Homology Requirements and Competition between Gene Conversion and Break-Induced Replication during Double-Strand Break Repair.

Saccharomyces cerevisiae mating-type switching is initiated by a double-strand break (DSB) at MATa, leaving one cut end perfectly homologous to the HMLα donor, while the second end must be processed to remove a non-homologous tail before completing repair by gene conversion (GC). When homology at the matched end is ≤150 bp, efficient repair depends on the recombination enhancer, which tethers HMLα near the DSB. Thus, homology shorter than an apparent minimum efficient processing segment can be rescued by tethering the donor near the break. When homology at the second end is ≤150 bp, second-end capture becomes inefficient and repair shifts from GC to break-induced replication (BIR). But when pol32 or pif1 mutants block BIR, GC increases 3-fold, indicating that the steps blocked by these mutations are reversible. With short second-end homology, absence of the RecQ helicase Sgs1 promotes gene conversion, whereas deletion of the FANCM-related Mph1 helicase promotes BIR.

Identifying, Addressing, and Eliminating Microaggressions in Healthcare.

Description Microaggressions are pervasive throughout society, including in healthcare and academic institutions. They are often unconscious but accumulate over time, and they negatively impact the recipients' productivity and achievement by creating a sense of inadequacy as well as a lack of belonging. We outline several evidence-based strategies and teaching frameworks that institutions and training programs can adopt to reduce the prevalence and impact of microaggressions against trainees from historically marginalized groups, and that can promote psychological safety for everyone.

Analysis of post-operative changes in serum protein expression profiles from colorectal cancer patients by MALDI-TOF mass spectrometry: a pilot methodological study.

BACKGROUND: Mass spectrometry-based protein expression profiling of blood sera can be used to discriminate colorectal cancer (CRC) patients from unaffected individuals. In a pilot methodological study, we have evaluated the changes in protein expression profiles of sera from CRC patients that occur following surgery to establish the potential of this approach for monitoring post-surgical response and possible early prediction of disease recurrence. METHODS: In this initial pilot study, serum specimens from 11 cancer patients taken immediately prior to surgery and at approximately 6 weeks following surgery were analysed alongside 10 normal control sera by matrix-assisted laser desorption ionisation time of-flight-mass spectrometry (MALDI-TOF MS). Using a two-sided t-test the top 20 ranked protein peaks that discriminate normal from pre-operative sera were identified. These were used to classify post-operative sera by hierarchical clustering analysis (Spearman's Rank correlation) and, as an independent 'test' dataset, by k-nearest neighbour and weighted voting supervised learning algorithms. RESULTS: Hierarchical cluster analysis classified post-operative sera from all six early Dukes' stage (A and B) patients as normal. The remaining five post-operative sera from more advanced Dukes' stages (C1 and C2) were classified as cancer. Analysis by supervised learning algorithms similarly grouped all advanced Dukes' stages as cancer, with four of the six post-operative sera from early Dukes' stages being classified as normal (P = 0.045; Fisher's exact test). CONCLUSIONS: The results of this pilot methodological study illustrate the proof-of-concept of using protein expression profiling of post-surgical blood sera from individual patients to monitor disease course. Further validation on a larger patient cohort and using an independent post-operative sera dataset would be required to evaluate the potential clinical relevance of this approach. Prospective data, including follow-up on patient survival, could in the future, then be evaluated to inform decisions on individualised treatment modalities.

A Pilot Study Examining Biofeedback and Structured Napping to Promote Medical Student Wellbeing.

This article was migrated. The article was marked as recommended. Objective: To examine medical students' engagement in wellness activities and evaluate the effects of biofeedback and structured napping on measures of stress, burnout and wellbeing. Method: A randomized trial of heart-rate variability (HRV) biofeedback and structured napping used by pre-clinical medical students at the University of Central Florida College of Medicine compared with a control group was conducted. Baseline measurement occurred in August 2016 with the follow-up period in March 2017. To measure biofeedback, participants used Heartmath Biofeedback® with Inner Balance® software to record HRV measurements while they engaged in self-guided breathing three times weekly. The biofeedback device connected to participants' iPhone or iPad with a sensor that clipped to users' earlobes. HRV recordings were stored in a heart-cloud database, and participants had the option to share their recordings with the researchers. Participants used sleep pods (MetroNaps Energy Pods®) to engage in 20-minute structured naps three times weekly. Participants completed six psychosocial self-report questionnaires at baseline (T1) and two follow-up points (T2, T3). The questionnaires included the Interpersonal Reactivity Index; Perceived Stress Scale; Quality of life scale; Oldenburg Burnout Inventory; and the Physician Well-Being Index. Results: Forty-two students enrolled in the study. Throughout the study, participants recorded 276 structured naps lasting approximately 20 minutes in duration and shared 24 personalized biofeedback recordings. Conclusions: Promotion of structured napping offers promise as an institution-initiated wellness intervention to promote medical students' mental health and wellbeing. HRV biofeedback warrants further study given the lack of conclusive findings in this study.

Genetic screening for novel Drosophila mutants with discrepancies in iron metabolism.

Ferritin, a symmetrical 24-subunit heteropolymer composed of heavy and light chains, is the primary iron-storage molecule in bacteria, plants and animals. We used a genetically engineered strain of the model organism Drosophila melanogaster which expresses a GFP (green fluorescent protein)-tagged ferritin 1 heavy chain homologue from its native chromosomal locus and incorporated it into endogenous functional ferritin, enabling in vivo visualization of the protein and permitting easy assessment of ferritin status following environmental or genetic perturbations. Random mutagenesis was induced, and individual mutagenized chromosomes were recovered by classic crossing schemes involving phenotypical markers and balancer chromosomes. In wild-type larvae, ferritin is predominantly localized in the brain, in regions of the intestine, in wreath cells and in pericardial cells. A pilot genetic screen revealed a mutant fruitfly strain expressing GFP-ferritin in the anal pads, a pair of organs located ventrally in the posterior end of the fruitfly larva, possibly involved in ion absorption and osmoregulation, which are normally devoid of ferritin. Our continuing genetic screen could reveal transcription factors involved in ferritin regulation and novel proteins important in iron metabolism, hopefully with conserved functions in evolution.

Sgs1 and Mph1 Helicases Enforce the Recombination Execution Checkpoint During DNA Double-Strand Break Repair in Saccharomyces cerevisiae.

We have previously shown that a recombination execution checkpoint (REC) regulates the choice of the homologous recombination pathway used to repair a given DNA double-strand break (DSB) based on the homology status of the DSB ends. If the two DSB ends are synapsed with closely-positioned and correctly-oriented homologous donors, repair proceeds rapidly by the gene conversion (GC) pathway. If, however, homology to only one of the ends is present, or if homologies to the two ends are situated far away from each other or in the wrong orientation, REC blocks the rapid initiation of new DNA synthesis from the synapsed end(s) and repair is carried out by the break-induced replication (BIR) machinery after a long pause. Here we report that the simultaneous deletion of two 3'→5' helicases, Sgs1 and Mph1, largely abolishes the REC-mediated lag normally observed during the repair of large gaps and BIR substrates, which now get repaired nearly as rapidly and efficiently as GC substrates. Deletion of SGS1 and MPH1 also produces a nearly additive increase in the efficiency of both BIR and long gap repair; this increase is epistatic to that seen upon Rad51 overexpression. However, Rad51 overexpression fails to mimic the acceleration in repair kinetics that is produced by sgs1Δ mph1Δ double deletion.

Recurrence of conversion disorder symptoms in a successfully treated 16-year-old female.

We present a case of a 16-year-old Caucasian female with a history of major depressive disorder and post-traumatic stress disorder who was admitted to an inpatient adolescent psychiatric unit with symptoms of conversion disorder, including non-epileptic seizures, an inability to speak or walk, and not eating on her own. She has a history of multiple previous medical and psychiatric hospitalizations without any significant resolution of symptoms, and extensive medical workups have all been negative. Treatment ultimately involved reassuring the patient and family that there was no underlying medical condition and emphasizing the conversion disorder diagnosis. The patient participated daily in physical therapy to improve mobility, deconditioning, and functioning. Hospital staff was instructed on the nature of the non-epileptic seizures, which continued to occur during the hospitalization. After one month, the patient was discharged home fully functional: walking, speaking, and eating on her own. One week after discharge, the patient presented with the same symptoms and was readmitted to the psychiatric facility. She subsequently never regained her previous level of functioning, and she was ultimately transferred to a residential treatment facility. We will discuss factors that led to the initial improvement and the factors that led to recurrence and persistence of symptoms.

Sources of DNA double-strand breaks and models of recombinational DNA repair.

DNA is subject to many endogenous and exogenous insults that impair DNA replication and proper chromosome segregation. DNA double-strand breaks (DSBs) are one of the most toxic of these lesions and must be repaired to preserve chromosomal integrity. Eukaryotes are equipped with several different, but related, repair mechanisms involving homologous recombination, including single-strand annealing, gene conversion, and break-induced replication. In this review, we highlight the chief sources of DSBs and crucial requirements for each of these repair processes, as well as the methods to identify and study intermediate steps in DSB repair by homologous recombination.