Search

Movie 1. Time-lapse imaging of MDA-MB-231 cells in complete medium.

Movie 2. Time-lapse imaging of MDA-MB-231 cells in glucose-depleted medium.

Movie 3. Time-lapse imaging of MDA-MB-231/shKAP1/S473A-KAP1 cells in glucose-depleted medium.

Mitochondrial dynamics during nutrient starvation of cancer cells likely exert profound effects on their capability for metastatic progression. Here, we report that KAP1 (TRIM28), a transcriptional coadaptor protein implicated in metastatic progression in breast cancer, is a pivotal regulator of mitochondrial fusion in glucose-starved cancer cells. Diverse metabolic stresses induced Ser473 phosphorylation of KAP1 (pS473-KAP1) in a ROS- and p38-dependent manner. Results from live-cell imaging and molecular studies revealed that during the first 6 to 8 hours of glucose starvation, mitochondria initially underwent extensive fusion, but then subsequently fragmented in a pS473-KAP1-dependent manner. Mechanistic investigations using phosphorylation-defective mutants revealed that KAP1 Ser473 phosphorylation limited mitochondrial hyperfusion in glucose-starved breast cancer cells, as driven by downregulation of the mitofusin protein MFN2, leading to reduced oxidative phosphorylation and ROS production. In clinical specimens of breast cancer, reduced expression of MFN2 corresponded to poor prognosis in patients. In a mouse xenograft model of human breast cancer, there was an association in the core region of tumors between MFN2 downregulation and the presence of highly fragmented mitochondria. Collectively, our results suggest that KAP1 Ser473 phosphorylation acts through MFN2 reduction to restrict mitochondrial hyperfusion, thereby contributing to cancer cell survival under conditions of sustained metabolic stress. Cancer Res; 76(17); 5006–18. ©2016 AACR.

Movie 4. Time-lapse imaging of MDA-MB-231/shKAP1/S473D-KAP1 cells in glucose-depleted medium.

Supplementary Figure S1. KAP1 transcript level is positively associated with some of mitochondrial complex genes expression in breast cancer patients. Supplementary Figure S2. KAP1 overexpression is associated with breast cancer poor prognosis. Supplementary Figure S3. p38MAPK, but not AMPK or ATM, activates metabolic stress-induced KAP1 phosphorylation at Ser473. Supplementary Figure S4. Time-dependent alterations of Mitochondrial dynamics upon glucose depletion. Supplementary Figure S5. S473A-KAP1 promotes mitochondrial network and sensitizes MDA-MB-231/shKAP1 cells to glucose starvation.

This file includes supplementary figure legends and movie legends.

Physicians, judges, teachers, and agents in many other settings differ systematically in the decisions they make when faced with similar cases. Standard approaches to interpreting and exploiting such differences assume they arise solely from variation in preferences. We develop an alternative framework that allows variation in preferences and diagnostic skill and show that both dimensions may be partially identified in standard settings under quasi-random assignment. We apply this framework to study pneumonia diagnoses by radiologists. Diagnosis rates vary widely among radiologists, and descriptive evidence suggests that a large component of this variation is due to differences in diagnostic skill. Our estimated model suggests that radiologists view failing to diagnose a patient with pneumonia as more costly than incorrectly diagnosing one without, and that this leads less skilled radiologists to optimally choose lower diagnostic thresholds. Variation in skill can explain 39% of the variation in diagnostic decisions, and policies that improve skill perform better than uniform decision guidelines. Failing to account for skill variation can lead to highly misleading results in research designs that use agent assignments as instruments.

Cristae membrane state plays a central role in regulating mitochondrial function and cellular metabolism. The protein Optic atrophy 1 (Opa1) is an important crista remodeler that exists as two forms in the mitochondrion, a membrane-anchored long form (l-Opa1) and a processed short form (s-Opa1). The mechanisms for how Opa1 influences cristae shape have remained unclear due to the lack of native 3D views of cristae morphology. We performin situcryo-electron tomography of cryo-focused ion beam milled mouse embryonic fibroblasts with well-defined Opa1 states to understand how each form of Opa1 influences cristae architecture. In our tomograms, we observe elongated mitochondria with a notable stacking phenotype, as well as an absence of tubular cristae, when only l-Opa1 is present. In contrast, when mitochondria contain mainly s-Opa1, we observe irregular cristae packing, an increase in globular cristae, and decreased matrix condensation. Notably, we find the absence of l-Opa1 results in mitochondria with wider cristae junctions. BH3 profiling reveals that absence of l-Opa1 reduces cytochrome c release in response to pro-apoptotic stimuli and protects cells from apoptosis induced by anti-cancer agents. We discuss the implications Opa1-dependent cristae morphologies in cell death initiation.HighlightsIn situultrastructural characterization of mitochondrial cristae with different forms of Opa1.Mitochondria with predominantly l-Opa1 show cristae stacking, longer cristae compared to WT, but also a reduction of globular cristae and no tubular cristae.Mitochondria with mostly s-Opa1 showed irregular cristae packing with wider cristae junctions and more narrow cristae than WT.BH3 profiling show Opa1-knock-out cells have reduced apoptotic priming and reduced sensitivity to apoptosis-inducing agents, and the presence l-Opa1 restores a WT protective apoptotic response.

I study team decisions among physician trainees. Exploiting a discontinuity in team roles across trainee tenure, I find evidence that teams alter decision-making, concentrating influence in the hands of senior trainees. I also demonstrate little convergence in variation of trainee effects despite intensive training. This general pattern of trainee effects on team decision-making exists in all types of decisions and settings that I examine. In analyses evaluating mechanisms behind this pattern, I find support for the idea that significant experiential learning occurs during training and that teams place more weight on judgments of senior trainees in order to aggregate information. (JEL D83, I11, J44, M53, M54)

Diabetic kidney disease (DKD) is a major complication of diabetes. Expression of members of the microRNA (miRNA) miR-379 cluster is increased in DKD. miR-379, the most upstream 5′-miRNA in the cluster, functions in endoplasmic reticulum (ER) stress by targeting EDEM3. However, the in vivo functions of miR-379 remain unclear. We created miR-379 knockout (KO) mice using CRISPR-Cas9 nickase and dual guide RNA technique and characterized their phenotype in diabetes. We screened for miR-379 targets in renal mesangial cells from WT vs. miR-379KO mice using AGO2-immunopreciptation and CLASH (cross-linking, ligation, sequencing hybrids) and identified the redox protein thioredoxin and mitochondrial fission-1 protein. miR-379KO mice were protected from features of DKD as well as body weight loss associated with mitochondrial dysfunction, ER- and oxidative stress. These results reveal a role for miR-379 in DKD and metabolic processes via reducing adaptive mitophagy. Strategies targeting miR-379 could offer therapeutic options for DKD., Kato, Abdollahi et al. identify a redox protein thioredoxin and mitochondrial fission-1 (FIS1) protein as miR-379 targets in mouse kidney. They find that miR-379 knockout mice are protected from diabetic kidney disease by enhancing mitophagy via FIS1, suggesting miR-379 as a potential therapeutic target for diabetic kidney disease.

One of the most challenging environments in health care is the emergency department (ED) A key decision-maker in that context is triage nurses, who assess patient illness severity and influence wait times for medical attention. We gather novel data on the triage process across 108 EDs, including wait times, triage nurse identities and assessments, and detailed patient information and outcomes. Using quasi-random assignment to ED, we find a striking rate of “inversions,” where patients who are sicker based on either ex ante information or ex post outcomes are scored as sicker and wait longer than their healthier counterparts.

Male germline development involves choreographed changes to mitochondrial number, morphology and organization. Mitochondrial reorganization during spermatogenesis was recently shown to require mitochondrial fusion and fission. Mitophagy, the autophagic degradation of mitochondria, is another mechanism for controlling mitochondrial number and physiology, but its role during spermatogenesis is largely unknown. During post-meiotic spermatid development, restructuring of the mitochondrial network results in packing of mitochondria into a tight array in the sperm midpiece to fuel motility. Here, we show that disruption of mouse Fis1 in the male germline results in early spermatid arrest that is associated with increased mitochondrial content. Mutant spermatids coalesce into multinucleated giant cells that accumulate mitochondria of aberrant ultrastructure and numerous mitophagic and autophagic intermediates, suggesting a defect in mitophagy. We conclude that Fis1 regulates mitochondrial morphology and turnover to promote spermatid maturation., Summary: Analysis of germ cell-specific Fis1 knockout mice and mitophagy reporter mice reveals that the mitochondrial dynamics gene Fis1 regulates mitochondrial morphology and turnover during spermatid maturation.

In setting prices for physician services, Medicare solicits input from a committee that evaluates proposals from industry. The committee itself comprises members from industry; we investigate whether this arrangement leads to regulatory capture with prices biased toward industry interests. We find that increasing a measure of affiliation between the committee and proposers by one standard deviation increases prices by 10%. We then evaluate whether employing a biased committee as an intermediary may nonetheless be desirable, if greater affiliation allows the committee to extract information needed for regulation. We find industry proposers more affiliated with the committee produce less hard evidence in their proposals. However, on soft information, we find evidence of a trade-off: private insurers set prices that more closely track Medicare prices generated under higher affiliation.

Diabetic kidney disease (DKD) is a major complication of diabetes. Expression of members of the microRNA (miRNA) miR-379 cluster is increased in DKD. miR-379, the most upstream 5'-miRNA in the cluster, functions in endoplasmic reticulum (ER) stress by targeting EDEM3. However, the in vivo functions of miR-379 remain unclear. We created miR-379 knockout (KO) mice using CRISPR-Cas9 nickase and dual guide RNA technique and characterized their phenotype in diabetes. We screened for miR-379 targets in renal mesangial cells from WT vs. miR-379KO mice using AGO2-immunopreciptation and CLASH (cross-linking, ligation, sequencing hybrids) and identified the redox protein thioredoxin and mitochondrial fission-1 protein. miR-379KO mice were protected from features of DKD as well as body weight loss associated with mitochondrial dysfunction, ER- and oxidative stress. These results reveal a role for miR-379 in DKD and metabolic processes via reducing adaptive mitophagy. Strategies targeting miR-379 could offer therapeutic options for DKD.

OPA1, a large GTPase of the dynamin superfamily, mediates fusion of the mitochondrial inner membranes, regulates cristae morphology, and maintains respiratory chain function. Inner membrane–anchored long forms of OPA1 (l-OPA1) are proteolytically processed by the OMA1 or YME1L proteases, acting at cleavage sites S1 and S2, respectively, to produce short forms (s-OPA1). In both mice and humans, half of the mRNA splice forms of Opa1 are constitutively processed to yield exclusively s-OPA1. However, the function of s-OPA1 in mitochondrial fusion has been debated, because in some stress conditions, s-OPA1 is dispensable for fusion. By constructing cells in which the Opa1 locus no longer produces transcripts with S2 cleavage sites, we generated a simplified system to identify the new YME1L-dependent site S3 that mediates constitutive and complete cleavage of OPA1. We show that mitochondrial morphology is highly sensitive to the ratio of l-OPA1 to s-OPA1, indicating that s-OPA1 regulates mitochondrial fusion.

ObjectiveTo measure and compare mortality outcomes between dually eligible veterans transported by ambulance to a Veterans Affairs hospital and those transported to a non-Veterans Affairs hospital.DesignRetrospective cohort study using data from medical charts and administrative files.SettingEmergency visits by ambulance to 140 Veteran Affairs and 2622 non-Veteran Affairs hospitals across 46 US states and the District of Columbia in 2001-18.ParticipantsNational cohort of 583 248 veterans (aged ≥65 years) enrolled in both the Veterans Health Administration and Medicare programs, who resided within 20 miles of at least one Veterans Affairs hospital and at least one non-Veterans Affairs hospital, in areas where ambulances regularly transported patients to both types of hospitals.InterventionEmergency treatment at a Veterans Affairs hospital.Main outcome measureDeaths in the 30 day period after the ambulance ride. Linear probability models of mortality were used, with adjustment for patients’ demographic characteristics, residential zip codes, comorbid conditions, and other variables.ResultsOf 1 470 157 ambulance rides, 231 611 (15.8%) went to Veterans Affairs hospitals and 1 238 546 (84.2%) went to non-Veterans Affairs hospitals. The adjusted mortality rate at 30 days was 20.1% lower among patients taken to Veterans Affairs hospitals than among patients taken to non-Veterans Affairs hospitals (9.32 deaths per 100 patients (95% confidence interval 9.15 to 9.50)v11.67 (11.58 to 11.76)). The mortality advantage associated with Veterans Affairs hospitals was particularly large for patients who were black (−25.8%), were Hispanic (−22.7%), and had received care at the same hospital in the previous year.ConclusionsThese findings indicate that within a month of being treated with emergency care at Veterans Affairs hospitals, dually eligible veterans had substantially lower risk of death than those treated at non-Veterans Affairs hospitals. The nature of this mortality advantage warrants further investigation, as does its generalizability to other types of patients and care. Nonetheless, the finding is relevant to assessments of the merit of policies that encourage private healthcare alternatives for veterans.

Mitochondrial fusion and fission (mitochondrial dynamics) are homeostatic processes that safeguard normal cellular function. This relationship is especially strong in tissues with constitutively high energy demands, such as brain, heart and skeletal muscle. Less is known about the role of mitochondrial dynamics in developmental systems that involve changes in metabolic function. One such system is spermatogenesis. The first mitochondrial dynamics gene, Fuzzy onions (Fzo), was discovered in 1997 to mediate mitochondrial fusion during Drosophila spermatogenesis. In mammals, however, the role of mitochondrial fusion during spermatogenesis remained unknown for nearly two decades after discovery of Fzo. Mammalian spermatogenesis is one of the most complex and lengthy differentiation processes in biology, transforming spermatogonial stem cells into highly specialized sperm cells over a 5-week period. This elaborate differentiation process requires several developmentally regulated mitochondrial and metabolic transitions, making it an attractive model system for studying mitochondrial dynamics in vivo. We review the emerging role of mitochondrial biology, and especially its dynamics, during the development of the male germ line.

Expert decision-makers often deviate from evidence-based guidelines. If experts lack awareness of guidelines, dissemination may improve outcomes. If experts who are aware of guidelines continue to deviate, promoting stricter adherence has ambiguous effects on outcomes, depending on whether experts use information not encoded in guidelines. We study the role of guidelines in physician decisions to prescribe anticoagulants for atrial fibrillation patients. Anticoagulation reduces stroke risk but increases hemorrhage risk. By text-mining physician notes for guideline mentions, we identify when each physician becomes aware of a new guideline. Once aware of the guideline, physicians pivot their practice patterns to align more closely with the recommendations, but guideline adherence remains far from perfect. To evaluate whether continued non-adherence reflects physicians’ superior information, we combine observational data on treatment choices with new machine learning estimates of heterogeneous treatment effects from eight randomized controlled trials. When physicians depart from guidelines, they are not responding to measurable treatment effect heterogeneity. Improving adherence to the current guideline produces much larger gains than making all physicians aware of the guideline. Our estimates imply that adherence to an optimal rule would prevent 47% more strokes without increasing hemorrhages. Institutional subscribers to the NBER working paper series, and residents of developing countries may download this paper without additional charge at www.nber.org.

Background Mitochondrial fission counterbalances fusion to maintain organelle morphology, but its role during development remains poorly characterized. Mammalian spermatogenesis is a complex developmental process involving several drastic changes to mitochondrial shape and organization. Mitochondria are generally small and spherical in spermatogonia, elongate during meiosis, and fragment in haploid round spermatids. Near the end of spermatid maturation, small mitochondrial spheres line the axoneme, elongate, and tightly wrap around the midpiece to form the mitochondrial sheath, which is critical for fueling flagellar movements. It remains unclear how these changes in mitochondrial morphology are regulated and how they affect sperm development. Methods We used genetic ablation of Mff (mitochondrial fission factor) in mice to investigate the role of mitochondrial fission during mammalian spermatogenesis. Results Our analysis indicates that Mff is required for mitochondrial fragmentation in haploid round spermatids and for organizing mitochondria in the midpiece in elongating spermatids. In Mff mutant mice, round spermatids have aberrantly elongated mitochondria that often show central constrictions, suggestive of failed fission events. In elongating spermatids and spermatozoa, mitochondrial sheaths are disjointed, containing swollen mitochondria with large gaps between organelles. These mitochondrial abnormalities in Mff mutant sperm are associated with reduced respiratory chain Complex IV activity, aberrant sperm morphology and motility, and reduced fertility. Conclusions Mff is required for organization of the mitochondrial sheath in mouse sperm. General Significance Mitochondrial fission plays an important role in regulating mitochondrial organization during a complex developmental process.

Importance Many employers use workplace wellness programs to improve employee health and reduce medical costs, but randomized evaluations of their efficacy are rare. Objective To evaluate the effect of a comprehensive workplace wellness program on employee health, health beliefs, and medical use after 12 and 24 months. Design, Setting, and Participants This randomized clinical trial of 4834 employees of the University of Illinois at Urbana-Champaign was conducted from August 9, 2016, to April 26, 2018. Members of the treatment group (n = 3300) received incentives to participate in the workplace wellness program. Members of the control group (n = 1534) did not participate in the wellness program. Statistical analysis was performed on April 9, 2020. Interventions The 2-year workplace wellness program included financial incentives and paid time off for annual on-site biometric screenings, annual health risk assessments, and ongoing wellness activities (eg, physical activity, smoking cessation, and disease management). Main Outcomes and Measures Measures taken at 12 and 24 months included clinician-collected biometrics (16 outcomes), administrative claims related to medical diagnoses (diabetes, hypertension, and hyperlipidemia) and medical use (office visits, inpatient visits, and emergency department visits), and self-reported health behaviors and health beliefs (14 outcomes). Results Among the 4834 participants (2770 women; mean [SD] age, 43.9 [11.3] years), no significant effects of the program on biometrics, medical diagnoses, or medical use were seen after 12 or 24 months. A significantly higher proportion of employees in the treatment group than in the control group reported having a primary care physician after 24 months (1106 of 1200 [92.2%] vs 477 of 554 [86.1%]; adjustedP = .002). The intervention significantly improved a set of employee health beliefs on average: participant beliefs about their chance of having a body mass index greater than 30, high cholesterol, high blood pressure, and impaired glucose level jointly decreased by 0.07 SDs (95% CI, −0.12 to −0.01 SDs;P = .02); however, effects on individual belief measures were not significant. Conclusions and Relevance This randomized clinical trial showed that a comprehensive workplace wellness program had no significant effects on measured physical health outcomes, rates of medical diagnoses, or the use of health care services after 24 months, but it increased the proportion of employees reporting that they have a primary care physician and improved employee beliefs about their own health. Trial Registration American Economic Association Randomized Controlled Trial Registry number:AEARCTR-0001368

Mitochondria are highly dynamic organelles that continuously grow, divide, and fuse. The division of mitochondria is crucial for human health. During mitochondrial division, the mechano-guanosine triphosphatase (GTPase) dynamin-related protein (Drp1) severs mitochondria at endoplasmic reticulum (ER)-mitochondria contact sites, where peripheral ER tubules interact with mitochondria. Here, we report that Drp1 directly shapes peripheral ER tubules in human and mouse cells. This ER-shaping activity is independent of GTP hydrolysis and located in a highly conserved peptide of 18 amino acids (termed D-octadecapeptide), which is predicted to form an amphipathic α helix. Synthetic D-octadecapeptide tubulates liposomes in vitro and the ER in cells. ER tubules formed by Drp1 promote mitochondrial division by facilitating ER-mitochondria interactions. Thus, Drp1 functions as a two-in-one protein during mitochondrial division, with ER tubulation and mechano-GTPase activities.

After birth, cardiomyocytes (CM) acquire numerous adaptations in order to efficiently pump blood throughout an animal’s lifespan. How this maturation process is regulated and coordinated is poorly understood. Here, we perform a CRISPR/Cas9 screen in mice and identify serum response factor (SRF) as a key regulator of CM maturation. Mosaic SRF depletion in neonatal CMs disrupts many aspects of their maturation, including sarcomere expansion, mitochondrial biogenesis, transverse-tubule formation, and cellular hypertrophy. Maintenance of maturity in adult CMs is less dependent on SRF. This stage-specific activity is associated with developmentally regulated SRF chromatin occupancy and transcriptional regulation. SRF directly activates genes that regulate sarcomere assembly and mitochondrial dynamics. Perturbation of sarcomere assembly but not mitochondrial dynamics recapitulates SRF knockout phenotypes. SRF overexpression also perturbs CM maturation. Together, these data indicate that carefully balanced SRF activity is essential to promote CM maturation through a hierarchy of cellular processes orchestrated by sarcomere assembly.

Mitochondrial dysfunction is associated with a number of human diseases. As an example, we recently established in vivo Drosophila models of IBMPFD (Inclusion body myopathy, Paget disease, and frontotemporal dementia), and uncovered that human disease mutations of the p97/VCP (Valosin Containing Protein) gene behave as hyperactive alleles associated with mitochondrial defects. Pharmacologic inhibition of VCP strongly suppressed disease and mitochondrial pathology in these animal models. In this protocol, we describe a method to evaluate mitochondrial respiratory function in IBMPFD patient-derived fibroblasts, as well as investigate the role of pharmacologic treatments. These experiments complement work done in animal models by investigating mitochondrial biology and the pharmacologic response in a human cell-based model of the disease. In principle, this technique can be used to investigate mitochondrial respiratory function for any disease in which patient-derived fibroblasts are available.

Background: The dynamin 1-like gene ( DNM1L) encodes a GTPase that mediates mitochondrial and peroxisomal fission and fusion. We report a new clinical presentation associated with a DNM1L pathogenic variant and review the literature. Results: A 13-year-old boy with mild developmental delays and paroxysmal dystonia presented acutely with multifocal myoclonic super-refractory status epilepticus. Despite sustained and aggressive treatment, seizures persisted and care was ultimately withdrawn in the context of extensive global cortical atrophy. Rapid trio-whole exome sequencing revealed a de novo heterozygous c.1207C>T (p.R403C) pathogenic variant in DNM1L. Immunofluorescence staining of fibroblast mitochondria revealed abnormally elongated and tubular morphology. Conclusions: This case highlights the diagnostic importance of rapid whole exome sequencing within a critical care setting and reveals the expanding phenotypic spectrum associated with DNM1L variants. This now includes progressive paroxysmal dystonia and adolescent-onset super-refractory myoclonic status epilepticus contributing to strikingly rapid and progressive cortical atrophy and death.

Summary PINK1 and Parkin mediate mitophagy, the cellular process that clears dysfunctional mitochondria. Mitophagy is regulated by mitochondrial dynamics, but the molecules linking these two processes remain poorly understood. Here, we show that Sam50, the core component of the sorting and assembly machinery (SAM), is a critical regulator of mitochondrial dynamics and PINK1-Parkin-mediated mitophagy. In response to Sam50 depletion, normal tubular mitochondria are first fragmented and subsequently merged into large spheres. Sam50 interacts with PINK1 to facilitate its processing and degradation. Depletion of Sam50 results in PINK1 accumulation, Parkin recruitment, and mitophagy. Interestingly, Sam50 deficiency induces a piecemeal mode of mitophagy that eliminates mitochondria “bit by bit” but spares mtDNA. In C. elegans, the Sam50 homolog gop-3 is required for the maintenance of mitochondrial morphology and mass. Our findings reveal that Sam50 directly links mitochondrial dynamics and mitophagy and that Sam50 depletion induces elimination of mitochondria without affecting mtDNA content.

Summary: Mitochondria shape cytosolic calcium ([Ca2+]c) transients and utilize the mitochondrial Ca2+ ([Ca2+]m) in exchange for bioenergetics output. Conversely, dysregulated [Ca2+]c causes [Ca2+]m overload and induces permeability transition pore and cell death. Ablation of MCU-mediated Ca2+ uptake exhibited elevated [Ca2+]c and failed to prevent stress-induced cell death. The mechanisms for these effects remain elusive. Here, we report that mitochondria undergo a cytosolic Ca2+-induced shape change that is distinct from mitochondrial fission and swelling. [Ca2+]c elevation, but not MCU-mediated Ca2+ uptake, appears to be essential for the process we term mitochondrial shape transition (MiST). MiST is mediated by the mitochondrial protein Miro1 through its EF-hand domain 1 in multiple cell types. Moreover, Ca2+-dependent disruption of Miro1/KIF5B/tubulin complex is determined by Miro1 EF1 domain. Functionally, Miro1-dependent MiST is essential for autophagy/mitophagy that is attenuated in Miro1 EF1 mutants. Thus, Miro1 is a cytosolic Ca2+ sensor that decodes metazoan Ca2+ signals as MiST. : Metazoan Ca2+ signal determines mitochondrial shape transition (MiST) and cellular quality control. Nemani et al. find that mitochondria undergo shape changes upon Ca2+ stress. MiST is distinct from matrix Ca2+-induced swelling and mitochondrial dynamics. The conserved Ca2+ sensor Miro1 enables MiST and promotes autophagy/mitophagy. Keywords: mitochondrial shape, MiST, calcium, Miro, EF hand, PTP, MCU, mitophagy, autophagy, mitochondrial dynamics

Work schedules play an important role in utilizing labor in organizations. In this study of emergency department physicians in shift work, schedules induce two distortions: First, physicians “slack off†by accepting fewer patients near end of shift (EOS). Second, physicians distort patient care, incurring higher costs as they spend less time on patients assigned near EOS. Examining how these effects change with shift overlap reveals a tradeoff between the two. Within an hour after the normal time of work completion, physicians are willing to spend hospital resources more than six times their market wage to preserve their leisure. Accounting for overall costs, I find that physicians slack off at approximately second†best optimal levels.

Mitochondria are double-membrane organelles with varying shapes influenced by metabolic conditions, developmental stage, and environmental stimuli1–4. Their dynamic morphology is realized through regulated and balanced fusion and fission processes5, 6. Fusion is crucial for the health and physiological functions of mitochondria, including complementation of damaged mitochondrial DNAs and maintenance of membrane potential6–8. Mitofusins (Mfns) are dynamin-related GTPases essential for mitochondrial fusion9, 10. They are embedded in the mitochondrial outer membrane and thought to fuse adjacent mitochondria via concerted oligomerization and GTP hydrolysis11–13. However, the molecular mechanisms behind this process remains elusive. Here we present crystal structures of engineered human Mfn1 containing the GTPase domain and a helical domain in different stages of GTP hydrolysis. The helical domain is composed of elements from widely dispersed sequence regions of Mfn1 and resembles the Neck of the bacterial dynamin-like protein. The structures reveal unique features of its catalytic machinery and explain how GTP binding induces conformational changes to promote G domain dimerization in the transition state. Disruption of G domain dimerization abolishes the fusogenic activity of Mfn1. Moreover, a conserved aspartate trigger was found in Mfn1 to affect mitochondrial elongation, likely through a GTP-loading-dependent domain rearrangement. Based on these results, we propose a mechanistic model for Mfn1-mediated mitochondrial tethering. Our study provides important insights in the molecular basis of mitochondrial fusion and mitofusin-related human neuromuscular disorders14.

Differentiating cells tailor their metabolism to fulfill their specialized functions. We examined whether mitochondrial fusion is important for metabolic tailoring during spermatogenesis. Acutely after depletion of mitofusins Mfn1 and Mfn2, spermatogenesis arrests due to failure to accomplish a metabolic shift during meiosis. This metabolic shift includes increased mitochondrial content, mitochondrial elongation, and upregulation of oxidative phosphorylation (OXPHOS). With long-term mitofusin loss, all differentiating germ cell types are depleted, but proliferation of stem-like undifferentiated spermatogonia remains unaffected. Thus, compared with undifferentiated spermatogonia, differentiating spermatogonia and meiotic spermatocytes have cell physiologies that require high levels of mitochondrial fusion. Proteomics in fibroblasts reveals that mitofusin-null cells downregulate respiratory chain complexes and mitochondrial ribosomal subunits. Similarly, mitofusin depletion in immortalized spermatocytes or germ cells in vivo results in reduced OXPHOS subunits and activity. We reveal that by promoting OXPHOS, mitofusins enable spermatogonial differentiation and a metabolic shift during meiosis.

BACKGROUND: The Relative Value Scale Update Committee (RUC) of the American Medical Association plays a central role in determining physician reimbursement. The RUC’s role and performance have been criticized but subjected to little empirical evaluation. METHODS: We analyzed the accuracy of valuations of 293 common surgical procedures from 2005 through 2015. We compared the RUC’s estimates of procedure time with “benchmark” times for the same procedures derived from the clinical registry maintained by the American College of Surgeons National Surgical Quality Improvement Program (NSQIP). We characterized inaccuracies, quantified their effect on physician revenue, and examined whether re-review corrected them. RESULTS: At the time of 108 RUC reviews, the mean absolute discrepancy between RUC time estimates and benchmark times was 18.5 minutes, or 19.8% of the RUC time. However, RUC time estimates were neither systematically shorter nor longer than benchmark times overall (β, 0.97; 95% confidence interval, 0.94 to 1.01; P = 0.10). Our analyses suggest that whereas orthopedic surgeons and urologists received higher payments than they would have if benchmark times had been used ($160 million and $40 million more, respectively, in Medicare reimbursement in 2011 through 2015), cardiothoracic surgeons, neurosurgeons, and vascular surgeons received lower payments ($130 million, $60 million, and $30 million less, respectively). The accuracy of RUC time estimates improved in 47% of RUC revaluations, worsened in 27%, and was unchanged in 25%. (Percentages do not sum to 100 because of rounding.) CONCLUSIONS: In this analysis of frequently conducted operations, we found substantial absolute discrepancies between intraoperative times as estimated by the RUC and the times recorded for the same procedures in a surgical registry, but the RUC did not systematically overestimate or underestimate times. (Funded by the National Institutes of Health.)

I investigate how teamwork may reduce moral hazard by joint monitoring and management. I study two organizational systems differing in the extent to which physicians may mutually manage work: Physicians are assigned patients in a “nurse-managed” system but divide patients between themselves in a “self-managed” system. The self-managed system increases throughput productivity by reducing a “foot-dragging” moral hazard, in which physicians prolong patient stays as expected future work increases. I find evidence that physicians in the same location have better information about each other and that, in the self-managed system, they use this information to assign patients.

DNM1L encodes dynamin-related protein 1 (DRP1/DLP1), a key component of the mitochondrial fission machinery that is essential for proper functioning of the mammalian brain. Previously reported probands with de novo missense mutations in DNM1L presented in the first year of life with severe encephalopathy and refractory epilepsy, with several dying within the first several weeks after birth. In contrast, we report identical novel missense mutations in DNM1L in two unrelated probands who experienced normal development for several years before presenting with refractory focal status epilepticus and subsequent rapid neurological decline. We expand the phenotype of DNM1L-related mitochondrial fission defects, reveal common unique clinical characteristics and imaging findings, and compare the cellular impact of this novel mutation to the previously reported A395D lethal variant. We demonstrate that our R403C mutation, which resides in the assembly region of DRP1, acts by a dominant-negative mechanism and reduces oligomerization, mitochondrial fission activity, and mitochondrial recruitment of DRP1, but to a lesser extent compared to the A395D mutation. In contrast to the initial report of neonatal lethality resulting from DNM1L mutation and DRP1 dysfunction, our results show that milder DRP1 impairment is compatible with normal early development and subsequently results in a distinct set of neurological findings. In addition, we identify a common pathogenic mechanism whereby DNM1L mutations impair mitochondrial fission. © 2016 Wiley Periodicals, Inc.

Background: Liquid biopsies have potential clinical utility as dynamic biomarkers for treatment response in advanced breast cancer. We evaluated a plasma-only assay to track serial changes in WG cfDNA to identify disease progression prior to routine imaging. Methods: We prospectively enrolled and serially collected blood from 25 patients with advanced breast cancer. Blood was drawn and collected in Streck tubes prior to start of a new treatment, after the first cycle (median 22 days), and/or second cycle (median 52 days). 4 mL of plasma was separated from peripheral blood, after which cfDNA was isolated from plasma and used to prepare libraries (11 with bisulfite conversion) for WG sequencing (median 20x depth). Based on a patient-specific profile of WG features, including copy-number alterations and cfDNA fragment length, the fraction of tumor-derived cfDNA (ctDNA) was quantified over the initial course of treatment. For a subset of the cohort (11 of 25 patients), we also quantified changes in genome-wide methylation levels from baseline to subsequent timepoints to classify patients as progressors or non-progressors. Imaging was performed per standard practice with treatment response determined by an independent radiologist according to RECIST 1.1 guidelines. Results: Median age of patients in the cohort was 65 (range 30-83) and included all subtypes—HR+ (n=14), HR+ & HER2+ (n=6), and triple negative (n=5). 52% of patients were on their third line of therapy (range 1-5). On-study therapies were chemotherapy alone (8), targeted + hormone therapy (5), hormone therapy alone (4), targeted + chemotherapy (5), targeted therapy alone (1), or immune checkpoint + HDAC inhibitors (2). Patients with predicted progression by cfDNA (n=5), indicated by an increase in tumor fraction at either post-treatment blood collection, had worse progression-free survival (median 67 days) compared to patients who did not show an increase (n=20; median 207 days) (hazard ratio 7.9, [95% CI 2.2-28.5], log-rank p=3 × 10-4). For the patients who were predicted to progress, the ctDNA assay preceded clinical evaluation by a median of 53 days. All patients with predicted progression were later confirmed to progress at the first follow-up evaluation (5/5, 100% positive predictive value). For the remaining patients, 15 of 20 did not progress (75% negative predictive value). Therefore, sensitivity for the assay was 50% and specificity was 100%. Comparing molecular predictions for 11 of 25 patients based on genomic versus methylation features, 8 non-progressors were classified correctly by both types of features. For the 3 progressors, 2 were predicted correctly based solely on methylation, increasing the sensitivity to detect progression early in the treatment course. Conclusions: Analyzing ctDNA early in the course of a new therapy holds promise to identify patients with early disease progression across multiple types of treatment. The assay accurately identified patients with no durable clinical benefit earlier, indicating potential clinical utility to change therapy and to limit unnecessary side effects and costs associated with ineffective treatments, if validated in a prospective clinical trial.​ Finally, integrating methylation-based changes with information about genomic alterations may increase performance of ctDNA-based response monitoring. Citation Format: Andrew A Davis, David Chan, Michael S Oh, Robert W Lentz, Neil Peterman, Alex Robertson, Rohith Srivas, Abhik Shah, Nicole Lambert, Ayse Tezcan, Haluk Tezcan, Young Kwang Chae. Longitudinal changes in whole-genome (WG) cell-free DNA (cfDNA) and methylation as a blood-based biomarker to identify early disease progression in advanced breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P5-01-13.

Objective: Excessive mitochondrial fission has been associated with several neurodegenerative diseases, including Huntington’s disease (HD). Consequently, mitochondrial dynamics has been suggested to be a promising therapeutic target for Huntington’s disease. Mitochondrial fission depends on recruitment of Drp1 to mitochondria, and Mff (mitochondrial fission factor) is one of the key adaptor proteins for this process. Removal of Mff therefore greatly reduces mitochondrial fission. Here we investigate whether removal of Mff can mitigate HD-associated pathologies in HD transgenic mice (R6/2) expressing mutant Htt. Method: We compared the phenotype of HD mice with and without Mff. The mice were monitored for lifespan, neurological phenotypes, Htt aggregate formation, and brain histology. Results: We found that HD mice lacking Mff display more severe neurological phenotypes and have shortened lifespans. Loss of Mff does not affect mutant Htt aggregation, but it accelerates HD pathology, including neuronal loss and neuroinflammation. Conclusions: Our data indicate a protective role for mitochondrial fission in HD and suggest that more studies are needed before manipulation of mitochondrial dynamics can be applied to HD therapy.