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Recruitment of DNA repair proteins to DNA damage sites is a critical step for DNA repair. Post-translational modifications of proteins at DNA damage sites serve as DNA damage codes to recruit specific DNA repair factors. Here, we show that mRNA is locally modified by m 5 C at sites of DNA damage. The RNA methyltransferase TRDMT1 is recruited to DNA damage sites to promote m 5 C induction. Loss of TRDMT1 compromises homologous recombination (HR) and increases cellular sensitivity to DNA double-strand breaks (DSBs). In the absence of TRDMT1, RAD51 and RAD52 fail to localize to sites of reactive oxygen species (ROS)-induced DNA damage. In vitro, RAD52 displays an increased affinity for DNA:RNA hybrids containing m 5 C-modified RNA. Loss of TRDMT1 in cancer cells confers sensitivity to PARP inhibitors in vitro and in vivo. These results reveal an unexpected TRDMT1-m 5 C axis that promotes HR, suggesting that post-transcriptional modifications of RNA can also serve as DNA damage codes to regulate DNA repair. Post-translational modifications of proteins at DNA damage sites can facilitate the recruitment of DNA repair factors. Here, the authors show that mRNA is locally modified with m 5 C at sites of DNA damage by the RNA methyltransferase TRDMT1 to promote homologous recombination repair.

Actively transcribed regions of the genome are protected by transcription-coupled DNA repair mechanisms, including transcription-coupled homologous recombination (TC-HR). Here we used reactive oxygen species (ROS) to induce and characterize TC-HR at a transcribed locus in human cells. As canonical HR, TC-HR requires RAD51. However, the localization of RAD51 to damage sites during TC-HR does not require BRCA1 and BRCA2, but relies on RAD52 and Cockayne Syndrome Protein B (CSB). During TC-HR, RAD52 is recruited by CSB through an acidic domain. CSB in turn is recruited by R loops, which are strongly induced by ROS in transcribed regions. Notably, CSB displays a strong affinity for DNA:RNA hybrids in vitro, suggesting that it is a sensor of ROS-induced R loops. Thus, TC-HR is triggered by R loops, initiated by CSB, and carried out by the CSB-RAD52-RAD51 axis, establishing a BRCA1/2-independent alternative HR pathway protecting the transcribed genome. Transcription-coupled homologous recombination (TC-HR) is activated by reactive oxygen species-induced DNA damage to maintain transcribed genome stability. The authors demonstrate that R loops are induced by ROS at the transcribed genome, triggering a CSB-RAD52- dependent but BRCA1/2-independent RAD51 loading for repair.

Damage repair mechanisms at transcriptionally active sites during the G0/G1 phase are largely unknown. To elucidate these mechanisms, we introduced genome site-specific oxidative DNA damage and determined the role of transcription in repair factor assembly. We find that KU and NBS1 are recruited to damage sites independent of transcription. However, assembly of RPA1, RAD51C, RAD51, and RAD52 at such sites is strictly governed by active transcription and requires both wild-type Cockayne syndrome protein B (CSB) function and the presence of RNA in the G0/G1 phase. We show that the ATPase activity of CSB is indispensable for loading and binding of the recombination factors. CSB counters radiation-induced DNA damage in both cells and zebrafish models. Taken together, our results have uncovered a novel, RNA-based recombination mechanism by which CSB protects genome stability from strand breaks at transcriptionally active sites and may provide insight into the clinical manifestations of Cockayne syndrome.

During the DNA damage response (DDR), ubiquitination plays an important role in the recruitment and regulation of repair proteins. However, little is known about elimination of the ubiquitination signal after repair is completed. Here we show that the ubiquitin-specific protease 5 (USP5), a deubiquitinating enzyme, is involved in the elimination of the ubiquitin signal from damaged sites and is required for efficient DNA double-strand break (DSB) repair. Depletion of USP5 sensitizes cells to DNA damaging agents, produces DSBs, causes delayed disappearance of γH2AX foci after Bleocin treatment, and influences DSB repair efficiency in the homologous recombination pathway but not in the non-homologous end joining pathway. USP5 co-localizes to DSBs induced by laser micro-irradiation in a RAD18-dependent manner. Importantly, polyubiquitin chains at sites of DNA damage remained for longer periods in USP5-depleted cells. Our results show that disassembly of polyubiquitin chains by USP5 at sites of damage is important for efficient DSB repair.

Obtaining a tenure track faculty position (TTFP) after postdoctoral appointment (PDA) completion is considered an indicator of successful transition to independence (TTI). Whether cross-institutional mobility (CIM)-moving to a different institution from that of the PDA-contributes to TTI is unclear, as data evaluating retention and mobility is lacking. We tested the hypothesis that, for postdocs (PDs) at R1 institutions, CIM is a significant predictor of successful TTI defined as TTFP-status 3 years post-PDA. Using University of Pittsburgh data for health sciences PDs we tested the association of CIM at PDA completion (moved to a different institution (CIM = 1) or retained at Pitt (CIM = 0)) with TTFP-status 3 years post-PDA (TTFP, non-TTFP, or left faculty position) using multinomial logistic regression models. Among all 622 Pitt PDs, 3-year retention in a faculty position at Pitt was 21%, while 14% had a faculty position outside of Pitt. Among the analytic sample of PDs with an academic career outcome during the study period (N = 238; 50% women, 8% underrepresented minorities (URM)), at baseline PDA completion 39% moved to a different institution (CIM = 1), and 61% remained at Pitt (CIM = 0) in any job type. Those with CIM = 1 had greater odds of having a TTFP at follow-up than those with CIM = 0 [adjusted OR (95% CI): 4.4 (2.1, 9.2)]. One fifth of Pitt PDs were retained by Pitt as faculty. While Pitt PDs were equally likely to get a faculty position whether they were retained at Pitt or left, those who left had greater odds of obtaining a TTFP. Future work with longer follow-up times, expanded markers of TTI, and samples from other R1 institutions is needed to better understand the reason for these results. This knowledge can lead to better support for the next generation of PDs as they successfully transition to faculty.

UV light-induced photoproducts are recognized and removed by the nucleotide-excision repair (NER) pathway. In humans, the UV-damaged DNA-binding protein (UV-DDB) is part of a ubiquitin E3 ligase complex (DDB1-CUL4A ᴰᴰᴮ²) that initiates NER by recognizing damaged chromatin with concomitant ubiquitination of core histones at the lesion. We report the X-ray crystal structure of the human UV-DDB in a complex with damaged DNA and show that the N-terminal domain of DDB2 makes critical contacts with two molecules of DNA, driving N-terminal-domain folding and promoting UV-DDB dimerization. The functional significance of the dimeric UV-DDB [(DDB1-DDB2) ₂], in a complex with damaged DNA, is validated by electron microscopy, atomic force microscopy, solution biophysical, and functional analyses. We propose that the binding of UV-damaged DNA results in conformational changes in the N-terminal domain of DDB2, inducing helical folding in the context of the bound DNA and inducing dimerization as a function of nucleotide binding. The temporal and spatial interplay between domain ordering and dimerization provides an elegant molecular rationale for the unprecedented binding affinities and selectivities exhibited by UV-DDB for UV-damaged DNA. Modeling the DDB1-CUL4A ᴰᴰᴮ² complex according to the dimeric UV-DDB-AP24 architecture results in a mechanistically consistent alignment of the E3 ligase bound to a nucleosome harboring damaged DNA. Our findings provide unique structural and conformational insights into the molecular architecture of the DDB1-CUL4A ᴰᴰᴮ² E3 ligase, with significant implications for the regulation and overall organization of the proteins responsible for initiation of NER in the context of chromatin and for the consequent maintenance of genomic integrity.

Medical students have higher rates of depression, suicidal ideation, and burnout than the general population. Schools have begun revisiting their procedures to ensure that they are doing everything possible to meet the mental health care needs of their students.

The patient-physician relationship, especially in the case of severely ill patients, is often fraught with anxiety, grief, and guilt in the physician who may come to feel that he or she has failed the patient and thereby becomes a “second victim.” This notion was first explored in a 1973 publication (Artiss and Levine N Engl J Med 288(23):1210-4, 1973 ) that described a novel interactive seminar series for oncology fellows that had been designed to address and possibly remedy the frequent disquiet experienced by young physicians in this setting. Fifty years later, the medical student co-authors of this Perspective enrolled in an elective course that comprised a similar series of interactive seminars, now addressing the contemporary patient-physician relationship. The earlier paper was employed as a historical background, and the framework of the course then broadened such that the students considered the current environmental changes in medical practice (social, cultural, financial, legal, policy) that may be linked to the character of individual patient-physician relationships. This essay reports on the students’ perception of such relationships, and on the environmental elements that may be helpful or harmful to the well-being of both patients and physicians.

Oxidative damage to telomeres leads to telomere attrition and genomic instability, resulting in poor cell viability. Telomere dynamics contribute to the maintenance of telomere integrity; however, whether oxidative damage induces telomere movement and how telomere mobility is regulated remain poorly understood. Here, we show that oxidative damage at telomeres triggers directional telomere movement. The presence of the human Sir2 homolog, Sirtuin 6 (SIRT6) is required for oxidative damage-induced telomeric movement. SIRT6 knock out (KO) cells show neither damage-induced telomere movement nor chromatin decondensation at damaged telomeres; both are observed in wild type (WT) cells. A deacetylation mutant of SIRT6 increases damage-induced telomeric movement in SIRT6 KO cells as well as WT SIRT6. SIRT6 recruits the chromatin-remodeling protein SNF2H to damaged telomeres, which appears to promote chromatin decondensation independent of its deacetylase activity. Together, our results suggest that SIRT6 plays a role in the regulation of telomere movement upon oxidative damage, shedding new light onto the function of SIRT6 in telomere maintenance.

PurposeBuilding leadership skills among faculty in academic medicine is essential, yet professional development programs focused on leadership are not always attentive to the needs of faculty on diverse career pathways or at differing career stages—nor are they often rigorously assessed. Evaluations commonly focus on participant satisfaction and short-term learning but not behavior change and institutional impact, which are difficult to assess but arguably more meaningful. Given the substantial time and money invested in these programs, more rigorous evaluation is critical.Design/methodology/approachThe authors evaluated an intensive, shared leadership-focused training program for early-career and mid-career faculty, offered by the University of Pittsburgh’s School of Medicine over the course of a year. They administered a pre/post-program assessment of confidence in key skill areas, and conducted semi-structured interviews with 21 participants between 1–4 years after program completion.FindingsParticipants in both programs showed statistically significant improvement (p < 0.001) on every item measured in the pre/post-test. Analysis of the interviews revealed indications of substantial behavior change as well as institutional impact. The evaluation also suggested particular benefits for female professionals.Originality/valueThe authors conducted a long-term assessment of leadership training focused on career pathway and career stage and found that it (a) prompted both positive behavioral change and institutional impact and (b) suggested benefits for female faculty in particular, which could potentially help to eliminate gender-based disparities in leadership in academic medical centers.