed1978

Adenovirus core protein synthesis in the absence of viral DNA synthesis late in infection.

The acid extraction of the adenovirus type 5 core proteins V, VII, and pVII (the precursor to VII) from infected cells and the subsequent electrophoresis on a 15% acrylamide-2.5 M urea-0.9 N acetic acid (pH 2.7) gel, revealed that peptide VII has a similar electrophoretic mobility to that of histone H1. The core proteins, which are coded by late adenovirus mRNA, continued to be synthesized late in infection when viral DNA synthesis was inhibited either by cytosine arabinoside in wild-type infections or by shifting adenovirus H5 ts 125-infected cells to the nonpermissive temperature (40 degree C). Only the initiation, not the continuation, of viral DNA replication was essential for core protein synthesis. The synthesis of viral core proteins continued for over 8 h after the cassation of DNA synthesis. This was in contrast to the rapid shutdown of cellular histone synthesis in the absence of cellular DNA synthesis.

Adenoviruses, Human↗
PubMed1977

Fate of histone messenger RNA in synchronized HeLa cells in the absence of initiation of protein synthesis.

The fate of cytoplasmic histone mRNA was studied under conditions in which initiation of protein synthesis in synchronized HeLa cells is S phase was blocked by increasing the osmolarity of the growth medium with NaCl. In contrast to the interruption of DNA replication with hydroxyurea, which results in an exponential degradation of translatable histone mRNA with a half-life of about 10-13 min, blocking the initiation of protein synthesis leads to only a marginal loss of biologically active histone mRNA in the cytoplasm. When the initiation of protein synthesis was interrupted by treating cells with 150 mM NaCl, 40-50% of the total cytoplasmic histone mRNA previously translated in polyribosomes appears in the cytoplasm integrated into mRNA-protein particle(s) sedimenting between 15 S and 30 S. On the other hand, in untreated S-phase cells or in cells blocked with hydroxyurea only 3-6% of the total translatable histone mRNA is found in the cytoplasm not bound to ribosomes or their subunits. In addition, the degradation of histone mRNA in hydroxyurea-blocked S-phase cells is prevented when the initiation of protein synthesis is inhibited with NaCl. These studies clearly indicate that the inhibition of initiation of protein synthesis per se is not the cause for the rapid degradation of cytoplasmic histone mRNA observed when DNA replication is turned off and that the inactivation of these mRNAs is a process dependent on continuous protein synthesis.

Cell Division↗
PubMed2025

Proximity interactome of alphavirus replicase component nsP3 includes proviral host factors eIF4G and AHNAK.

All positive-strand RNA viruses replicate their genomes in association with modified intracellular membranes, inducing either membrane invaginations termed spherules, or double-membrane vesicles. Alphaviruses encode four non-structural proteins nsP1-nsP4, all of which are essential for RNA replication and spherule formation. To understand the host factors associated with the replication complex, we fused the efficient biotin ligase miniTurbo with Semliki Forest virus (SFV) nsP3, which is located on the cytoplasmic surface of the spherules. We characterized the proximal proteome of nsP3 in three cell lines, including cells unable to form stress granules, and identified >300 host proteins constituting the microenvironment of nsP3. These included all the nsPs, as well as several previously characterized nsP3 binding proteins. However, the majority of the identified interactors had no previously identified roles in alphavirus replication, including 39 of the top 50 interacting proteins. The most prominent biological processes involving the proximal proteins were nucleic acid metabolism, translational regulation, cytoskeletal rearrangement and membrane remodeling. siRNA silencing confirmed six novel proviral factors, USP10, AHNAK, eIF4G1, SH3GL1, XAB2 and ANKRD17, which are associated with distinct cellular functions. All of these except SH3GL1 were also important for the replication of chikungunya virus. We discovered that the small molecule 4E1RCat, which inhibits the interaction between the canonical translation initiation factors eIF4G and eIF4E, exhibits antiviral activity against SFV. Since the same molecule was previously found to inhibit coronaviruses, this suggest the possibility that translation initiation factors could be considered as targets for broadly acting antivirals.

Viral Nonstructural Proteins↗
PubMed2026

MCM10 and RECQL4 have cooperative and redundant roles in activating the CMG helicase during the replication initiation.

DNA replication initiation requires activation of the CMG helicase to establish the replisome. This process involves the extrusion of single-stranded DNA (ssDNA) from the central channel of MCM double hexamers, allowing the two CMG helicases to pass each other; however, the factors that mediate this process in human cells remain unclear. We show that degron-mediated depletion of either MCM10 or RECQL4 alone causes mild replication defects, whereas simultaneous depletion of both proteins severely impairs CMG activation. ChIP-seq analyses demonstrate that RECQL4 localizes to replication initiation zones (IZs) independently of MCM10, whereas MCM10 recruitment to IZs is enhanced upon RECQL4 depletion, consistent with partially redundant roles during CMG activation. Rescue experiments further indicate that RECQL4 cooperates with MCM10 through direct interaction, and that their ssDNA-binding activity underlies their functional overlap. We propose that MCM10 and RECQL4 act cooperatively and redundantly to promote CMG activation.

CMG activation↗
PubMed1976

Isolation of an intermediate which precedes dnaG RNA polymerase participation in enzymatic replication of bacteriophage phi X174 DNA.

Conversion of phi X174 single-stranded DNA to the duplex replicative form (RF) in vitro requires at least 10 purified proteins. Three stages - strand initiation, elongation, and termination - comprise this conversion. We now identify a separate stage in strand initiation which precedes dnaG RNA polymerase participation. Incubation of five proteins - protein i, protein n, DNA unwinding protein, dnaB protein, and dnaC protein - with ATP and phi X174 DNA forms an intermediate which enables subsequent stages measured by DNA synthesis to proceed 20 times faster. The intermediate can be isolated in quantitative yield by gel filtration or by ultracentrifugation. Protein i and protein n are required in less than stoichiometric amounts and appear to be absent from the isolated intermediate. Whereas formation of the intermediate is sensitive to antibody to protein i and to N-ethylmaleimide (an inhibitor of protein n and dnaC protein), the intermediate itself is resistant to these reagents. DNA unwinding protein complexes the DNA in a ratio of 60 molecules per circle. Synthesis of the intermediate appears to require stoichiometric quantities of dnaB protein and dnaC PROTEin but their presence in the intermediate has not been established as yet.

Bacterial Proteins↗
PubMed1975

Characteristics of a Bacillus subtilis W23 mutant temperature sensitive for initiation of chromosome replication.

A temperature-sensitive mutant of Bacillus subtilis W23, dna-20 (Ts), has been isolated and shown to be defective in initiation of rounds of chromosome replication at the nonpermissive temperature. Upon transfer of dna-20(Ts) from 30 to 45 C, deoxyribonucleic acid synthesis, as measured by [3H]thymine incorporation, gradually ceases. The distribution of genetic markers among unreplicated and replicated deoxyribonucleic acid, isolated from dna-20(Ts) after a period at 43 C in a medium containing 5-bromouracil, and fractionated in a CsCl gradient, shows that the cessation of initiation at the higher temperature is immediate. On the other hand, ribonucleic acid and protein synthesis continues at elevated or unaltered rates for some time after the shift to 45 C. Marker frequency analysis shows that all rounds of replication in progress at the time of the temperature shift terminate rapidly (within 40 min), even when chromosomes are replicating dichotomously in rich media. dna-20(Ts) remains 100% viable for at least 2 h at 45 C. Over a 5-h period at 45 C the nuclear bodies remain compact; a small number (less than 5%) of deoxyribonucleic acid-less cells are produced, but there is no morphological distortion of the cells. When the cells are returned to 30 C after 2 h at 45 C, chromosome replication is initiated rapidly at the normal origin and then proceeds in the normal established sequence. However, a second round of replication is initiated soon after the first. dna-20(Ts) has been shown to map as a B-group mutation, the major class of initiation mutants identified in B. subtillus 168.

Autoradiography↗