About
65
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Introduction
Our current research interests aim at studying the molecular bases of the DNA damage response in proliferating cells and translate these findings into innovative treatments for cancer.
Additional affiliations
January 2007 - present
CNRS-Institute of Human Genetics
Position
- Principal Investigator
October 2001 - December 2006
CNRS-Institute of Human Genetics
Position
- Researcher
June 1998 - September 2001
Publications
Publications (65)
Pluripotency of embryonic stem cells (ESC) is tightly regulated by a network of transcription factors among which the estrogen-related receptor β (Esrrb). Esrrb contributes to the relaxation of the G1 to S-phase (G1/S) checkpoint in mouse ESCs by transcriptional control of the deubiquitylase Dub3 gene, contributing to Cdc25A persistence after DNA d...
Proliferating cell nuclear antigen (PCNA) is a well-known scaffold for many DNA replication and repair proteins, but how the
switch between partners is regulated is currently unclear. Interaction with PCNA occurs via a domain known as a PCNA-Interacting
Protein motif (PIP box). More recently, an additional specialized PIP box has been described, th...
The molecular mechanism underlying G1/S checkpoint bypass in mouse embryonic stem cells (ESCs) remains unknown. DNA damage blocks S phase entry by inhibiting the CDK2 kinase through destruction of its activator, the Cdc25A phosphatase. We observed high Cdc25A levels in G1 that persist even after DNA damage in mouse ESCs. We also found higher expres...
Uncoupling between DNA polymerases and helicase activities at replication forks, induced by diverse DNA lesions or replication
inhibitors, generate long stretches of primed single-stranded DNA that is implicated in activation of the S-phase checkpoint.
It is currently unclear whether nucleation of the essential replication factor RPA onto this subs...
In early embryogenesis of fast cleaving embryos, DNA synthesis is short and surveillance mechanisms preserving genome integrity are inefficient, implying the possible generation of mutations. We have analyzed mutagenesis in Xenopus laevis and Drosophila melanogaster early embryos. We report the occurrence of a high mutation rate in Xenopus and show...
DNA can experience “replication stress”, an important source of genome instability, induced by various external or endogenous impediments that slow down or stall DNA synthesis. While genome instability is largely documented to favor both tumor formation and heterogeneity, as well as drug resistance, conversely, excessive instability appears to supp...
Cisplatin is a widely used chemotherapy drug, despite its significant
ototoxic side effects. To date, the mechanism of cisplatininduced
ototoxicity remains unclear, and hearing preservation
during cisplatin-based chemotherapy in patients is lacking. We
found activation of the ATM-Chk2-p53 pathway to be a major
determinant of cisplatin ototoxicity....
Maintenance and surveillance of genome integrity is crucial during the very early steps of embryonic development, since de novo mutations generated during this stage can be propagated in differentiated adult cells and may lead to predisposition to diseases including cancer. Surprisingly, early embryos are characterized by a relaxed control of genom...
In early embryogenesis of fast cleaving embryos DNA synthesis is short and surveillance mechanisms preserving genome integrity are inefficient implying the possible generation of mutations. We have analyzed mutagenesis in Xenopus laevis and Drosophila melanogaster early embryos. We report the occurrence of a high mutation rate in Xenopus and show t...
Imbalance in the level of the pyrimidine degradation products dihydrouracil and dihydrothymine is associated with cellular transformation and cancer progression. Dihydropyrimidines are degraded by dihy-dropyrimidinase (DHP), a zinc metalloenzyme that is upregulated in solid tumors but not in the corresponding normal tissues. How dihydropyrimidine m...
During the very early stages of embryonic development chromosome replication occurs under rather challenging conditions, including a very short cell cycle, absence of transcription, a relaxed DNA damage response and, in certain animal species, a highly contracted S-phase. This raises the puzzling question of how the genome can be faithfully replica...
Metabolic alterations support cellular transformation. Noticeably, cellular transformation and cancer progression is associated with accumulation of dihydrouracil and dihydrothymine. However, it remains elusive how dihydropyrimidine metabolites affect cellular phenotypes. Dihydropyrimidines are degraded by dihydropyrimidinase (DHP). This zinc metal...
DNA replication is an essential process occurring prior to cell division. Cell division coupled to proliferation ensures the growth and renewal of a large variety of specialized cell types generated during embryonic development. Changes in the DNA replication program occur during development. Embryonic undifferentiated cells show a high replication...
The DEAD-box Helicase 19 (Ddx19) gene codes for an RNA helicase involved in both messenger RNA (mRNA) export from the nucleus into the cytoplasm and in mRNA translation. In unperturbed cells, Ddx19 localizes in the cytoplasm and at the cytoplasmic face of the nuclear pore. Here we review recent findings related to an additional Ddx19 function in th...
Coordination between transcription and replication is crucial in the maintenance of genome integrity. Disturbance of these processes leads to accumulation of aberrant DNA:RNA hybrids (R-loops) that, if unresolved, generate DNA damage and genomic instability. Here we report a novel, unexpected role for the nucleopore-associated mRNA export factor Dd...
Early embryonic cleavages are characterized by short and highly synchronous cell cycles made of alternating S- and M-phases with virtually absent gap phases. In this contracted cell cycle, the duration of DNA synthesis can be extraordinarily short. Depending on the organism, the whole genome of an embryo is replicated at a speed that is between 20...
Early embryonic cleavages are characterized by short and highly synchronous cell cycles made of alternating S- and M-phases with virtually absent gap phases. In this contracted cell cycle, the duration of DNA synthesis can be extraordinarily short. Depending on the organism, the whole genome of an embryo is replicated at a speed that is between 20...
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Cisplatin is a widely used chemotherapy drug, despite its significant ototoxic side effects. To date, the mechanism of cisplatin-induced ototoxicity remains unclear, and hearing preservation during cisplatin-based chemotherapy in patients is lacking. We found activation of the ATM-Chk2-p53 pathway to be a major determinant of cisplatin ototoxicity....
Maintenance of genomic stability is crucial in ensuring cellular homeostasis and perpetuation of life. Perpetuation of the genetic information relies upon faithful replication of the genome. Mutations, generated during DNA synthesis and/or cell division and induced by exposure to external chemical agents, are drivers of genetic and associated genom...
In early embryos, the DNA damage checkpoint is silent until the midblastula transition (MBT) because of maternal limiting factors of unknown identity. Here we identify the RAD18 ubiquitin ligase as one such factor in Xenopus. We show, in vitro and in vivo, that inactivation of RAD18 function leads to DNA damage-dependent checkpoint activation, moni...
The major challenge of the cell cycle is to deliver an intact, and fully duplicated, genetic material to the daughter cells. To this end, progression of DNA synthesis is monitored by a feedback mechanism known as replication checkpoint that is untimely linked to DNA replication. This signaling pathway ensures coordination of DNA synthesis with cell...
The use of the human or animal MCM9 gene, or parts of the gene, or transcripts thereof, or antisense nucleic acids able to hybridize with part of the gene or transcripts, or silencing RNA derived from parts of the transcripts and able to repress the MCM9 gene, or proteins or peptidic fragments translated from the transcripts, or antibodies directed...
Formation of primed single-stranded DNA at stalled replication forks triggers activation of the replication checkpoint signalling cascade resulting in the ATR-mediated phosphorylation of the Chk1 protein kinase, thus preventing genomic instability. By using siRNA-mediated depletion in human cells and immunodepletion and reconstitution experiments i...
Activation of the replication checkpoint relies upon uncoupling of DNA polymerases and helicase activities at replication forks, which in multicellular organism results in production of long stretches of single-stranded DNA bound by the trimeric, single stranded DNA binding protein, the RPA complex. Binding of RPA to this substrate promotes synthes...
Repair of single-stranded DNA breaks before DNA replication is critical in maintaining genomic stability; however, how cells
deal with these lesions during S phase is not clear. Using combined approaches of proteomics and in vitro and in vivo protein–protein interaction, we identified the p58 subunit of DNA Pol α-primase as a new binding partner of...
Geminin is an important cell cycle regulator having a dual role in cell proliferation and differentiation. During proliferation, Geminin controls DNA synthesis by interacting with the licensing factor Cdt1 and interferes with the onset of differentiation by inhibiting the activity of transcription factors such as Hox and Six3. During early developm...
Initiation of DNA synthesis involves the loading of the MCM2-7 helicase onto chromatin by Cdt1 (origin licensing). Geminin is thought to prevent relicensing by binding and inhibiting Cdt1. Here we show, using Xenopus egg extracts, that geminin binding to Cdt1 is not sufficient to block its activity and that a Cdt1-geminin complex licenses chromatin...
The MCM proteins identify a group of ten conserved factors functioning in the replication of the genomes of archae and eukaryotic organisms. Among these, MCM2-7 proteins are related to each other and form a family of DNA helicases implicated at the initiation step of DNA synthesis. Recently this family expanded by the identification of two addition...
MCM2-7 proteins are conserved replication factors functioning as DNA helicases during DNA synthesis. MCM8 is another member of this family, which appears to be specific for higher eukaryotes, as it is absent in worms and yeast. Here we report the complete identification of a novel member of this family, the MCM9 protein. Like MCM8, MCM9 is only pre...
MCM2-7 proteins are replication factors required to initiate DNA synthesis and are currently the best candidates for replicative helicases. We show that the MCM2-7-related protein MCM8 is required to efficiently replicate chromosomal DNA in Xenopus egg extracts. MCM8 does not associate with the soluble MCM2-7 complex and binds chromatin upon initia...
A crucial regulation for maintaining genome integrity in eukaryotes is to limit DNA replication in S phase to only one round. Several models have been proposed; one of which, the licensing model, predicted that formation of the nuclear membrane restricts access to chromatin to a positive replication factor. Cdt1, a factor binding to origins and rec...
Replication protein A (RPA) is a three subunit single-stranded DNA-binding protein required for DNA replication. In Xenopus, RPA assembles in nuclear foci that form before DNA synthesis, but their significance in the assembly of replication initiation complexes has been questioned. Here we show that the RPA34 regulatory subunit is dephosphorylated...
We have determined the crystal structure of the coiled-coil domain of human geminin, a DNA synthesis inhibitor in higher eukaryotes. We show that a peptide encompassing the five heptad repeats of the geminin leucine zipper (LZ) domain is a dimeric parallel coiled coil characterized by a unique pattern of internal polar residues and a negatively cha...
In early Xenopus development, transcription is repressed and DNA replication initiates at non-specific sites. Here, we show that a site-specific DNA replication origin can be induced in this context by the assembly of a transcription domain. Deletion of the promoter element abolishes site-specific initiation, and its relocalization to an ectopic si...
Cdt1 is a conserved replication factor required in licensing the chromosome for a single round of DNA synthesis. The activity of Cdt1 is inhibited by geminin. The mechanism by which geminin interferes with Cdt1 activity is unknown. It is thought that geminin binds to and sequestrate Cdt1. We show that geminin does not interfere with the chromatin a...
The assembly of the DNA helicase at replication origins is crucial in initiating DNA synthesis. This process requires the conserved protein Cdt1. Here, a new study identifies a functional homologue of Cdt1 in Saccharomyces cerevisiae. The regulation of its activity reveals an alternative way to assemble prereplicative complexes (pre-RCs) and regula...
Eukaryotic replication origins are 'licensed' for replication early in the cell cycle by loading Mcm(2-7) proteins. As chromatin replicates, Mcm(2-7) are removed, thus preventing the origin from firing again. Here we report the purification of the RLF-B component of the licensing system and show that it corresponds to Cdt1. RLF-B/Cdt1 was inhibited...
In eukaryotic cells, chromosomal DNA replication begins with the formation of pre-replication complexes at replication origins. Formation and maintenance of pre-replication complexes is dependent upon CDC6 (ref. 1), a protein which allows assembly of MCM2-7 proteins, which are putative replicative helicases. The functional assembly of MCM proteins...
Acquisition of the competence to replicate requires the assembly of the MCM2-7 (minichromosome maintenance) protein complex onto pre-replicative chromatin, a step of the licensing reaction. This step is thought to occur through binding of a heterohexameric MCM complex containing the six related MCM subunits. Here we show that assembly of the MCM co...
We report the identification of a novel nucleolar protein from fission yeast, p17nhp2, which is homologous to the recently identified Nhp2p core component of H+ACA snoRNPs in Saccharomyces cerevisiae. We show that the fission yeast p17nhp2 localizes to the nucleolus in live S. cerevisiae or Schizosaccharomyces pombe cells and is functionally conser...
Although proteins involved in DNA replication in yeast have counterparts in multicellular organisms, the definition of an origin of DNA replication and its control in higher eukaryotes might obey to different rules. Origins of DNA replication that are site-specific have been found, supporting the notion that specific DNA regions are used to initiat...
MCM proteins are molecular components of the DNA replication licensing system inXenopus.These proteins comprise a conserved family made up of six distinct members which have been found to associate in large protein complexes. We have used a combination of biochemical and cytological methods to study the association of soluble and chromatin-boundXen...
We have previously revealed that in the brine shrimp Artemia franciscana an AluI DNA family of repeats, 113 bp in length, is the major component of the constitutive heterochromatin and that this repetitive DNA shows a stable curvature that confers a solenoidal geometry on the double helix in vitro. It was suggested that this particular structure ma...
A clearer picture of replication control is emerging through the characterization of proteins, such as cdc 18/Cdc6 and members of the mini-chromosome maintenance (MCM) protein family, that are involved in the initiation step. Cyclin B dependent kinases have conserved roles in both Saccharomyces cerevisiae and Schizosaccharomyces pombe, switching on...
The regulatory mechanism which ensures that eukaryotic chromosomes replicate precisely once per cell cycle is a basic and essential cellular property of eukaryotes. This fundamental aspect of DNA replication is still poorly understood, but recent advances encourage the view that we may soon have a clearer picture of how this regulation is achieved....
The fission yeast cdc21 protein belongs to the MCM family, implicated in the once per cell cycle regulation of chromosome replication. In budding yeast, proteins in this family are eliminated from the nucleus during S phase, which has led to the suggestion that they may serve to distinguish unreplicated from replicated DNA, as in the licensing fact...
The study of the structural organization of the eukaryotic genome is one of the most important tools for disclosing the evolutionary relationships between species.Artemia (Crustacea, Phyllopoda) offers a very interesting model for speciation studies. The genus, distributed all over the world, comprises both bisexual sibling species and parthenogene...
DNA bending has been suggested to play a role in the regulation of gene expression, initiation of DNA replication, site specific recombination and DNA packaging. In Artemia franciscana (Phillopoda anostraca) cells we have revealed that an AluI DNA family of repeats, 113-bp in length, is the major component of the constitutive heterochromatin found...
Thesis (Ph. D.)--University of Oxford, 1995.
Projects
Project (1)
We're studying the functional interaction between the transcription and replication machineries, focussing on the role of the Ddx19 RNA helicase, which we have recently shown to be involved in resolution of RNA:DNA hybrids, also known as R-loops, generated upon interference between transcription and replication (Hodroj et al., EMBO J. 2017).
We're also studying the molecular basis of cancer resistance to therapeutic treatments and have recently identified a new target strongly involved in this process.
Two postdoctoral positions are available immediately to support highly motivated candidates with a strong background in biochemistry and cell biology.