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Division of Labor Between PCNA Loaders in DNA Replication and Sister Chromatid Cohesion Establishment [Affymetrix ChIP-chip]

ABSTRACT: Analysis of PCNA levels at stalled Saccharomyces cerevisiae replication forks upon Replication Factor C (RFC) Removal Analysis of nascent DNA incorporation at progressing Saccharomyces cerevisiae replication forks upon Rfc1 depletion

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE138054 | GEO | 2020/03/29

REPOSITORIES: GEO

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Division of Labor between PCNA Loaders in DNA Replication and Sister Chromatid Cohesion Establishment [BrdU-IP-ssSeq, ChIP-ssSeq and eSPAN analysis]

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Division of Labor between PCNA Loaders in DNA Replication and Sister Chromatid Cohesion Establishment.

Project description:Concomitant with DNA replication, the chromosomal cohesin complex establishes cohesion between newly replicated sister chromatids. Several replication-fork-associated "cohesion establishment factors," including the multifunctional Ctf18-RFC complex, aid this process in as yet unknown ways. Here, we show that Ctf18-RFC's role in sister chromatid cohesion correlates with PCNA loading but is separable from its role in the replication checkpoint. Ctf18-RFC loads PCNA with a slight preference for the leading strand, which is dispensable for DNA replication. Conversely, the canonical Rfc1-RFC complex preferentially loads PCNA onto the lagging strand, which is crucial for DNA replication but dispensable for sister chromatid cohesion. The downstream effector of Ctf18-RFC is cohesin acetylation, which we place toward a late step during replication maturation. Our results suggest that Ctf18-RFC enriches and balances PCNA levels at the replication fork, beyond the needs of DNA replication, to promote establishment of sister chromatid cohesion and possibly other post-replicative processes.

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Budding yeast Wapl controls sister chromatid cohesion maintenance and the chromosome condensation status [BrdU-ChIP].

Project description:Cohesin acetylation by Eco1 during DNA replication establishes sister chromatid cohesion. We show that acetylation makes cohesin resistant to Wapl activity from S-phase until mitosis. Wapl turns out to be a key regulator of cohesin dynamics on chromosomes by controling cohesin maintenance following its establishment in S-phase and its role in chromosome condensation. The Affymetrix Saccharomyces cerevisiae Chip Tiling 1.0F Arrays were used to analyze the incorporation of BrdU in Saccharomyces cerevisiae in S-phase arrested cells.

2012-11-22 | E-GEOD-42456 | biostudies-arrayexpress

Division of Labor Between PCNA Loaders in DNA Replication and Sister Chromatid Cohesion Establishment

Project description:This SuperSeries is composed of the SubSeries listed below.

2020-03-28 | GSE138056 | GEO

Budding yeast Wapl controls sister chromatid cohesion maintenance and the chromosome condensation status [PK-ChIP]

Project description:Cohesin acetylation by Eco1 during DNA replication establishes sister chromatid cohesion. We show that acetylation makes cohesin resistant to Wapl activity from S-phase until mitosis. Wapl turns out to be a key regulator of cohesin dynamics on chromosomes by controling cohesin maintenance following its establishment in S-phase and its role in chromosome condensation. The Affymetrix Saccharomyces cerevisiae Chip Tiling 1.0R Arrays were used to analyze the binding pattern of Scc1 along the genome of Saccharomyces cerevisiae in late G1 and metaphase arrested cells.

2012-11-22 | E-GEOD-42457 | biostudies-arrayexpress

Transcription profiling of reduced folate carrier (RFC1-/-) mouse knockouts at E9.5 reveals altered gene expression of the cubulin-megalin complex

Project description:The reduced folate carrier (RFC1) is an integral membrane protein and facilitative anion exchanger that mediates delivery of 5-methyltetrahydrofolate into mammalian cells. Adequate maternal-fetal transport of folate is necessary for normal embryogenesis. Targeted inactivation of the murine RFC1 gene results in post-implantation embryo lethality, but daily folic acid supplementation of pregnant dams prolongs survival of homozygous embryos until mid-gestation. At E10.5 RFC1-/- embryos are developmentally delayed relative to wildtype littermates, have multiple malformations, including neural tube defects, and die due to failure of chorioallantoic fusion. The mesoderm is sparse and disorganized, and there is a marked absence of erythrocytes in yolk sac blood islands. Affymetrix microarray analysis and quantitative RT-PCR validation of the relative gene expression profiles in E9.5 RFC1-/- vs. RFC1+/+ embryos indicates a dramatic downregulation of multiple genes involved in erythropoiesis, and upregulation of several genes that form the cubilin-megalin multiligand endocytic receptor complex. Megalin protein expression disappears from the visceral yolk sac of RFC1-/- embryos, and cubilin protein is widely misexpressed. Inactivation of RFC1 impacts the expression of several ligands and interacting proteins in the cubilin-amnionless-megalin complex that are involved in the maternal-fetal transport of folate, vitamin B12, and other nutrients, lipids and morphogens required for normal embryogenesis. Comparison of RFC KO, wildtype normal embryos vs. RFC KO, nullizygous affected embryos Experiment Overall Design: 6 samples RFC KO mouse embryos, E9.5, folic acid treated: 3 Control, Wildtype, normal; 3 Affected, Nullizygous, CR/chorioallantoic defect; as paired-littermates with one normal and one affected embryo per set from each of three separate litters for RNA extraction and hybridization on Affymetrix microarrays.

2008-04-05 | E-GEOD-10659 | biostudies-arrayexpress

Division of Labor Between PCNA Loaders in DNA Replication and Sister Chromatid Cohesion Establishment

Project description:Division of Labor Between PCNA Loaders in DNA Replication and Sister Chromatid Cohesion Establishment

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Nuclear pore complexes mediate subtelomeric gene silencing by regulating PCNA levels on chro atin

Project description:The nuclear pore complex (NPC) physically interacts with chromatin and regulates gene expression. The Saccharomyces cerevisiae inner ring nucleoporin Nup170 has been implicated in chromatin organization and the maintenance of gene silencing in subtelomeric regions. To gain insight into how Nup170 regulates this process, we used protein-protein interactions, genetic interactions, and transcriptome correlation analyses to identify the Ctf18-RFC complex, an alternative proliferating cell nuclear antigen (PCNA) loader, as a facilitator of the gene regulatory functions of Nup170. The Ctf18-RFC complex is recruited to a subpopulation of NPCs that lack the nuclear basket proteins Mlp1 and Mlp2. In the absence of Nup170, PCNA levels on DNA are reduced, resulting in the loss of silencing of subtelomeric genes. Increasing PCNA levels on DNA by removing Elg1, which is required for PCNA unloading, rescues subtelomeric silencing defects in nup170Δ. The NPC therefore mediates subtelomeric gene silencing by regulating PCNA levels on DNA.

2023-06-13 | PXD041496 | Pride

AFFYMETRIX ANALYSIS OF E9.5 RFC MOUSE KO EMBRYOS REVEALS ALTERED EXPR’N OF GENES IN THE CUBILIN-MEGALIN COMPLEX

Project description:The reduced folate carrier (RFC1) is an integral membrane protein and facilitative anion exchanger that mediates delivery of 5-methyltetrahydrofolate into mammalian cells. Adequate maternal-fetal transport of folate is necessary for normal embryogenesis. Targeted inactivation of the murine RFC1 gene results in post-implantation embryo lethality, but daily folic acid supplementation of pregnant dams prolongs survival of homozygous embryos until mid-gestation. At E10.5 RFC1-/- embryos are developmentally delayed relative to wildtype littermates, have multiple malformations, including neural tube defects, and die due to failure of chorioallantoic fusion. The mesoderm is sparse and disorganized, and there is a marked absence of erythrocytes in yolk sac blood islands. Affymetrix microarray analysis and quantitative RT-PCR validation of the relative gene expression profiles in E9.5 RFC1-/- vs. RFC1+/+ embryos indicates a dramatic downregulation of multiple genes involved in erythropoiesis, and upregulation of several genes that form the cubilin-megalin multiligand endocytic receptor complex. Megalin protein expression disappears from the visceral yolk sac of RFC1-/- embryos, and cubilin protein is widely misexpressed. Inactivation of RFC1 impacts the expression of several ligands and interacting proteins in the cubilin-amnionless-megalin complex that are involved in the maternal-fetal transport of folate, vitamin B12, and other nutrients, lipids and morphogens required for normal embryogenesis. Comparison of RFC KO, wildtype normal embryos vs. RFC KO, nullizygous affected embryos Keywords: reduced folate carrier knockout, folate receptor, cubilin, megalin, embryos, gene expression, neural tube defect, chorioallantoic fusion

2008-03-17 | GSE10659 | GEO

Genome-wide mapping proteins at replication forks in the yeast S. cerevisiae

Project description:Faithful duplication of DNA is essential for the maintenance of genomic stability in all organisms. DNA synthesis proceeds bi-directionally with continuous synthesis of leading strand DNA and discontinuous synthesis of lagging strand DNA. Herein, we describe a method of enriching and Sequencing of Protein-Associated Nascent strand DNA (eSPAN) to detect whether a protein binds the leading- and lagging-strands of DNA replication forks. We show that Pol-epsilon, PCNA, Cdc45, Mcm6 and Mcm10 preferentially associate with leading strands, whereas Pol-alpha, Pol32, Pol-delta, Rfa1 and Rfc1 associate with lagging strands of hydroxyurea (HU)-stalled replication forks. In contrast, PCNA is enriched at lagging strands of normal replication forks in wild type cells and HU-stalled forks in cells lacking Elg1. These studies demonstrate a strategy to reveal proteins at leading and lagging strands of DNA replication forks, and suggest that the unloading of PCNA from lagging strands of HU-stalled replication forks helps maintain genome integrity.

2014-10-29 | GSE52614 | GEO

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