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.

SourceResearch connectionFrom yearThrough year

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

At least 37 records · Page 2Linked to original sources

PubMed1977

Induction of protein X in Escherichia coli.

Certain treatments that damage DNA and/or inhibit replication in E. coli have been reported to induce synthesis of a new protein, termed protein X, in recA+ lexA+ strains. We have examined some of the treatments that might induce protein X and we have, in particular, tested the hypothesis of Gudas and Pardee (1975) that DNA degradation products play an essential role in the induction process. We confirmed that UV irradiation, nalidixic acid treatment, or thymine starvation result in protein X synthesis in wild type strains. However, we found that UV irradiation, unlike nalidixic acid, also induced protein X in recB strains, in which little DNA degradation occurs. Furthermore, we found that the presence of DNA fragments resulting from host-controlled restriction of phage lambda DNA did not affect protein X synthesis. We conclude that no causal relationship exists between the production of DNA fragments and induction of protein X. The presence of the plasmid R46, which confers enhanced mutagenesis and UV resistance on its host, did not affect protein X synthesis. Growth in the presence of 5-bromouracil, which does not result in production of degradation fragments, resulted eventually in a low rate of protein X synthesis. In dnaA mutants, deficient in the initiation of new rounds of replication, UV irradiation induced protein X, again unlike nalidixic acid. Thus, the inhibition of active replication forks is not an essential requirement for protein X induction.

Bacterial Proteins↗

PubMed2025

Altered histone modifications in Aedes aegypti following Rift Valley fever virus exposure.

When arthropod-borne viruses (arboviruses) are delivered to vector mosquitoes in an infectious bloodmeal, viral components interact with host proteins to hijack cells and initiate replication. The extent to which arbovirus infection alters mosquito host transcriptional and genomic regulatory processes is currently unknown. We hypothesized that histone modifications would be altered in mosquitoes exposed to Rift Valley fever virus (RVFV MP12, Phlebovirus riftense, family Phleboviridae). We interrogated transcriptome and chromatin landscapes in Aedes aegypti midguts by performing Cleavage Under Targets and Release Using Nuclease (CUT&RUN), using H3K27ac and H3K9me3 marks. Altered H3K27ac marks were identified following RVFV MP12 exposure, as well as upon bloodfeeding alone. It took several days for differential H3K27ac marks to be associated with differentially expressed genes (DEGs) in RVFV-exposed midguts. H3K27ac peaks showed progressive depletion as infection progressed. Gene set enrichment analysis revealed that immune response transcripts were enriched at 1 and 3 dpf (days post-feeding) but depleted by 7 dpf. Hedgehog/Gli (glioma-associated oncogene homolog) signaling pathway transcripts were depleted, indicating possible viral manipulation of cellular polarization. Moreover, at 7 dpf, 7 of 102 DEGs were proximal to differentially acetylated sites in a pattern expected to favor viral propagation. However, one transcript coding for an antiviral effector (LysM-TLDc domain protein) showed significant depletion of both H3K9me3 and H3K27ac marks. Analysis of midguts after a non-infectious bloodmeal versus sugar-fed controls revealed global changes to H3K27ac and H3K9me3 marks during and following the period of bloodmeal digestion. Differential H3K27ac marks were proximal to one quarter of all DEGs at 1 dpf, consistent with an important role of H3K27ac in bloodmeal digestion. These results demonstrate that H3K27ac and H3K9me3 patterns are altered upon virus exposure in a complex interplay that favors viral replication but is also countered by host responses to limit replication.

ChIP-Seq↗

PubMed1978

Site-specific initiation of a DNA fragment: nucleotide sequence of the bacteriophage G4 negative-strand initiation site.

The synthesis of the bacteriophage G4 negative strand is an example of the de novo initiation of a polynucleotide chain. This initiation is performed by the Escherichia coli replication protein dna G which selects a unique site on 5400-base positive-strand template. In this paper we present the nucleotide sequence of the G4 negative-strand initiation site. This is the template element recognized by the dna G priming protein. In conjunction with the sequence of the nascent negative strand, obtained by Bouché, Rowen, and Kornberg [Bouché, J.-P., Rowen, L. & Kornberg, A. (1978) J. Biol. Chem. 253, 765-769], the present data provide a description of a dna G-dependent origin of replication in which one knows the place at which polymerization starts at the nucleotide level.

Base Sequence↗

PubMed1976

The specific role of ribosomal protein S1 in the recognition of native phage RNA.

The previously reported requirement of ribosomal protein S1 for translation of phage RNA is now shown to be related to the involvement of the protein in initiation complex formation. The structure of the messenger RNA appears to be uniquely related to S1 function, since translation and initiation and midly unfolded phage RNA (by modification with formaldehyde) are independent of S1. It is proposed that S1 functions in conjunction with initiation factor IF-3 by recognizing and unfolding elements of the tertiary structure of phage RNA. A model is suggested for S1 function in both initiation of protein synthesis and initiation of phage RNA replication.

Antigen-Antibody Reactions↗

PubMed1979

The recovery of mammalian cells treated with methyl methanesulfonate, nitrogen mustard or UV light. I. The effect of alkylation products on DNA replication.

CHO cells were synchronized in G1 phase and treated with MMS or HN2. The subsequent rate of DNA replication was found to be reduced in a dose-dependent manner. In addition, 2 X 10(-3 M and 3 X 10(-3) M MMS resulted in a 3--4 h delay prior to the initiation of S phase. If the cells were held for 8 h in hydroxyurea after MMS treatment, no subsequent lag in DNA synthesis was seen after removal of the hydroxyurea. The entry of confluent cells into S phase was found to be delayed 7 h upon trypsinizing and replating. Treatment of these cells with MMS resulted in a reduced rate of DNA replication, but no further delay in its initiation. Repair replication was found to continue at a constant rate for at least 12 h following MMS treatment of cells under all of these conditions. At the concentrations used in these experiments MMS severely inhibited the rate of protein synthesis, but HN2 had little effect. By comparing both the kinetics of repair replication and recovery of protein synthesis with the rate of DNA replication, it was concluded that the initial, severe reduction in rate following MMS treatment was probably due to an inhibition of protein synthesis.

Animals↗

PubMed1977

phiX174 cistron A protein is a multifunctional enzyme in DNA replication.

The cistron A protein induced by phage varphiX174 nicks (produces a single-strand break in) the viral strand of the superhelical varphiX duplex DNA, thereby forming a complex with the DNA. The protein, seen bound to the DNA in the electron microscope, was located in the restriction endonuclease fragment between nucleotides 4290 and 4330 on the varphiX map [Sanger, F., Air, G. M., Barrel, B. G., Brown, N. L., Coulson, A. R., Fiddes, J. C., Hutchison, C. A., III, Slocomb, P. M. Y. & Smith, M. (1977) Nature 265, 687-695]. Replication also was initiated at this point, thus identifying the site of cistron A protein nicking and binding as the origin of replication. The cisA-DNA complex (separated from free cistron A protein), upon the addition of Escherichia coli rep protein, ATP, and DNA binding protein, is unwound to generate a single-stranded linear [presumably the nicked (+) strand] and a circular [presumably the (-) strand] molecule. The cisA-DNA complex, upon the further addition of DNA polymerase III holoenzyme and deoxynucleoside triphosphates, supports replication to generate viral, single-stranded circles, as many as 15 circles per cisA-DNA complex. The replicating intermediates seen in the electron microscope are a novel form of "rolling circle" [Gilbert, W. & Dressler, D. H. (1969) Cold Spring Harbor Symp. Quant. Biol. 33, 473-485]. The 5' end (presumably with the cistron A protein bound to it) is locked in the replication fork and loops back to accompany the strand-separation and replication fork around the template [(-) strand] circle. Thus, the multiple functions of cistron A protein include: (i) nicking the viral strand at the origin of replication to initiate a round of replication, (ii) participating in a complex which supports fork movement in strand separation and replication, (iii) nicking again at the regenerated origin to produce a unit-length DNA, and (iv) ligating the newly generated 3'-OH end to the 5'-phosphate-complexed end to form a circular viral molecule.

Coliphages↗

PubMed1977

[Regulation of DNA replication (author's transl)].

Two aspects of the regulatory mechanisms of DNA replication are discussed: 1. When resting mammalian cells are stimulated to proliferate a genetic program is expressed which provides proteins and other replication factors; 2. the interaction of proteins with specific sections on the DNA leads to initiation; most probably, structural changes in chromatin are also involved. Once initiated, replication proceeds almost automatically through the elongation phase. Lymphocytes stimulated to proliferate by concanavalin A are used as an example to demonstrate the relative importance of the two aspects of DNA replication.

Animals↗

PubMed2027

Production of Viral Particles from a Chikungunya Virus Infectious Clone.

Chikungunya virus (CHIKV) is a positive-sense single-stranded RNA virus, which poses challenges for its study and genetic manipulation. Because direct mutagenesis of viral RNA genomes is technically impractical, reverse genetics systems are essential tools for investigating viral biology. To enable such approaches, infectious clones containing a full-length cDNA copy of the viral genome are constructed. The cDNA is positioned under the control of a bacteriophage RNA polymerase promoter, allowing commercial RNA polymerases to use the linearized plasmid as a template for the in vitro transcription of full-length viral genomic RNA (gRNA). Importantly, positive-sense viral genomes serve as mRNAs for the translation of viral proteins in a cellular environment, meaning that these transcripts contain all the information required to initiate viral replication. Following transfection into permissive cultured cells, viral proteins are expressed, enabling genome replication and, ultimately, the recovery of infectious particles from the cell supernatant. Here, we describe a detailed procedure for generating CHIKV particles through plasmid linearization, in vitro transcription, and subsequent RNA transfection.

Chikungunya virus↗

PubMed2026

The cellular protein TIAR mediates rapid initiation of West Nile virus genome RNA synthesis.

During the intracellular replication cycle of West Nile virus (WNV), genome RNA synthesis is initially inefficient but increases exponentially as viral replication complexes are sequestered in invaginations in the endoplasmic reticulum. In this study, we investigated the functional role of the cellular protein TIAR (T-cell intracellular antigen-related protein) in the transcription of WNV genome RNA. Close colocalization of cytoplasmic TIAR with viral double-stranded RNA was detected by a proximity ligation assay in WNV-infected cells. TIAR binds specifically to the WNV 3'(-) SL but not to the complementary WNV 5'(+) SL in in vitro RNA binding assays. Only the 3' end of the WNV minus-strand RNA was enriched by immunoprecipitation of infected cell lysates with anti-TIAR antibody. Stable overexpression of TIAR in clonal A549 cells increased the ratio of intracellular viral plus-strand to minus-strand RNA in a dose-dependent manner. TIAR contains three RNA recognition motifs (RRMs). Biophysical data indicated that only RRM2 directly contacts RNA and that up to three TIAR molecules can bind cooperatively to the WNV 3'(-) SL RNA. These data provide additional evidence that TIAR functions as a proviral host factor facilitating exponential amplification of WNV genome production in infected cells.IMPORTANCEWest Nile virus (WNV) is a mosquito-borne orthoflavivirus associated with increasing global human disease incidence. The molecular mechanisms underlying viral replication are not fully understood. In early stages of infection, viral genome transcription is inefficient; however, in late stages, viral genome transcription increases exponentially. T-cell intracellular antigen-related (TIAR) protein is a cellular protein that has been shown to interact with the 3' end of the WNV negative-sense antigenomic RNA. We obtained data showing colocalization of cellular TIAR with viral replication complexes in infected cells and an increased ratio of intracellular genomic to antigenomic viral RNA in TIAR-overexpressing cells, and confirmed preferential binding of TIAR to the 3' end of the WNV antigenome both in vitro and in infected cell extracts. We also demonstrated that multiple TIAR proteins can bind cooperatively to the WNV 3'(-) stem-loop RNA. These data provide supporting evidence for a model of TIAR-mediated rapid initiation of nascent genome RNA synthesis in infected cells.

TIAR↗

PubMed1979

dnaG (primase)-dependent origins of DNA replication. Nucleotide sequences of the negative strand initiation sites of bacteriophages St-1, phi K, and alpha 3.

The simplest known origins of DNA replication occur in the single-stranded bacteriophages. In one set of phages, negative strand synthesis is initiated by a single protein, the product of the Escherichia coli replication gene dnaG. Evidently, in these phages--G4, St-1, phi K, and alpha 3--the origin for negative strand synthesis consists of a nucleic acid element capable of direct recognition by the dnaG priming protein. We have located and sequenced the origins of negative strand synthesis in St-1, phi K, and alpha 3, and compared them with the origin sequence previously determined for G4. In each case, the point at which the negative strand is initiated can be identified at the nucleotide level. The data lead to the following conclusions: 1. In all four phages, the negative strand initiation site occurs within an intercistronic region of approximately 135 bases. While in G4, the origin lies between genes specifying the viral coat proteins F and G, the origin is shifted in St-1, phi K, and alpha 3 to a position between coat protein genes G and H. 2. Extensive nucleotide conservation exists at the negative strand origin, but does not extend into the adjacent coding regions. The conserved origin DNA occurs in two regions, 42 and 45 bases long, which are separated by 13 bases of divergent sequence. 3. Correlated with the two stretches of conserved nucleotide sequence are two regions of potential secondary structure. The start point of negative strand synthesis lies just prior to one of these hairpins. Similarities in both primary sequence and secondary structure can be found between the negative strand origins of G4, St-1, phi K, and alpha 3 and the general origin regions of bacteriophage lambda and of E. coli.

Base Sequence↗

PubMed1979

Inhibition of initiation of bacteriophage T4 DNA replication by perturbation of Escherichia coli host membrane composition.

3-Decynoyl-N-acetylcysteamine (3-decynoyl-NAC) is an analog which specifically causes the immediate cessation of the biosynthesis of unsaturated fatty acids in Escherichia coli, whereas the synthesis of saturated fatty acids is actually stimulated. As a result, the cell membrane accumulates saturated fatty acids in its phospholipid. Addition of the inhibitor at the time of infection of E. coli by T4 phage had no effect on normal phage replication and development, implying that the synthesis of unsaturated fatty acids per se has little effect on T4 DNA replication. However, if the integrity and composition of the bacterial membrane was grossly perturbed by first treating the cells with the inhibitor for 60 min before infection, the proper initiation and the attainment of a rapid rate of T4 DNA synthesis were not observed. Under these conditions, a full complement of T4 early proteins was synthesized. The membrane associability of the known DNA delay proteins induced by wild-type T4 phage in the treated cells resembled that expected of a culture of untreated cells infected with a DNA delay mutant. When any one of three DNA delay mutants was used to infect 3-decynoyl-NAC-treated cells, T4 DNA replication was aborted. These findings suggest that some kind of specific interactions among the initiation proteins defined by the DNA delay mutants and the bacterial membrane may be necessary to facilitate the normal initiation and rapid rate of T4 DNA replication. A model for the involvement of the three different initiation proteins and the subsequent attainment of rapid DNA synthesis is discussed.

Cell Membrane↗

PubMed1979

Mutational analysis of the simian virus 40 replicon: pseudorevertants of mutants with a defective replication origin.

The circular genome of simian virus 40 is a model mammalian replicon, containing a unique origin of replication (ori) and coding for a protein (SV40 T antigen) known to be involved in initiation of viral DNA replication and to bind in vitro to the origin region. Mutations within the ori sequence lead to defective viral DNA replication and the formation of small viral plaques after infection of a cell monolayer. Second-site revertants (pseudorevertants) of ori mutants were isolated by random local mutagenesis of mutant DNA followed by transfection of cultured cells and the selection of large plaques. In each case, reversion of the plaque phenotype was associated with an increased rate of viral DNA replication. The second-site mutations that suppressed the replication defects were localized by in vitro recombination or marker rescue experiments to the gene for T antigen. Their map positions differ from those of previously described T antigen mutants, possibly reflecting a specific ori-binding domain of T antigen. From these results we infer that T antigen interacts with the ori signal during virus development as it does in vitro and that this interaction regulates the rate of viral DNA replication.

Antigens, Neoplasm↗

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↗

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↗