A Bacterial Cytotoxin Identifies the RhoA Exchange Factor Net1 as a Key Effector in the Response to DNA Damage

Background: Exposure of adherent cells to DNA damaging agents, such as the bacterial cytolethal distending toxin (CDT) or ionizing radiations (IR), activates the small GTPase RhoA, which promotes the formation of actin stress fibers and delays cell death. The signalling intermediates that regulate RhoA activation and promote cell survival are unknown. Principal Findings: We demonstrate that the nuclear RhoA-specific Guanine nucleotide Exchange Factor (GEF) Net1 becomes dephosphorylated at a critical inhibitory site in cells exposed to CDT or IR. Expression of a dominant negative Net1 or Net1 knock down by iRNA prevented RhoA activation, inhibited the formation of stress fibers, and enhanced cell death, indicating that Net1 activation is required for this RhoA-mediated responses to genotoxic stress. The Net1 and RhoAdependent signals involved activation of the Mitogen-Activated Protein Kinase p38 and its downstream target MAPKactivated protein kinase 2. Significance: Our data highlight the importance of Net1 in controlling RhoA and p38 MAPK mediated cell survival in cells exposed to DNA damaging agents and illustrate a molecular pathway whereby chronic exposure to a bacterial toxin ma

Reliability and Validity of the Disability Assessment Structured Interview (DASI): A Tool for Assessing Functional Limitations in Claimants

Objective The aim of this study is to investigate the reliability and validity of the Disability Assessment Structured Interview (DASI). The DASI is a semi-structured interview for assessing long-term functional limitations concerning the work disability assessment of claimants. Methods A randomized controlled trial was conducted. Patients applying for a work-disability pension after 21 months of sick leave were independently interviewed and examined either by two physicians who had completed a DASI training period (n = 32) or by two physicians from a control group (n = 30) without any DASI training. Agreement percentages within both groups of physicians, eligibility for a disability benefit, and differences between the groups in terms of the scores given on the work-limitation items from the Functional Ability List (FAL) were measured to investigate reliability and concurrent validity. To determine the content validity, the insurance physicians who completed DASI training (n = 8) were asked to fill out a questionnaire concerning their opinion of the DASI. Additionally, patients filled out a questionnaire to measure their satisfaction as to the behavioral aspects of the physicians. Results The groups showed no important differences in agreement percentages (mean percentage about 80%) and eligibility for a disability benefit. In 9 out of 21 items the physicians of the control group indicated fewer work limitations compared to physicians using the DASI. All physicians agreed on the fact that the DASI was an acceptable tool in daily practice, one that provided a realistic picture of the patient and provided sufficient information to assess functional limitations. In addition, between the two groups, no differences were found as to the satisfaction of patients concerning the behavioral aspects of the physicians. Conclusion The DASI is a tool with a reasonable to good inter-rater reliability and content validity, and it appears to be acceptable to both patients and physicians. It did not improve inter-observer agreement beyond that of usual interview procedures used in the Netherlands. The DASI would seem to be a worthwhile tool for collecting self-reported information in order to assess functional limitations in claimants

Predisposition to Cancer Caused by Genetic and Functional Defects of Mammalian Atad5

ATAD5, the human ortholog of yeast Elg1, plays a role in PCNA deubiquitination. Since PCNA modification is important to regulate DNA damage bypass, ATAD5 may be important for suppression of genomic instability in mammals in vivo. To test this hypothesis, we generated heterozygous (Atad5+/m) mice that were haploinsuffficient for Atad5. Atad5+/m mice displayed high levels of genomic instability in vivo, and Atad5+/m mouse embryonic fibroblasts (MEFs) exhibited molecular defects in PCNA deubiquitination in response to DNA damage, as well as DNA damage hypersensitivity and high levels of genomic instability, apoptosis, and aneuploidy. Importantly, 90% of haploinsufficient Atad5+/m mice developed tumors, including sarcomas, carcinomas, and adenocarcinomas, between 11 and 20 months of age. High levels of genomic alterations were evident in tumors that arose in the Atad5+/m mice. Consistent with a role for Atad5 in suppressing tumorigenesis, we also identified somatic mutations of ATAD5 in 4.6% of sporadic human endometrial tumors, including two nonsense mutations that resulted in loss of proper ATAD5 function. Taken together, our findings indicate that loss-of-function mutations in mammalian Atad5 are sufficient to cause genomic instability and tumorigenesis