REC-1 and HIM-5 distribute meiotic crossovers and function redundantly in meiotic double-strand break formation in Caenorhabditis elegans.

The Caenorhabditis elegans gene rec-1 was the first genetic locus identified in metazoa to affect the distribution of meiotic crossovers along the chromosome. We report that rec-1 encodes a distant paralog of HIM-5, which was discovered by whole-genome sequencing and confirmed by multiple genome-edited alleles. REC-1 is phosphorylated by cyclin-dependent kinase (CDK) in vitro, and mutation of the CDK consensus sites in REC-1 compromises meiotic crossover distribution in vivo. Unexpectedly, rec-1; him-5 double mutants are synthetic-lethal due to a defect in meiotic double-strand break formation. Thus, we uncovered an unexpected robustness to meiotic DSB formation and crossover positioning that is executed by HIM-5 and REC-1 and regulated by phosphorylation.

Methodological considerations for mutagen exposure in C. elegans.

Maintenance of the genome requires the continual repair of DNA lesions. Exposure of nematodes to DNA damage-inducing agents is a powerful method to rapidly ascribe a role for specific genes in DNA repair and to define epistatic relationships to other repair genes which allows for the construction of repair pathways. Despite the extensive use of these agents, however, differences in dosing, timing, and handling makes it difficult to compare results across laboratories. We provide herein a consideration of the parameters that influence the results of these exposures and detailed protocols for the exposure to mutagenic inducing agents.

Idelalisib for Treatment of Relapsed Follicular Lymphoma and Chronic Lymphocytic Leukemia: A Comparison of Treatment Outcomes in Clinical Trial Participants vs Medicare Beneficiaries.

Importance: Idelalisib (IDEL) is approved as monotherapy in relapsed follicular lymphoma (FL) and with rituximab (IDEL+R) for relapsed chronic lymphocytic leukemia (CLL). Toxic effects can be severe and treatment-limiting. Outcomes in a real-world population are not yet characterized. Objective: We compared IDEL treatment outcomes in the clinical setting with outcomes in clinical trial data. Design, Setting, and Participants: This cohort study compared clinical trial participants treated with IDEL, aged 65 years or older, in studies 101-09 and 312-0116 with Medicare beneficiaries treated with IDEL of the same disease state and treatment regimen. Study 101-09 was a phase 2, single-group, open-label trial supporting accelerated approval of IDEL for relapsed or refractory FL. Study 312-0116 was a phase 3, multicenter, randomized, double-blind trial supporting approval of IDEL+R for relapsed CLL. Analyses were conducted between February and December 2018. Main Outcomes and Measures: Treatment duration, on-treatment and overall mortality, and serious and fatal infections were compared between trial participants and Medicare beneficiaries. Cox proportional hazards models quantified differences by cohort. Results: We identified 26 trial participants (mean [SD] age, 73 [4.9] years; 12 [46.2%] women) and 305 Medicare beneficiaries (mean [SD] age, 76 [6.9] years; 103 [54.8%] women) receiving IDEL for FL and 89 trial participants (mean [SD] age, 74 [6.0] years; 30 [33.7%] women) and 294 Medicare beneficiaries (mean age, 76 [6.3] years; 111 [37.8%] women) receiving IDEL+R for CLL. Medicare beneficiaries were older with higher comorbidity; had a shorter median treatment duration for CLL (173 days vs 473 days, P < .001) but not FL (114, days vs 160 days, P = .38); a numerically higher mortality rate (CLL: HR, 1.40; 95% CI, 0.93-2.11; FL: HR, 1.39; 95% CI, 0.69-2.78); and a significantly higher fatal infection rate per 100 person-years for CLL (18.4 vs 9.8, P = .04) and a numerically higher rate for FL (27.6 vs 18.6, P = .54), compared with trial participants. Trial participants had approximately twice as many dose reductions (CLL: 32.6% vs 18.0%; P = .003; FL: 38.5% vs 16.1%; P = .02). Among Medicare beneficiaries, a hospitalized infection within 6 months prior to IDEL initiation was associated with a 2.11-fold increased risk for on-treatment fatal infections (95% CI, 1.44-3.10). Despite a March 2016 recommendation for Pneumocystis jirovecii pneumonia prophylaxis in patients treated with IDEL, prophylaxis rates were low after March 2016 (FL: 25%, CLL: 37%). Conclusions and Relevance: We observed substantial imbalances in baseline comorbidities and treatment outcomes between Medicare beneficiaries and trial participants aged 65 years or older. Immunosuppression-related toxic effects, including infections, may have been somewhat reduced in trials by more frequent dose reductions and exclusion of patients with ongoing infections. Selective eligibility criteria and closer monitoring of trial patients may be responsible for limited generalizability of trial data to clinical practice.

Meiotic Double-Strand Break Proteins Influence Repair Pathway Utilization.

Double-strand breaks (DSBs) are among the most deleterious lesions DNA can endure. Yet, DSBs are programmed at the onset of meiosis, and are required to facilitate appropriate reduction of ploidy in daughter cells. Repair of these breaks is tightly controlled to favor homologous recombination (HR)-the only repair pathway that can form crossovers. However, little is known about how the activities of alternative repair pathways are regulated at these stages. We discovered an unexpected synthetic interaction between the DSB machinery and strand-exchange proteins. Depleting the Caenorhabditis elegans DSB-promoting factors HIM-5 and DSB-2 suppresses the formation of chromosome fusions that arise in the absence of RAD-51 or other strand-exchange mediators. Our investigations reveal that nonhomologous and theta-mediated end joining (c-NHEJ and TMEJ, respectively) and single strand annealing (SSA) function redundantly to repair DSBs when HR is compromised, and that HIM-5 influences the utilization of TMEJ and SSA.

The p53-like Protein CEP-1 Is Required for Meiotic Fidelity in C. elegans.

The passage of genetic information during meiosis requires exceptionally high fidelity to prevent birth defects and infertility. Accurate chromosome segregation during the first meiotic division relies on the formation of crossovers between homologous chromosomes and a series of precisely controlled steps to exchange genetic information. Many studies have hinted at a role for p53 in meiosis, but how it functions in this process is poorly understood. Here, we have identified a cooperative role for the p53-like protein CEP-1 and the meiotic protein HIM-5 in maintaining genome stability in the C. elegans germline. Loss of cep-1 and him-5 results in synthetic lethality that is dependent on the upstream DNA damage checkpoint but independent of the downstream core apoptotic pathway. We show that this synthetic lethality is the result of defective crossover formation due to reduced SPO-11-dependent double-strand breaks. Using cep-1 separation-of-function alleles, we show that cep-1 and him-5 also suppress inappropriate activation of the nonhomologous end joining (NHEJ) pathway. This work reveals an ancestral function for the p53 family in ensuring the fidelity of meiosis and establishes CEP-1 as a critical determinant of repair pathway choice.