Replication protein A
About: Replication protein A is a research topic. Over the lifetime, 5172 publications have been published within this topic receiving 377645 citations. The topic is also known as: RPA & replication protein A.
Papers published on a yearly basis
TL;DR: The amount and length of telomeric DNA in human fibroblasts does in fact decrease as a function of serial passage during ageing in vitro and possibly in vivo.
Abstract: The terminus of a DNA helix has been called its Achilles' heel. Thus to prevent possible incomplete replication and instability of the termini of linear DNA, eukaryotic chromosomes end in characteristic repetitive DNA sequences within specialized structures called telomeres. In immortal cells, loss of telomeric DNA due to degradation or incomplete replication is apparently balanced by telomere elongation, which may involve de novo synthesis of additional repeats by novel DNA polymerase called telomerase. Such a polymerase has been recently detected in HeLa cells. It has been proposed that the finite doubling capacity of normal mammalian cells is due to a loss of telomeric DNA and eventual deletion of essential sequences. In yeast, the est1 mutation causes gradual loss of telomeric DNA and eventual cell death mimicking senescence in higher eukaryotic cells. Here, we show that the amount and length of telomeric DNA in human fibroblasts does in fact decrease as a function of serial passage during ageing in vitro and possibly in vivo. It is not known whether this loss of DNA has a causal role in senescence.
TL;DR: The data suggest that RPA-coated ssDNA is the critical structure at sites of DNA damage that recruits the ATR-ATRIP complex and facilitates its recognition of substrates for phosphorylation and the initiation of checkpoint signaling.
Abstract: The function of the ATR (ataxia-telangiectasia mutated- and Rad3-related)-ATRIP (ATR-interacting protein) protein kinase complex is crucial for the cellular response to replication stress and DNA damage. Here, we show that replication protein A (RPA), a protein complex that associates with single-stranded DNA (ssDNA), is required for the recruitment of ATR to sites of DNA damage and for ATR-mediated Chk1 activation in human cells. In vitro, RPA stimulates the binding of ATRIP to ssDNA. The binding of ATRIP to RPA-coated ssDNA enables the ATR-ATRIP complex to associate with DNA and stimulates phosphorylation of the Rad17 protein that is bound to DNA. Furthermore, Ddc2, the budding yeast homolog of ATRIP, is specifically recruited to double-strand DNA breaks in an RPA-dependent manner. A checkpoint-deficient mutant of RPA, rfa1-t11, is defective for recruiting Ddc2 to ssDNA both in vivo and in vitro. Our data suggest that RPA-coated ssDNA is the critical structure at sites of DNA damage that recruits the ATR-ATRIP complex and facilitates its recognition of substrates for phosphorylation and the initiation of checkpoint signaling.
TL;DR: Recent progress is described in understanding of how cells detect and signal the presence and repair of one particularly important form of DNA damage induced by ionizing radiation—the DNA double-strand break (DSB).
Abstract: To ensure the high-fidelity transmission of genetic information, cells have evolved mechanisms to monitor genome integrity. Cells respond to DNA damage by activating a complex DNA-damage-response pathway that includes cell-cycle arrest, the transcriptional and post-transcriptional activation of a subset of genes including those associated with DNA repair, and, under some circumstances, the triggering of programmed cell death. An inability to respond properly to, or to repair, DNA damage leads to genetic instability, which in turn may enhance the rate of cancer development. Indeed, it is becoming increasingly clear that deficiencies in DNA-damage signaling and repair pathways are fundamental to the etiology of most, if not all, human cancers. Here we describe recent progress in our understanding of how cells detect and signal the presence and repair of one particularly important form of DNA damage induced by ionizing radiation-the DNA double-strand break (DSB). Moreover, we discuss how tumor suppressor proteins such as p53, ATM, Brca1 and Brca2 have been linked to such pathways, and how accumulating evidence is connecting deficiencies in cellular responses to DNA DSBs with tumorigenesis.
TL;DR: The evidence presented strongly supports a role for the gamma-H2AX and the PI-3 protein kinase family in focus formation at sites of double-strand breaks and suggests the possibility of a change in chromatin structure accompanying double-Strand break repair.
Abstract: Background: The response of eukaryotic cells to double-strand breaks in genomic DNA includes the sequestration of many factors into nuclear foci. Recently it has been reported that a member of the histone H2A family, H2AX, becomes extensively phosphorylated within 1–3 minutes of DNA damage and forms foci at break sites. Results: In this work, we examine the role of H2AX phosphorylation in focus formation by several repair-related complexes, and investigate what factors may be involved in initiating this response. Using two different methods to create DNA double-strand breaks in human cells, we found that the repair factors Rad50 and Rad51 each colocalized with phosphorylated H2AX (γ-H2AX) foci after DNA damage. The product of the tumor suppressor gene BRCA1 also colocalized with γ-H2AX and was recruited to these sites before Rad50 or Rad51. Exposure of cells to the fungal inhibitor wortmannin eliminated focus formation by all repair factors examined, suggesting a role for the phosphoinositide (PI)-3 family of protein kinases in mediating this response. Wortmannin treatment was effective only when it was added early enough to prevent γ-H2AX formation, indicating that γ-H2AX is necessary for the recruitment of other factors to the sites of DNA damage. DNA repair-deficient cells exhibit a substantially reduced ability to increase the phosphorylation of H2AX in response to ionizing radiation, consistent with a role for γ-H2AX in DNA repair. Conclusions: The pattern of γ-H2AX foci that is established within a few minutes of DNA damage accounts for the patterns of Rad50, Rad51, and Brca1 foci seen much later during recovery from damage. The evidence presented strongly supports a role for the γ-H2AX and the PI-3 protein kinase family in focus formation at sites of double-strand breaks and suggests the possibility of a change in chromatin structure accompanying double-strand break repair.
TL;DR: It is found that purified Sp1 requires Zn(II) for sequence-specific binding to DNA, and it is likely that Sp1 interacts with DNA by binding of the Zn (II) fingers.
Abstract: Transcription factor Sp1 is a protein present in mammalian cells that binds to GC box promoter elements and selectively activates mRNA synthesis from genes that contain functional recognition sites. We have isolated a cDNA that encodes the 696 C-terminal amino acid residues of human Sp1. By expression of truncated fragments of Sp1 in E. coli, we have localized the DNA binding activity to the C-terminal 168 amino acid residues. In this region, Sp1 has three contiguous Zn(II) finger motifs, which are believed to be metalloprotein structures that interact with DNA. We have found that purified Sp1 requires Zn(II) for sequence-specific binding to DNA. Thus, it is likely that Sp1 interacts with DNA by binding of the Zn(II) fingers. To facilitate the identification of mutant variants of Sp1 that are defective in DNA binding, we have also devised a bacterial colony assay for detection of Sp1 binding to DNA.
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