A programmable reaction-diffusion system for spatiotemporal cell signaling circuit design.

Cells self-organize molecules in space and time to generate complex behaviors, but we lack synthetic strategies for engineering spatiotemporal signaling. We present a programmable reaction-diffusion platform for designing protein oscillations, patterns, and circuits in mammalian cells using two bacterial proteins, MinD and MinE (MinDE). MinDE circuits act like "single-cell radios," emitting frequency-barcoded fluorescence signals that can be spectrally isolated and analyzed using digital signal processing tools. We define how to genetically program these signals and connect their spatiotemporal dynamics to cell biology using engineerable protein-protein interactions. This enabled us to construct sensitive reporter circuits that broadcast endogenous cell signaling dynamics on a frequency-barcoded imaging channel and to build control signal circuits that synthetically pattern activities in the cell, such as protein condensate assembly and actin filamentation. Our work establishes a paradigm for visualizing, probing, and engineering cellular activities at length and timescales critical for biological function.

WMSA 2: a multiple DNA/RNA sequence alignment tool implemented with accurate progressive mode and a fast win-win mode combining the center star and progressive strategies.

Multiple sequence alignment is widely used for sequence analysis, such as identifying important sites and phylogenetic analysis. Traditional methods, such as progressive alignment, are time-consuming. To address this issue, we introduce StarTree, a novel method to fast construct a guide tree by combining sequence clustering and hierarchical clustering. Furthermore, we develop a new heuristic similar region detection algorithm using the FM-index and apply the k-banded dynamic program to the profile alignment. We also introduce a win-win alignment algorithm that applies the central star strategy within the clusters to fast the alignment process, then uses the progressive strategy to align the central-aligned profiles, guaranteeing the final alignment's accuracy. We present WMSA 2 based on these improvements and compare the speed and accuracy with other popular methods. The results show that the guide tree made by the StarTree clustering method can lead to better accuracy than that of PartTree while consuming less time and memory than that of UPGMA and mBed methods on datasets with thousands of sequences. During the alignment of simulated data sets, WMSA 2 can consume less time and memory while ranking at the top of Q and TC scores. The WMSA 2 is still better at the time, and memory efficiency on the real datasets and ranks at the top on the average sum of pairs score. For the alignment of 1 million SARS-CoV-2 genomes, the win-win mode of WMSA 2 significantly decreased the consumption time than the former version. The source code and data are available at https://github.com/malabz/WMSA2.

Identification and functional analysis of circulating small extracellular vesicle lncRNA signatures in children with fulminant myocarditis.

Fulminant myocarditis (FM) is the most serious type of myocarditis. However, the molecular mechanism underlying the pathogenesis of FM has not been fully elucidated. Small extracellular vesicles (sEVs) play important roles in many diseases, but any potential role in paediatric FM has not been reported. Here, the differential signatures of lncRNAs in plasma sEVs were studied in FM children and healthy children using transcriptome sequencing followed by functional analysis. Then immune-related lncRNAs were screened to study their role in immune mechanisms, the levels and clinical relevance of core immune-related lncRNAs were verified by qRT-PCR in a large sample size. Sixty-eight lncRNAs had increased levels of plasma sEVs in children with FM and 11 had decreased levels. Functional analysis showed that the sEVs-lncRNAs with different levels were mainly related to immunity, apoptosis and protein efflux. Seventeen core immune-related sEVs-lncRNAs were screened, functional enrichment analysis showed that these lncRNAs were closely related to immune activation, immune cell migration and cytokine pathway signal transduction. The results of the study show that sEVs-lncRNAs may play an important role in the pathogenesis of fulminant myocarditis in children, especially in the mechanism of immune regulation.

A novel fast multiple nucleotide sequence alignment method based on FM-index.

Multiple sequence alignment (MSA) is fundamental to many biological applications. But most classical MSA algorithms are difficult to handle large-scale multiple sequences, especially long sequences. Therefore, some recent aligners adopt an efficient divide-and-conquer strategy to divide long sequences into several short sub-sequences. Selecting the common segments (i.e. anchors) for division of sequences is very critical as it directly affects the accuracy and time cost. So, we proposed a novel algorithm, FMAlign, to improve the performance of multiple nucleotide sequence alignment. We use FM-index to extract long common segments at a low cost rather than using a space-consuming hash table. Moreover, after finding the longer optimal common segments, the sequences are divided by the longer common segments. FMAlign has been tested on virus and bacteria genome and human mitochondrial genome datasets, and compared with existing MSA methods such as MAFFT, HAlign and FAME. The experiments show that our method outperforms the existing methods in terms of running time, and has a high accuracy on long sequence sets. All the results demonstrate that our method is applicable to the large-scale nucleotide sequences in terms of sequence length and sequence number. The source code and related data are accessible in https://github.com/iliuh/FMAlign.

Trends for opioid prescribing and the impact of the COVID-19 pandemic in patients with rheumatic and musculoskeletal diseases between 2006 and 2021.

OBJECTIVE: To investigate opioid prescribing trends and assess the impact of the COVID-19 pandemic on opioid prescribing in rheumatic and musculoskeletal diseases (RMDs). METHODS: Adult patients with RA, PsA, axial spondyloarthritis (AxSpA), SLE, OA and FM with opioid prescriptions between 1 January 2006 and 31 August 2021 without cancer in UK primary care were included. Age- and gender-standardized yearly rates of new and prevalent opioid users were calculated between 2006 and 2021. For prevalent users, monthly measures of mean morphine milligram equivalents (MME)/day were calculated between 2006 and 2021. To assess the impact of the pandemic, we fitted regression models to the monthly number of prevalent opioid users between January 2015 and August 2021. The time coefficient reflects the trend pre-pandemic and the interaction term coefficient represents the change in the trend during the pandemic. RESULTS: The study included 1 313 519 RMD patients. New opioid users for RA, PsA and FM increased from 2.6, 1.0 and 3.4/10 000 persons in 2006 to 4.5, 1.8 and 8.7, respectively, in 2018 or 2019. This was followed by a fall to 2.4, 1.2 and 5.9, respectively, in 2021. Prevalent opioid users for all RMDs increased from 2006 but plateaued or dropped beyond 2018, with a 4.5-fold increase in FM between 2006 and 2021. In this period, MME/day increased for all RMDs, with the highest for FM (≥35). During COVID-19 lockdowns, RA, PsA and FM showed significant changes in the trend of prevalent opioid users. The trend for FM increased pre-pandemic and started decreasing during the pandemic. CONCLUSION: The plateauing or decreasing trend of opioid users for RMDs after 2018 may reflect the efforts to tackle rising opioid prescribing in the UK. The pandemic led to fewer people on opioids for most RMDs, providing reassurance that there was no sudden increase in opioid prescribing during the pandemic.

A convenient fluorimetry-based degranulation assay using RBL-2H3 cells.

Type I hypersensitivity is triggered by mast cell degranulation, a stimulus-induced exocytosis of preformed secretory granules (SGs) containing various inflammatory mediators. The degree of degranulation is generally expressed as a percentage of secretory granule markers (such as ß-hexosaminidase and histamine) released into the external solution, and considerable time and labor are required for the quantification of markers in both the supernatants and cell lysates. In this study, we developed a simple fluorimetry-based degranulation assay using rat basophilic leukemia (RBL-2H3) mast cells. During degranulation, the styryl dye FM1-43 in the external solution fluorescently labeled the newly exocytosed SGs, whose increase in intensity was successively measured using a fluorescence microplate reader. In addition to the rate of ß-hexosaminidase secretion, the cellular FM1-43 intensity successfully represented the degree and kinetics of degranulation under various conditions, suggesting that this method facilitates multi-sample and/or multi-time-point analyses required for screening substances regulating mast cell degranulation.

The effect of eye movement desensitization and reprocessing on fibromyalgia: A multiple-baseline experimental case study across ten participants.

Fibromyalgia (FM) is a chronic pain disorder characterized by widespread musculoskeletal pain, fatigue, and stiffness in muscles and joints. Traumatic life experiences and post-traumatic stress symptoms play a role in its development and persistence. Although previous research suggests that pain and FM symptoms decrease after eye movement desensitization and reprocessing (EMDR) therapy, its effectiveness has not been investigated in a controlled manner. The present study investigated the effectiveness of a six-session, 90-minute EMDR therapy using a multiple baseline single-case experimental design (SCED) across ten adult females with FM. The SCED involved a baseline, intervention, one- and three-month follow-up phase. The primary outcome was pain. Secondary outcomes included post-traumatic stress symptoms, other FM symptoms (fatigue, stiffness in muscles and joints), and the impact of FM on daily activities and sleep. Data were statistically analyzed by primarily testing means across phases on an individual and group level. Post-traumatic stress symptoms improved significantly in seven participants. Pain severity decreased significantly in six participants, with three of them maintaining significant improvement three months later. One participant showed clinically relevant change one month later. Furthermore, improvements were observed in secondary outcome measures. The findings support the efficacy of EMDR in reducing FM symptoms.

Exploring levels of TSH and FT4 in patients with chronic fatigue syndrome (CFS), fibromyalgia (FM) and healthy controls did not reveal any associations between fatigue score and level of thyroid hormones.

AIM: The diagnoses of Chronic Fatigue Syndrome (CFS) and Fibromyalgia (FM) are highly associated with fatigue and pain, respectively. Physiologically and clinically an effect of thyroid status on fatigue and pain is expected. There may be clinically relevant differences in thyroid hormone axes though within values of reference in both patients with normal thyroid hormones, or in patients with well-regulated thyroid disease. These potential differences are explored in this study. MATERIALS AND METHODS: In the present study, female patients with CFS (n = 49) and FM (n = 58) as well as female healthy controls (n = 53) were included. We explored plasma levels of TSH and FT4 between the groups using Kruskall-Wallis, and the relation between fatigue score and levels of TSH and FT4 by means of Spearman's rho. RESULTS: There were no group differences between CFS patients, FM patients, and healthy controls in levels of TSH and FT4. CONCLUSION: As one might clinically and physiologically expect an association between thyroid function and fatigue, which may be associated with clinical disorders such as CFS and FM, we suggest future studies to examine the field further by exploring the influence of thyroid receptors and responses of the thyroid hormone cascade.

Singlet oxygen production by photosystem II is caused by misses of the oxygen evolving complex.

Singlet oxygen (1 O2 ) is a harmful species that functions also as a signaling molecule. In chloroplasts, 1 O2 is produced via charge recombination reactions in photosystem II, but which recombination pathway(s) produce triplet Chl and 1 O2 remains open. Furthermore, the role of 1 O2 in photoinhibition is not clear. We compared temperature dependences of 1 O2 production, photoinhibition, and recombination pathways. 1 O2 production by pumpkin thylakoids increased from -2 to +35°C, ruling out recombination of the primary charge pair as a main contributor. S2 QA - or S2 QB - recombination pathways, in turn, had too steep temperature dependences. Instead, the temperature dependence of 1 O2 production matched that of misses (failures of the oxygen (O2 ) evolving complex to advance an S-state). Photoinhibition in vitro and in vivo (also in Synechocystis), and in the presence or absence of O2 , had the same temperature dependence, but ultraviolet (UV)-radiation-caused photoinhibition showed a weaker temperature response. We suggest that the miss-associated recombination of P680 + QA - is the main producer of 1 O2 . Our results indicate three parallel photoinhibition mechanisms. The manganese mechanism dominates in UV radiation but also functions in white light. Mechanisms that depend on light absorption by Chls, having 1 O2 or long-lived P680 + as damaging agents, dominate in red light.

Evaluation of the agreement between the ACR 1990 fibromyalgia tender points and an enthesitis score in patients with axial spondyloarthritis.

OBJECTIVES: Coexistence of FM represents a challenge in the evaluation of enthesitis in patients with axial spondyloarthritis (axSpA) due to a possible overlap between the tender points (TP) due to enthesitis and those of FM. The objective was to assess the agreement between the MASES enthesitis score and the tender points of the ACR 1990 criteria in patients with axSpA. METHODS: This was a cross-sectional ancillary analysis of the Predict-SpA study (NCT03039088). Patients had a diagnosis of axSpA according to their rheumatologist and an indication to start a TNFα blocker. All patients were screened for FM according to the FiRST questionnaire. A physician was asked to assess 31 anatomically described sites in a random order without knowing to which instrument the site belonged (i.e. the 18 ACR 1990 TP and the 13 MASES sites). Agreement between the MASES and the ACR 1990 TPs by the intraclass correlation coefficient (ICC), also stratified by the presence/absence of concomitant FM according to the FiRST. RESULTS: Among the 526 patients, 53% were men and 202 (38%) had FM. Radiographic sacroiliitis and MRI sacroiliitis were present in 56% and 68% patients, respectively. Patients were mostly men (53.4%) with radiographic and MRI sacroiliitis in 56% and 68% patients, respectively. Mean number of ACR 1990 TP was 5.4 (s.d. 4.6) and mean MASES was 4.2 (s.d. 3.6). ICC between both scores was 0.7 [95% CI (0.6, 0.8)]. ICC between both scores was 0.6 [95% CI (0.3, 0.8)] and 0.7 [95% CI (0.6, 0.7)] for patients with and without FM, respectively. CONCLUSION: These results suggest a significant overlap between both scores in patients with axSpA, including in those without concomitant FM. TRIAL REGISTRATION: clinicaltrials.gov, https://clinicaltrials.gov, NCT03039088.

New foundations for the physical mechanism of variable chlorophyll a fluorescence. Quantum efficiency versus the light-adapted state of photosystem II.

Photosystem II (PSII) uses solar energy to oxidize water and delivers electrons to fix CO2. Although the structure at atomic resolution and the basic photophysical and photochemical functions of PSII are well understood, many important questions remain. The activity of PSII in vitro and in vivo is routinely monitored by recording the induction kinetics of chlorophyll a fluorescence (ChlF). According to the 'mainstream' model, the rise from the minimum level (Fo) to the maximum (Fm) of ChlF of dark-adapted PSII reflects the closure of all functionally active reaction centers, and the Fv/Fm ratio is equated with the maximum photochemical quantum yield of PSII (where Fv=Fm-Fo). However, this model has never been free of controversies. Recent experimental data from a number of studies have confirmed that the first single-turnover saturating flash (STSF), which generates the closed state (PSIIC), produces F1

Prediction of the Contribution Ratio of a Target Metabolic Enzyme to Clearance from Chemical Structure Information.

The contribution ratio of metabolic enzymes such as cytochrome P450 to in vivo clearance (fraction metabolized: fm) is a pharmacokinetic index that is particularly important for the quantitative evaluation of drug-drug interactions. Since obtaining experimental in vivo fm values is challenging, those derived from in vitro experiments have often been used alternatively. This study aimed to explore the possibility of constructing machine learning models for predicting in vivo fm using chemical structure information alone. We collected in vivo fm values and chemical structures of 319 compounds from a public database with careful manual curation and constructed predictive models using several machine learning methods. The results showed that in vivo fm values can be obtained from structural information alone with a performance comparable to that based on in vitro experimental values and that the prediction accuracy for the compounds involved in CYP induction or inhibition is significantly higher than that by using in vitro values. Our new approach to predicting in vivo fm values in the early stages of drug discovery should help improve the efficiency of the drug optimization process.

Assessing the impact of non-nutritive sweeteners on anthropometric indices and leptin levels in adults: A GRADE-assessed systematic review, meta-analysis, and meta-regression of randomized clinical trials.

In today's world, non-nutritive sweeteners (NNSs) are recognized as substitutes for sugar or other high-calorie sweeteners, and their consumption is increasing dramatically. However, there is ongoing debate regarding the impact of NNSs on anthropometric indices. To fill this gap in knowledge, the current GRADE-assessed systematic review and meta-analysis of randomized controlled trials (RCTs) was conducted to evaluate the effects of artificial- and stevia-based sweeteners consumption on anthropometric indices and serum leptin level which is known as an appetite-regulating hormone. A comprehensive search was conducted on the Scopus, PubMed, and Embase databases up to November 2022 to identify randomized controlled trials (RCTs) investigating the effects of NNSs on anthropometric indices and serum leptin levels. Data extraction from qualified studies was performed independently by two researchers. A random- or fixed-effects model was used to estimate weighted mean differences (WMDs) and 95% confidence intervals (CIs) for anthropometric indices such as body weight (BW), body mass index (BMI), fat mass (FM), fat-free mass (FFM), waist circumference (WC) and serum leptin level. Heterogeneity between studies was assessed using Cochran's Q test and quantified using the I2 statistic. From a pool of 3212 studies initially identified, 20 studies with a total sample size of 2158 subjects were included in the analysis. Results of the pooled analysis showed that NNSs consumption had a significant reducing effect on BW (WMD: -1.02, 95% CI: -1.57, -0.46 Kg), FM (WMD: -1.09, 95% CI: -1.90, -0.29), and FFM (WMD: -0.83, 95% CI: -1.42, -0.23), but did not have any significant effect on BMI (WMD: -0.16, 95% CI: -0.35, 0.02), WC (WMD: -1.03, 95% CI: -2.77, 0.72), or serum leptin level (WMD: -2.17, 95% CI: -4.98, 0.65). The findings of this study indicate that the consumption of artificial- and stevia-based sweeteners may lead to a reduction in body weight, fat mass, and free fat mass.

Inositol hexakisphosphate primes syndapin I/PACSIN 1 activation in endocytosis.

Endocytosis is controlled by a well-orchestrated molecular machinery, where the individual players as well as their precise interactions are not fully understood. We now show that syndapin I/PACSIN 1 is expressed in pancreatic ß cells and that its knockdown abrogates ß cell endocytosis leading to disturbed plasma membrane protein homeostasis, as exemplified by an elevated density of L-type Ca2+ channels. Intriguingly, inositol hexakisphosphate (InsP6) activates casein kinase 2 (CK2) that phosphorylates syndapin I/PACSIN 1, thereby promoting interactions between syndapin I/PACSIN 1 and neural Wiskott-Aldrich syndrome protein (N-WASP) and driving ß cell endocytosis. Dominant-negative interference with endogenous syndapin I/PACSIN 1 protein complexes, by overexpression of the syndapin I/PACSIN 1 SH3 domain, decreases InsP6-stimulated endocytosis. InsP6 thus promotes syndapin I/PACSIN 1 priming by CK2-dependent phosphorylation, which endows the syndapin I/PACSIN 1 SH3 domain with the capability to interact with the endocytic machinery and thereby initiate endocytosis, as exemplified in ß cells.

Application of ferromanganese functionalized biochar simultaneously reduces Cd and Pb uptake of wheat in contaminated alkaline soils.

The reduction of Cd and Pb accumulation in wheat grains grown on Cd and Pb contaminated alkaline soils is a pressing issue that needs to be solved. In this study, ferromanganese functionalized biochar (FM-BC) was used to remediate Cd and Pb contaminated alkaline soils and mitigate Cd and Pb accumulation in wheat grains. The immobilization capacity and mechanism of FM-BC were investigated by Fourier transform infrared spectroscopy (FT-IR) and X-ray diffraction (XRD) characterization and chemical analysis. Fe and Mn loaded on FM-BC improved the removal efficiencies of DTPA-Cd and DTPA-Pb in soil with DTPA-Cd removal of 22.99%- 52.04% (JM22) and 25.54%- 53.32 (AK58) and DTPA-Pb removal of 11.39%- 22.36% (JM22) and 5.38%- 13.00% (AK58). The FT-IR and XRD results indicated that the complexation and precipitation of Cd and Pb with the Fe-Mn oxides and the oxygen-containing functional groups on biochar surface stabilized the Cd and Pb in soil for the observation of Cd2Mn3O8, PbHPO4, CdCO3, and PbO2 on FM-BC isolated from contaminated soils. FM-BC with excellent adsorption capacity reduced the available Cd and Pb in the soil, therefore, thereby inhibiting the Cd and Pb accumulation in wheat. In the 3% FM-BC treatment, Cd and Pb contents in wheat grains were lower than 0.10 mg/kg and 0.20 mg/kg, respectively, reaching the national safety standards. And FM-BC increased the Fe, Mn, Na and Zn contents in wheat grains, and improved the growth and yield of wheat. These findings suggest that FM-BC can be considered a prospective and effective material for remediation of alkaline soils contaminated with Cd and Pb.

Comparative study on plant growth-promoting bacterial inoculation by irrigation and spraying for promoting Bidens pilosa L. phytoremediation of cadmium-contaminated soil.

A field study was conducted to compare FM-1 inoculation by irrigation and spraying for promoting Bidens pilosa L. phytoremediation of cadmium (Cd)-contaminated soil. Cascading relationships between bacterial inoculation by irrigation and spraying and soil properties, plant growth-promoting traits, plant biomass and Cd concentrations in Bidens pilosa L. were explored based on the partial least squares path model (PLS-PM). The results indicated that inoculation with FM-1 not only improved the rhizosphere soil environment of B. pilosa L. but also increased the Cd extracted from the soil. Moreover, Fe and P in leaves play vital roles in promoting plant growth when FM-1 is inoculated by irrigation, while Fe in leaves and stems plays a vital role in promoting plant growth when FM-1 is inoculated by spraying. In addition, FM-1 inoculation decreased the soil pH by affecting soil dehydrogenase and oxalic acid in cases with irrigation and Fe in roots in cases with spraying. Thus, the soil bioavailable Cd content increased and promoted Cd uptake by Bidens pilosa L. To address Cd-induced oxidative stress, Fe in leaves helped to convert GSH into PCs, which played a vital role in ROS scavenging when FM-1 was inoculated by irrigation. The soil urease content effectively increased the POD and APX activities in the leaves of Bidens pilosa L., which helped alleviate Cd-induced oxidative stress when FM-1 was inoculated by spraying. This study compares and illustrates the potential mechanism by which FM-1 inoculation can improve the phytoremediation of Cd-contaminated soil by Bidens pilosa L., suggesting that FM-1 inoculation by irrigation and spraying is useful in the phytoremediation of Cd-contaminated sites.

Perspectives of family medicine residents in Riyadh on leadership training: a cross-sectional study.

BACKGROUND: Medical educators in academia have faced challenges incorporating leadership training into curricula while minimizing redundancy and assuring value and relevance for all learners. This study aims to assess the status of leadership training as perceived by family medicine residents in Riyadh to advise the development of a formal leadership training curriculum. METHOD: The research is cross-sectional and quantitative. Participants were asked via an electronic questionnaire about their leadership attitudes, perceived degree of training in various leadership domains, and where they could find additional training. RESULTS: The survey was completed by 270 family medicine residents in Riyadh. Residents rated the importance of physician leadership in their communities as high (6 out of 7 on a Likert scale). In contrast, agreement with the statement 'I am a leader' obtained the lowest grade (4.4 of 7 on a Likert scale). Overall, most of the residents participating in the study (50% or more) voiced a desire for more training in all leadership domains. Over 50% of residents indicated that leadership electives or selective lectures, workshops, or seminars as well as WADAs (Weekly Academic Day Activities), leadership mentors or coaches teaching junior learners (with training), and leadership courses could be incorporated into the curriculum to foster leadership skills. CONCLUSION: Residents were enthusiastic about family physicians being leaders, aligning with the current educational philosophy but requiring formal training. They also indicated areas where leadership training might be improved and developed in the current curriculum. This poll's results could be used to help residents build leadership skills by incorporating them into a formal leadership curriculum.

FM1-43 Dye Memorizes Piezo1 Activation in the Trigeminal Nociceptive System Implicated in Migraine Pain.

It has been proposed that mechanosensitive Piezo1 channels trigger migraine pain in trigeminal nociceptive neurons, but the mechanosensitivity of satellite glial cells (SGCs) supporting neuronal sensitization has not been tested before. Moreover, tools to monitor previous Piezo1 activation are not available. Therefore, by using live calcium imaging with Fluo-4 AM and labeling with FM1-43 dye, we explored a new strategy to identify Piezo channels' activity in mouse trigeminal neurons, SGCs, and isolated meninges. The specific Piezo1 agonist Yoda1 induced calcium transients in both neurons and SGCs, suggesting the functional expression of Piezo1 channels in both types of cells. In Piezo1-transfected HEK cells, FM1-43 produced only a transient fluorescent response, whereas co-application with Yoda1 provided higher transient signals and a remarkable long-lasting FM1-43 'tail response'. A similar Piezo1-related FM1-43 trapping was observed in neurons and SGCs. The non-specific Piezo channel blocker, Gadolinium, inhibited the transient peak, confirming the involvement of Piezo1 receptors. Finally, FM1-43 labeling demonstrated previous activity in meningeal tissues 3.5 h after Yoda1 washout. Our data indicated that trigeminal neurons and SGCs express functional Piezo channels, and their activation provides sustained labeling with FM1-43. This long-lasting labelling can be used to monitor the ongoing and previous activation of Piezo1 channels in the trigeminal nociceptive system, which is implicated in migraine pain.

REV7 Monomer Is Unable to Participate in Double Strand Break Repair and Translesion Synthesis but Suppresses Mitotic Errors.

Rev7 is a regulatory protein with roles in translesion synthesis (TLS), double strand break (DSB) repair, replication fork protection, and cell cycle regulation. Rev7 forms a homodimer in vitro using its HORMA (Hop, Rev7, Mad2) domain; however, the functional importance of Rev7 dimerization has been incompletely understood. We analyzed the functional properties of cells expressing either wild-type mouse Rev7 or Rev7K44A/R124A/A135D, a mutant that cannot dimerize. The expression of wild-type Rev7, but not the mutant, rescued the sensitivity of Rev7-/- cells to X-rays and several alkylating agents and reversed the olaparib resistance phenotype of Rev7-/- cells. Using a novel fluorescent host-cell reactivation assay, we found that Rev7K44A/R124A/A135D is unable to promote gap-filling TLS opposite an abasic site analog. The Rev7 dimerization interface is also required for shieldin function, as both Rev7-/- cells and Rev7-/- cells expressing Rev7K44A/R124A/A135D exhibit decreased proficiency in rejoining some types of double strand breaks, as well as increased homologous recombination. Interestingly, Rev7K44A/R124A/A135D retains some function in cell cycle regulation, as it maintains an interaction with Ras-related nuclear protein (Ran) and partially rescues the formation of micronuclei. The mutant Rev7 also rescues the G2/M accumulation observed in Rev7-/- cells but does not affect progression through mitosis following nocodazole release. We conclude that while Rev7 dimerization is required for its roles in TLS, DSB repair, and regulation of the anaphase promoting complex, dimerization is at least partially dispensable for promoting mitotic spindle assembly through its interaction with Ran.

Effects of Environmental Conditions on the Individual Architectures and Photosynthetic Performances of Three Species in Drosera.

The aim of this study was to determine the environmental conditions, individual architectures, and photosynthetic efficiencies of three sundew species: Drosera rotundifolia, D. anglica, and D. intermedia, found in well-preserved peatlands and sandy lake shores in NW Poland. Morphological traits and chlorophyll a fluorescence (Fv/Fm) were measured in 581 individuals of Drosera. D. anglica occupies the best-lit and warmest habitats, and also those that are the most heavily hydrated and the richest in organic matter; its rosettes are larger under conditions of higher pH, less organic matter, and less well-lit habitats. D. intermedia occupies substrates with the highest pH but the lowest conductivity, the poorest level of organic matter, and the least hydration. It is highly variable in terms of individual architecture. D. rotundifolia occupies habitats that are the most diverse, and that are often poorly lit, with the lowest pH but the highest conductivity. It is the least variable in terms of individual architecture. The value of the Fv/Fm ratio in Drosera is low (0.616 ± 0.137). The highest photosynthetic efficiency is achieved by D. rotundifolia (0.677 ± 0.111). It is significant for all substrates, indicating its high phenotypic plasticity. The other species have lower and similar Fv/Fm values (D. intermedia, 0.571 ± 0.118; D. anglica, 0.543 ± 0.154). Due to its very low photosynthetic efficiency, D. anglica avoids competition by occupying highly hydrated habitats. D. intermedia has adapted to the occupation of highly variable habitats in terms of hydration, while D. rotundifolia is primarily adapted to variable light conditions.