SARS-CoV-2 Mpro responds to oxidation by forming disulfide and NOS/SONOS bonds.

The main protease (Mpro) of SARS-CoV-2 is critical for viral function and a key drug target. Mpro is only active when reduced; turnover ceases upon oxidation but is restored by re-reduction. This suggests the system has evolved to survive periods in an oxidative environment, but the mechanism of this protection has not been confirmed. Here, we report a crystal structure of oxidized Mpro showing a disulfide bond between the active site cysteine, C145, and a distal cysteine, C117. Previous work proposed this disulfide provides the mechanism of protection from irreversible oxidation. Mpro forms an obligate homodimer, and the C117-C145 structure shows disruption of interactions bridging the dimer interface, implying a correlation between oxidation and dimerization. We confirm dimer stability is weakened in solution upon oxidation. Finally, we observe the protein's crystallization behavior is linked to its redox state. Oxidized Mpro spontaneously forms a distinct, more loosely packed lattice. Seeding with crystals of this lattice yields a structure with an oxidation pattern incorporating one cysteine-lysine-cysteine (SONOS) and two lysine-cysteine (NOS) bridges. These structures further our understanding of the oxidative regulation of Mpro and the crystallization conditions necessary to study this structurally.

Evaluation of the point sampling method and inter-comparison of remote emission sensing systems for screening real-world car emissions.

Emissions from internal combustion vehicles are currently not properly monitored throughout their life cycle. Remote emission sensing (RES) is a technology that can measure emissions under real driving conditions without contact. Current light extinction based RES systems are capable of providing emission factors for various gases, but lack accuracy for particulate matter (PM). Point Sampling (PS) is an extraction-based RES technique that can measure gases as well as various particle metrics such as black carbon or particle number. In this work, we evaluated the performance of a recently developed PS system and the state-of-the-art light extinction based remote sensing devices EDAR (HEAT) and ORSD (OPUS RSE) during co-location measurements. Validation measurements with portable emission measurement systems and emissions screening of several thousand cars in three European cities provide detailed insights into system's performance. Meteorological evaluations showed that the PS capture rate is strongly influenced by wind, but no other weather influences were found. Both light extinction based systems are unable to measure during rain. We found that all three systems tested were capable of screening NOx emissions from pre-Euro 6 diesel cars. Measurement results show the ability of the PS system to quantify high and low PM emitters equally well. The open-path RES systems (EDAR, ORSD) are capable of estimating PM emissions from pre-Euro 5 diesel cars. However, deficiencies of open-path RES systems are evident in the quantification of PM emissions from newer engine technologies (diesel Euro 5 and beyond) and from petrol cars. The PS system has a 2 to 5 times lower capture rate than open-path RES systems, but the PS measurement results are more accurate (more than 5 times for PM and more than 1.35 times for NOx). The good accuracy of individual measurements makes PS a powerful tool for reliable high emitter identification.

The sleep-circadian interface: A window into mental disorders.

Sleep, circadian rhythms, and mental health are reciprocally interlinked. Disruption to the quality, continuity, and timing of sleep can precipitate or exacerbate psychiatric symptoms in susceptible individuals, while treatments that target sleep-circadian disturbances can alleviate psychopathology. Conversely, psychiatric symptoms can reciprocally exacerbate poor sleep and disrupt clock-controlled processes. Despite progress in elucidating underlying mechanisms, a cohesive approach that integrates the dynamic interactions between psychiatric disorder with both sleep and circadian processes is lacking. This review synthesizes recent evidence for sleep-circadian dysfunction as a transdiagnostic contributor to a range of psychiatric disorders, with an emphasis on biological mechanisms. We highlight observations from adolescent and young adults, who are at greatest risk of developing mental disorders, and for whom early detection and intervention promise the greatest benefit. In particular, we aim to a) integrate sleep and circadian factors implicated in the pathophysiology and treatment of mood, anxiety, and psychosis spectrum disorders, with a transdiagnostic perspective; b) highlight the need to reframe existing knowledge and adopt an integrated approach which recognizes the interaction between sleep and circadian factors; and c) identify important gaps and opportunities for further research.

Engineering cannabinoid production in Saccharomyces cerevisiae.

Phytocannabinoids are natural products with highly interesting pharmacological properties mainly produced by plants. The production of cannabinoids in a heterologous host system has gained interest in recent years as a promising alternative to production from plant material. However, the systems reported so far do not achieve industrially relevant titers, highlighting the need for alternative systems. Here, we show the production of the cannabinoids cannabigerolic acid and cannabigerol from glucose and hexanoic acid in a heterologous yeast system using the aromatic prenyltransferase NphB from Streptomyces sp. strain CL190. The production was significantly increased by introducing a fusion protein consisting of ERG20WW and NphB. Furthermore, we improved the production of the precursor olivetolic acid to a titer of 56 mg L-1 . The implementation of the cannabinoid synthase genes enabled the production of Δ9 -tetrahydrocannabinolic acid, cannabidiolic acid as well as cannabichromenic acid, where the heterologous biosynthesis of cannabichromenic acid in a yeast system was demonstrated for the first time. In addition, we found that the product spectrum of the cannabinoid synthases localized to the vacuoles of the yeast cells was highly dependent on extracellular pH, allowing for easy manipulation. Finally, using a fed-batch approach, we showed cannabigerolic acid and olivetolic acid titers of up to 18.2 mg L-1 and 117 mg L-1 , respectively.

Revealing bactericidal events on graphene oxide nano films deposited on metal implant surfaces.

At the time when pathogens are developing robust resistance to antibiotics, the demand for implant surfaces with microbe-killing capabilities has significantly risen. To achieve this goal, profound understanding of the underlying mechanisms is crucial. Our study demonstrates that graphene oxide (GO) nano films deposited on stainless steel (SS316L) exhibit superior antibacterial features. The physicochemical properties of GO itself play a pivotal role in influencing biological events and their diversity may account for the contradictory results reported elsewhere. However, essential properties of GO coatings, such as oxygen content and the resulting electrical conductivity, have been overlooked so far. We hypothesize that the surface potential and electrical resistance of the oxygen content in the GO-nano films may induce bacteria-killing events on conductive metallic substrates. In our study, the GO applied contains 52 wt% of oxygen, and thus exhibits insulating properties. When deposited as a nano film on an electrically conducting steel substrate, GO flakes generate a Schottky barrier at the interface. This barrier, consequently, impedes the transfer of electrons to the underlying conductive substrate. As a result, this creates reactive oxygen species (ROS), leading to bacterial death. We confirmed the presence of GO coatings and their hydrolytic stability by using X-ray photoelectron spectroscopy (XPS), µRaman spectroscopy, scanning electron microscopy (SEM), and Kelvin probe force microscopy (KPFM) measurements. The biological evaluation was performed on the MG63 osteoblast-like cell line and two selected bacteria species: S. aureus and P. aeruginosa, demonstrating both the cytocompatibility and antibacterial behavior of GO-coated SS316L substrates. We propose a two-step bactericidal mechanism: electron transfer from the bacteria membrane to the substrate, followed by ROS generation. This mechanism finds support in changes observed in contact angle, surface potential, and work function, identified as decisive factors. By addressing overlooked factors and effectively bridging the gap between understanding and practicality, we present a transformative approach for implant surfaces, combating microbial resistance, and offering new application possibilitie.

Light-induced Trpin/Metout Switching During BLUF Domain Activation in ATP-bound Photoactivatable Adenylate Cyclase OaPAC.

The understanding of signal transduction mechanisms in photoreceptor proteins is essential for elucidating how living organisms respond to light as environmental stimuli. In this study, we investigated the ATP binding, photoactivation and signal transduction process in the photoactivatable adenylate cyclase from Oscillatoria acuminata (OaPAC) upon blue light excitation. Structural models with ATP bound in the active site of native OaPAC at cryogenic as well as room temperature are presented. ATP is found in one conformation at cryogenic- and in two conformations at ambient-temperature, and is bound in an energetically unfavorable conformation for the conversion to cAMP. However, FTIR spectroscopic experiments confirm that this conformation is the native binding mode in dark state OaPAC and that transition to a productive conformation for ATP turnover only occurs after light activation. A combination of time-resolved crystallography experiments at synchrotron and X-ray Free Electron Lasers sheds light on the early events around the Flavin Adenine Dinucleotide (FAD) chromophore in the light-sensitive BLUF domain of OaPAC. Early changes involve the highly conserved amino acids Tyr6, Gln48 and Met92. Crucially, the Gln48 side chain performs a 180° rotation during activation, leading to the stabilization of the FAD chromophore. Cryo-trapping experiments allowed us to investigate a late light-activated state of the reaction and revealed significant conformational changes in the BLUF domain around the FAD chromophore. In particular, a Trpin/Metout transition upon illumination is observed for the first time in the BLUF domain and its role in signal transmission via α-helix 3 and 4 in the linker region between sensor and effector domain is discussed.

Foldseek reveals a CBGA prenylating enzyme GlyMa_02G168000 from Glycine max.

The present research provides an application for an aromatic prenyltransferase from Glycine max for use in heterologous microorganism expression to generate cannabinoids. The known cannabinoid prenyltransferase CsPT04 was queried in FoldSeek. An enzyme derived from Glycine max known as GLYMA_02G168000, which is a predicted homogentisate solanyltransferase, was identified and found to have affinity for the prenylation of geranyldiphosphate (GPP) and olivetolic acid (OA) to produce cannabigerolic acid (CBGA) and cannabigerol (CBG). The in vitro production of CBGA was accomplished through the heterologous expression of this prenyltransferase in Saccharomyces cerevisiae. After growing the yeast cells, a purified microsomal fraction was harvested, which was rich in the membrane-bound prenyltransferase GlyMa_02G168000. Addition of purified microsomal fraction to a reaction matrix facilitated the successful prenylation of externally supplied OA with GPP, culminating in the production of CBGA. Structural comparisons revealed a notably closer similarity between GLYMA_02G168000 and CsPT04, compared to the similarity of other cannabinoid prenyltransferases with CsPT04. Herein, a novel application for a homogentisate solanyltransferase has been established towards the production of cannabinoids.

Trends in Attempted Vaginal Delivery among Pregnancies Complicated by Gastroschisis, 2014 to 2020.

OBJECTIVE: Gastroschisis is a full-thickness congenital defect of the abdominal wall through which intestines and other organs may herniate. In a prior analysis, attempted vaginal delivery with fetal gastroschisis appeared to increase through 2013, although cesarean delivery remained common. The objective of this analysis was to update current trends in attempted vaginal birth among pregnancies complicated by gastroschisis. STUDY DESIGN: We performed an updated cross-sectional analysis of live births from 2014 and 2020 using data from the U.S. National Vital Statistics System and evaluated trends in attempted vaginal deliveries among births with gastroschisis. Trends were evaluated using joinpoint regression. We constructed logistic regression models to evaluate the association between demographic and clinical variables and attempted vaginal delivery in the setting of gastroschisis. RESULTS: Among 5,355 deliveries with gastroschisis meeting inclusion criteria, attempted vaginal delivery increased significantly from 68.9% to 75.1%, an average annual percent change of 1.7% (95% confidence interval [CI], 0.8-2.5). Among gastroschisis-complicated pregnancies, patients 35 to 39 years old (adjusted odds ratio [aOR], 0.53; 95% CI, 0.37-0.79) and Hispanic race/ethnicity (aOR, 0.69; 95% CI, 0.58-0.62) were at lower likelihood of attempted vaginal delivery in adjusted analyses. CONCLUSION: These findings suggest that vaginal delivery continues to increase in the setting of gastroschisis. Further reduction of surgical delivery for this fetal defect may be possible. KEY POINTS: · Vaginal deliveries increased among gastroschisis pregnancies.. · Hispanic patients were less likely to attempt vaginal delivery.. · Some gastroschisis pregnancies still deliver surgically..

Ultradian sleep cycles: Frequency, duration, and associations with individual and environmental factors-A retrospective study.

OBJECTIVE: Sleep varies between individuals in response to sleep-wake history and various environmental factors, including light and noise. Here we report on the intranight variation of the ultradian nonrapid eye movement-rapid eye movement (NREM-REM) sleep cycle in 369 participants who have contributed to different laboratory studies from 1994 to 2020 at the Centre for Chronobiology, Basel, Switzerland. RESULTS: We observed a large interindividual variability in sleep cycle duration, including NREM and REM sleep episodes in healthy participants who were given an 8-hour sleep opportunity at habitual bedtime in controlled laboratory settings. The median sleep cycle duration was 96 minutes out of 6064 polysomnographically-recorded cycles. The number and duration of cycles were not normally distributed, and the distribution became narrower for NREM sleep and wider for REM sleep later in the night. The first cycle was consistently shorter than subsequent cycles, and moderate presleep light or nocturnal noise exposure had no significant effects on ultradian sleep cycle duration. Age and sex significantly affected NREM and REM sleep duration, with older individuals having longer NREM and shorter REM sleep particularly in the end of the night, and females having longer NREM sleep episodes. High sleep pressure (ie, sleep deprivation) and low sleep pressure (ie, multiple naps) altered ultradian sleep cycles, with high sleep pressure leading to longer NREM sleep in the first cycle, and low sleep pressure leading to longer REM sleep episodes. Positive correlations were observed between N2 and NREM duration, and between N1 and REM duration. Weak intrasleep REM sleep homeostasis was also evident in our data set. CONCLUSIONS: We conclude that ultradian sleep cycles are endogenous biological rhythms modulated by age, sex, and sleep homeostasis, but not directly responsive to (moderate levels of) environmental cues in healthy good sleepers.

Prediction of cognitive decline in healthy aging based on neuropsychiatric symptoms and PET-biomarkers of Alzheimer's disease.

Neuropsychiatric symptoms (NPS) have been associated with a risk of accelerated cognitive decline or conversion to dementia of the Alzheimer's Disease (AD) type. Moreover, the NPS were also associated with higher AD biomarkers (brain tau and amyloid burden) even in non-demented patients. But the effect of the relationship between NPS and biomarkers on cognitive decline has not yet been studied. This work aims to assess the relationship between longitudinal cognitive changes and NPS, specifically depression and anxiety, in association with AD biomarkers in healthy middle-aged to older participants. The cohort consisted of 101 healthy participants aged 50-70 years, 66 of whom had neuropsychological assessments of memory, executive functions, and global cognition at a 2-year follow-up. At baseline, NPS were assessed using the Beck Depression and Anxiety Inventories while brain tau and amyloid loads were measured using positron emission topography. For tau burden, THK5351 uptake is used as a proxy of tau and neuroinflammation. Participants, declining or remaining stable at follow-up, were categorized into groups for each cognitive domain. Group classification was investigated using binary logistic regressions based on combined AD biomarkers and the two NPS. The results showed that an association between anxiety and prefrontal amyloid burden significantly classified episodic memory decline, while the classification of global cognitive decline involved temporal and occipital amyloid burden but not NPS. Moreover, depression together with prefrontal and hippocampal tau burden were associated with a decline in memory. The classification of participants based on executive decline was related to depression and mainly prefrontal tau burden. These findings suggest that the combination of NPS and brain biomarkers of AD predicts the occurrence of cognitive decline in aging.

Mitochondrial dysfunction abrogates dietary lipid processing in enterocytes.

Digested dietary fats are taken up by enterocytes where they are assembled into pre-chylomicrons in the endoplasmic reticulum followed by transport to the Golgi for maturation and subsequent secretion to the circulation1. The role of mitochondria in dietary lipid processing is unclear. Here we show that mitochondrial dysfunction in enterocytes inhibits chylomicron production and the transport of dietary lipids to peripheral organs. Mice with specific ablation of the mitochondrial aspartyl-tRNA synthetase DARS2 (ref. 2), the respiratory chain subunit SDHA3 or the assembly factor COX10 (ref. 4) in intestinal epithelial cells showed accumulation of large lipid droplets (LDs) in enterocytes of the proximal small intestine and failed to thrive. Feeding a fat-free diet suppressed the build-up of LDs in DARS2-deficient enterocytes, which shows that the accumulating lipids derive mostly from digested fat. Furthermore, metabolic tracing studies revealed an impaired transport of dietary lipids to peripheral organs in mice lacking DARS2 in intestinal epithelial cells. DARS2 deficiency caused a distinct lack of mature chylomicrons concomitant with a progressive dispersal of the Golgi apparatus in proximal enterocytes. This finding suggests that mitochondrial dysfunction results in impaired trafficking of chylomicrons from the endoplasmic reticulum to the Golgi, which in turn leads to storage of dietary lipids in large cytoplasmic LDs. Taken together, these results reveal a role for mitochondria in dietary lipid transport in enterocytes, which might be relevant for understanding the intestinal defects observed in patients with mitochondrial disorders5.

Napping and circadian sleep-wake regulation during healthy aging.

STUDY OBJECTIVES: Daytime napping is frequently reported among the older population and has attracted increasing attention due to its association with multiple health conditions. Here, we tested whether napping in the aged is associated with altered circadian regulation of sleep, sleepiness and vigilance performance. METHODS: Sixty healthy older individuals (mean age: 69y., 39 women) were recruited with respect to their napping habits (30 nappers, 30 non-nappers). All participants underwent an in-lab 40-h multiple nap protocol (10 cycles of 80 mins of sleep opportunity alternating with 160 mins of wakefulness), preceded and followed by a baseline and recovery sleep period. Saliva samples for melatonin assessment, sleepiness and vigilance performance were collected during wakefulness and electrophysiological data were recorded to derive sleep parameters during scheduled sleep opportunities. RESULTS: The circadian amplitude of melatonin secretion was reduced in nappers, compared to non-nappers. Furthermore, nappers were characterized by higher sleep efficiencies and REM sleep proportion during day- compared to night-time naps. The nap group also presented altered modulation in sleepiness and vigilance performance at specific circadian phases. DISCUSSION: Our data indicate that napping is associated with an altered circadian sleep-wake propensity rhythm and thereby contribute to the understanding of the biological correlates underlying napping and/or sleep-wake cycle fragmentation during healthy aging. Altered circadian sleep-wake promotion can lead to a less distinct allocation of sleep into night-time and/or a reduced wakefulness drive during the day, thereby potentially triggering the need to sleep at adverse circadian phase.

In vivo marker of brainstem myelin is associated to quantitative sleep parameters in healthy young men.

The regional integrity of brain subcortical structures has been implicated in sleep-wake regulation, however, their associations with sleep parameters remain largely unexplored. Here, we assessed association between quantitative Magnetic Resonance Imaging (qMRI)-derived marker of the myelin content of the brainstem and the variability in the sleep electrophysiology in a large sample of 18-to-31 years healthy young men (N = 321; ~ 22 years). Separate Generalized Additive Model for Location, Scale and Shape (GAMLSS) revealed that sleep onset latency and slow wave energy were significantly associated with MTsat estimates in the brainstem (pcorrected ≤ 0.03), with overall higher MTsat value associated with values reflecting better sleep quality. The association changed with age, however (MTsat-by-age interaction-pcorrected ≤ 0.03), with higher MTsat value linked to better values in the two sleep metrics in the younger individuals of our sample aged ~ 18 to 20 years. Similar associations were detected across different parts of the brainstem (pcorrected ≤ 0.03), suggesting that the overall maturation and integrity of the brainstem was associated with both sleep metrics. Our results suggest that myelination of the brainstem nuclei essential to regulation of sleep is associated with inter-individual differences in sleep characteristics during early adulthood. They may have implications for sleep disorders or neurological diseases related to myelin.

Adapting the preterm birth phenotyping framework to a low-resource, rural setting and applying it to births from Migori County in western Kenya.

BACKGROUND: Preterm birth is the leading cause of neonatal and under-five mortality worldwide. It is a complex syndrome characterized by numerous etiologic pathways shaped by both maternal and fetal factors. To better understand preterm birth trends, the Global Alliance to Prevent Prematurity and Stillbirth published the preterm birth phenotyping framework in 2012 followed by an application of the model to a global dataset in 2015 by Barros, et al. Our objective was to adapt the preterm birth phenotyping framework to retrospective data from a low-resource, rural setting and then apply the adapted framework to a cohort of women from Migori, Kenya. METHODS: This was a single centre, observational, retrospective chart review of eligible births from November 2015 - March 2017 at Migori County Referral Hospital. Adaptations were made to accommodate limited diagnostic capabilities and data accuracy concerns. Prevalence of the phenotyping conditions were calculated as well as odds of adverse outcomes. RESULTS: Three hundred eighty-seven eligible births were included in our study. The largest phenotype group was none (no phenotype could be identified; 41.1%), followed by extrauterine infection (25.1%), and antepartum stillbirth (16.7%). Extrauterine infections included HIV (75.3%), urinary tract infections (24.7%), malaria (4.1%), syphilis (3.1%), and general infection (3.1%). Severe maternal condition was ranked fourth (15.6%) and included anaemia (69.5%), chronic respiratory distress (22.0%), chronic hypertension prior to pregnancy (5.1%), diabetes (3.4%), epilepsy (3.4%), and sickle cell disease (1.7%). Fetal anaemia cases were the most likely to transfer to the newborn unit (OR 5.1, 95% CI 0.8, 30.9) and fetal anomaly cases were the most likely to result in a pre-discharge mortality (OR 3.9, 95% CI 0.8, 19.2). CONCLUSIONS: Using routine data sources allowed for a retrospective analysis of an existing dataset, requiring less time and fewer resources than a prospective study and demonstrating a feasible approach to preterm phenotyping for use in low-resource settings to inform local prevention strategies.

Preterm birth is a complex syndrome, yet it is the leading cause of death in children worldwide. To help unravel the clinical complexities, preterm birth phenotyping is a framework that considers multiple diagnoses in the mother, helping to evaluate trends in causes of preterm birth in a given region. In our study, we adapted this international phenotyping framework to accommodate a rural, low-resource setting where obstetrical and neonatal technologies were limited, but preterm birth rates were high. We evaluated data from the patient records of a large hospital in Migori, Kenya, in the southwestern region of the country. By lowering the threshold of diagnostic criteria, we were able to apply this framework to our dataset and see that maternal infection and maternal chronic illness appear to be a significant driving forces of preterm birth. Given high rates of HIV and malaria in the region, this is not a surprising finding, but one that can inform antenatal care practices, mainly the need to test and treat for common infections (HIV, malaria, as well as urinary and reproductive tract infections), and to increase the frequency of antenatal care interactions per the World Health Organization recommendations.

Pediatric pain knowledge and attitudes among health care professionals-A National Danish Survey.

Aim: To explore and compare Danish health-care professionals' attitudes and knowledge towards pain management of children. Methods: The cross-sectional study was carried out using the Pediatric Pain Knowledge and Attitudes Questionnaire. The questionnaire was distributed to all health care professionals caring for children in three hospital settings, including nurses and physicians in departments of pediatric, emergency, and anesthesia and medical laboratory technologists. Results: The study was conducted in 2020 and 765 health care professionals participated. Within the six main categories included in the questionnaire, there were significant differences between nurses and physicians in three subcategories: view on the care of children in pain, using drugs to relieve pain, and the four mandatories. Comparing nurses with medical laboratory technicians, there were significant differences in the subcategory "view on the care of children in pain." Comparing types of clinical departments, there were significant differences in the subcategories' view on the care of children in pain, using drugs to relieve pain, and the four mandatories. Overall, we found that the participating health professionals did not have a uniform understanding of pain management and therefore might treat children differently. Conclusion: The present study highlights the need to align health care professionals' knowledge regarding pain assessment and management of children, as well as the need to develop and test interventions that support the use of knowledge in practice.

Mix-and-extrude: high-viscosity sample injection towards time-resolved protein crystallography.

Time-resolved crystallography enables the visualization of protein molecular motion during a reaction. Although light is often used to initiate reactions in time-resolved crystallography, only a small number of proteins can be activated by light. However, many biological reactions can be triggered by the interaction between proteins and ligands. The sample delivery method presented here uses a mix-and-extrude approach based on 3D-printed microchannels in conjunction with a micronozzle. The diffusive mixing enables the study of the dynamics of samples in viscous media. The device design allows mixing of the ligands and protein crystals in 2 to 20 s. The device characterization using a model system (fluorescence quenching of iq-mEmerald proteins by copper ions) demonstrated that ligand and protein crystals, each within lipidic cubic phase, can be mixed efficiently. The potential of this approach for time-resolved membrane protein crystallography to support the development of new drugs is discussed.

The ribosomal protein S6 kinase alpha-1 (RPS6KA1) induces resistance to venetoclax/azacitidine in acute myeloid leukemia.

Venetoclax/azacitidine combination therapy is effective in acute myeloid leukemia (AML) and tolerable for older, multimorbid patients. Despite promising response rates, many patients do not achieve sustained remission or are upfront refractory. Identification of resistance mechanisms and additional therapeutic targets represent unmet clinical needs. By using a genome-wide CRISPR/Cas9 library screen targeting 18,053 protein- coding genes in a human AML cell line, various genes conferring resistance to combined venetoclax/azacitidine treatment were identified. The ribosomal protein S6 kinase A1 (RPS6KA1) was among the most significantly depleted sgRNA-genes in venetoclax/azacitidine- treated AML cells. Addition of the RPS6KA1 inhibitor BI-D1870 to venetoclax/azacitidine decreased proliferation and colony forming potential compared to venetoclax/azacitidine alone. Furthermore, BI-D1870 was able to completely restore the sensitivity of OCI-AML2 cells with acquired resistance to venetoclax/azacitidine. Analysis of cell surface markers revealed that RPS6KA1 inhibition efficiently targeted monocytic blast subclones as a potential source of relapse upon venetoclax/azacitidine treatment. Taken together, our results suggest RPS6KA1 as mediator of resistance towards venetoclax/azacitidine and additional RPS6KA1 inhibition as strategy to prevent or overcome resistance.

FOXA2 controls the anti-oxidant response in FH-deficient cells.

Hereditary leiomyomatosis and renal cell cancer (HLRCC) is a cancer syndrome caused by inactivating germline mutations in fumarate hydratase (FH) and subsequent accumulation of fumarate. Fumarate accumulation leads to profound epigenetic changes and the activation of an anti-oxidant response via nuclear translocation of the transcription factor NRF2. The extent to which chromatin remodeling shapes this anti-oxidant response is currently unknown. Here, we explored the effects of FH loss on the chromatin landscape to identify transcription factor networks involved in the remodeled chromatin landscape of FH-deficient cells. We identify FOXA2 as a key transcription factor that regulates anti-oxidant response genes and subsequent metabolic rewiring cooperating without direct interaction with the anti-oxidant regulator NRF2. The identification of FOXA2 as an anti-oxidant regulator provides additional insights into the molecular mechanisms behind cell responses to fumarate accumulation and potentially provides further avenues for therapeutic intervention for HLRCC.