Mutation of the mouse Rad17 gene leads to embryonic lethality and reveals a role in DNA damage-dependent recombination

Genetic defects in DNA repair mechanisms and cell cycle checkpoint (CCC) genes result in increased genomic instability and cancer predisposition. Discovery of mammalian homologs of yeast CCC genes suggests conservation of checkpoint mechanisms between yeast and mammals. However, the role of many CCC genes in higher eukaryotes remains elusive. Here, we report that targeted deletion of an N-terminal part of mRad17, the mouse homolog of the Schizosaccharomyces pombe Rad17 checkpoint clamp-loader component, resulted in embryonic lethality during early/mid-gestation. In contrast to mouse embryos, embryonic stem (ES) cells, isolated from mRad17(5'Delta/5'Delta) embryos, produced truncated mRad17 and were viable. These cells displayed hypersensitivity to various DNA-damaging agents. Surprisingly, mRad17(5'Delta/5'Delta) ES cells were able to arrest cell cycle progression upon induction of DNA damage. However, they displayed impaired homologous recombination as evidenced by a strongly reduced gene targeting efficiency. In addition to a possible role in DNA damage-induced CCC, based on sequence homology, our results indicate that mRad17 has a function in DNA damage-dependent recombination that may be responsible for the sensitivity to DNA-damaging agents.     

The structure of the human ERCC1/XPF interaction domains reveals a complementary role for the two proteins in nucleotide excision repair

The human ERCC1/XPF complex is a structure-specific endonuclease with defined polarity that participates in multiple DNA repair pathways. We report the heterodimeric structure of the C-terminal domains of both proteins responsible for ERCC1/XPF complex formation. Both domains exhibit the double helix-hairpin-helix motif (HhH)2, and they are related by a pseudo-2-fold symmetry axis. In the XPF domain, the hairpin of the second motif is replaced by a short turn. The ERCC1 domain folds properly only in the presence of the XPF domain, which implies a role for XPF as a scaffold for the folding of ERCC1. The intersubunit interactions are largely hydrophobic in nature. NMR titration data show that only the ERCC1 domain of the ERCC1/XPF complex is involved in DNA binding. On the basis of these findings, we propose a model for the targeting of XPF nuclease via ERCC1-mediated interactions in the context of nucleotide excision repair.     

Attenuated XPC Expression Is Not Associated with Impaired DNA Repair in Bladder Cancer

Bladder cancer has a high incidence with significant morbidity and mortality. Attenuated expression of the DNA damage response protein Xeroderma Pigmentosum complementation group C (XPC) has been described in bladder cancer. XPC plays an essential role as the main initiator and damage-detector in global genome nucleotide excision repair (NER) of UV-induced lesions, bulky DNA adducts and intrastrand crosslinks, such as those made by the chemotherapeutic agent Cisplatin. Hence, XPC protein might be an informative biomarker to guide personalized therapy strategies in a subset of bladder cancer cases. Therefore, we measured the XPC protein expression level and functional NER activity of 36 bladder tumors in a standardized manner. We optimized conditions for dissociation and in vitro culture of primary bladder cancer cells and confirmed attenuated XPC expression in approximately 40% of the tumors. However, NER activity was similar to co-cultured wild type cells in all but one of 36 bladder tumors. We conclude, that (i) functional NER deficiency is a relatively rare phenomenon in bladder cancer and (ii) XPC protein levels are not useful as biomarker for NER activity in these tumors.     

Microbial biogeography of drinking water: patterns in phylogenetic diversity across space and time

In this study, we collected water from different locations in 32 drinking water distribution networks in the Netherlands and analysed the spatial and temporal variation in microbial community composition by high‐throughput sequencing of 16S rRNA gene amplicons. We observed that microbial community compositions of raw source and processed water were very different for each distribution network sampled. In each network, major differences in community compositions were observed between raw and processed water, although community structures of processed water did not differ substantially from end‐point tap water. End‐point water samples within the same distribution network revealed very similar community structures. Network‐specific communities were shown to be surprisingly stable in time. Biofilm communities sampled from domestic water metres varied distinctly between households and showed no resemblance to planktonic communities within the same distribution networks. Our findings demonstrate that high‐throughput sequencing provides a powerful and sensitive tool to probe microbial community composition in drinking water distribution systems. Furthermore, this approach can be used to quantitatively compare the microbial communities to match end‐point water samples to specific distribution networks. Insight in the ecology of drinking water distribution systems will facilitate the development of effective control strategies that will ensure safe and high‐quality drinking water.     

A systematic review of 3D scapular kinematics and muscle activity during elevation in stroke subjects and controls

Through the onset of post-stroke motor disorders, the normal scapular function is compromised. As a result, shoulder pain and associated upper limb dysfunctions frequently arise after stroke.This review aimed to provide a systematic overview of available literature on scapular function, i.e. scapular three-dimensional (3D) kinematics and muscle activity during elevation, in healthy persons, persons with primary shoulder disorders and post-stroke patients. 3D scapular kinematics have been widely reported in healthy persons and persons with primary shoulder disorders, whereby a general pattern of upward rotation and posterior tilt during elevation has been agreed upon. Results on scapular internal/external rotation are inconsistent. In a post-stroke population, 3D scapular kinematics are less frequently reported. Scapular muscle activity has thus far been studied to very limited extend and firm conclusions could not be drawn.Although 3D scapular kinematics and muscle activity registrations are being increasingly used, some general methodological aspects should be considered. While the International Society of Biomechanics already proposed recommendations on the definition of upper limb joint coordinate systems and rotation sequences, proper result comparison necessitates further guidelines on other methodological aspects, i.e. data collection, processing, analyzing, and reporting.