ABSTRACT
Rad18 functions at the cross-roads of three different DNA damage response (DDR) pathways involved in protecting stressed replication forks: homologous recombination repair, DNA inter-strand cross-link repair and DNA damage tolerance. Although Rad18 serves to facilitate replication of damaged genomes by promoting translesion synthesis (TLS), this comes at a cost of potentially error-prone lesion bypass. In contrast, loss of Rad18-dependent TLS potentiates the collapse of stalled forks and leads to incomplete genome replication. Given the pivotal nature with which Rad18 governs the fine balance between replication fidelity and genome stability, Rad18 levels and activity have a major impact on genomic integrity. Here, we identify the de-ubiquitylating enzyme USP7 as a critical regulator of Rad18 protein levels. Loss of USP7 destabilizes Rad18 and compromises UV-induced PCNA mono-ubiquitylation and Pol η recruitment to stalled replication forks. USP7-depleted cells also fail to elongate nascent daughter strand DNA following UV irradiation and show reduced DNA damage tolerance. We demonstrate that USP7 associates with Rad18 directly via a consensus USP7-binding motif and can disassemble Rad18-dependent poly-ubiquitin chains both in vitro and in vivo. Taken together, these observations identify USP7 as a novel component of the cellular DDR involved in preserving the genome stability.
Subject(s)
DNA Damage , DNA Repair , DNA-Binding Proteins/metabolism , Ubiquitin Thiolesterase/metabolism , Ubiquitin-Protein Ligases/metabolism , Amino Acid Motifs , Cell Line , HeLa Cells , Humans , Protein Binding , Protein Stability , Ubiquitin/metabolism , Ubiquitin-Specific Peptidase 7ABSTRACT
As part of 'EduSens', a project aiming to measure the effect of a sensory education program developed in France on the food behaviour of school children, the present paper shows the results regarding neophobia. One hundred and eighty children (8-10 years old) were involved in the study. Half of them (experimental group) were educated during school-time with the 12 sessions of taste lessons "Les classes du goût" by J. Puisais. The others served as a control group. Food neophobia was evaluated before and after the education period of the experimental group and once again 10 months later. An adapted food neophobia scale was used (AFNS) and the willingness to taste novel food (WTNF) was evaluated by the presentation of eight unknown foods. To improve involvement in the expressed willingness to taste new foods, the children were told that they would have to eat one of the not rejected unknown foods afterwards. Results revealed that, at the end of the education period, in the educated group, declarative food neophobia decreased significantly and participants' willingness to taste novel food seemed to increase compared to the control group. Nevertheless, these effects had disappeared 10 months later. Thus, we have shown that sensory education can influence childrens' food neophobia, but does so only temporarily. This is especially true for the WTNF test, which measures the expression of neophobia in concrete situations, whereas neophobia measured as a psychological trait by the AFNS test hardly changes.
Subject(s)
Child Nutrition Sciences/education , Feeding Behavior/psychology , Food Preferences/psychology , Phobic Disorders/psychology , Taste/physiology , Child , Exploratory Behavior , Feeding Behavior/physiology , Female , Food Preferences/physiology , France , Humans , Male , Psychology, Child , PsychometricsABSTRACT
With the availability of complete DNA sequences for many prokaryotic and eukaryotic genomes, and soon for the human genome itself, it is important to develop reliable proteome-wide approaches for a better understanding of protein function. As elementary constituents of cellular protein complexes and pathways, protein-protein interactions are key determinants of protein function. Here we have built a large-scale protein-protein interaction map of the human gastric pathogen Helicobacter pylori. We have used a high-throughput strategy of the yeast two-hybrid assay to screen 261 H. pylori proteins against a highly complex library of genome-encoded polypeptides. Over 1,200 interactions were identified between H. pylori proteins, connecting 46.6% of the proteome. The determination of a reliability score for every single protein-protein interaction and the identification of the actual interacting domains permitted the assignment of unannotated proteins to biological pathways.
Subject(s)
Bacterial Proteins/metabolism , Helicobacter pylori/metabolism , Amino Acid Sequence , Binding Sites , Databases, Factual , Escherichia coli/genetics , Gene Library , Humans , Internet , Molecular Sequence Data , Protein Binding , Proteome , Sequence Alignment , Software , Urease/metabolismABSTRACT
Mutations affecting mismatch repair result in elevated frequencies of microsatellite length alteration in prokaryotes and eukaryotes. However, the finding that microsatellite instability is found often in cells with a functional mismatch repair system prompted a search for other factors of tract alteration. In the present report, we show that, in Escherichia coli, poly(AC/TG) tracts are destabilized by mutations that induce SOS. These observations may have implications for eukaryotic cells because recent results suggest the existence of a mammalian SOS response analogous to that in prokaryotes. In addition, a defect in the 5'-3' exonuclease domain of DNA polymerase I, homologous to the mammalian FEN1 and the yeast RAD27 nucleases, leads to a marked increase in repeat expansions characteristic of several genetic disorders. Finally, we found that the combination of a proofreading defect with mismatch repair deficiency results in extreme microsatellite instability.