A comparative genomics examination of desiccation tolerance and sensitivity in two sister grass species.

Desiccation tolerance is an ancient and complex trait that spans all major lineages of life on earth. Although important in the evolution of land plants, the mechanisms that underlay this complex trait are poorly understood, especially for vegetative desiccation tolerance (VDT). The lack of suitable closely related plant models that offer a direct contrast between desiccation tolerance and sensitivity has hampered progress. We have assembled high-quality genomes for two closely related grasses, the desiccation-tolerant Sporobolus stapfianus and the desiccation-sensitive Sporobolus pyramidalis Both species are complex polyploids; S. stapfianus is primarily tetraploid, and S. pyramidalis is primarily hexaploid. S. pyramidalis undergoes a major transcriptome remodeling event during initial exposure to dehydration, while S. stapfianus has a muted early response, with peak remodeling during the transition between 1.5 and 1.0 grams of water (gH2O) g-1 dry weight (dw). Functionally, the dehydration transcriptome of S. stapfianus is unrelated to that for S. pyramidalis A comparative analysis of the transcriptomes of the hydrated controls for each species indicated that S. stapfianus is transcriptionally primed for desiccation. Cross-species comparative analyses indicated that VDT likely evolved from reprogramming of desiccation tolerance mechanisms that evolved in seeds and that the tolerance mechanism of S. stapfianus represents a recent evolution for VDT within the Chloridoideae. Orthogroup analyses of the significantly differentially abundant transcripts reconfirmed our present understanding of the response to dehydration, including the lack of an induction of senescence in resurrection angiosperms. The data also suggest that failure to maintain protein structure during dehydration is likely critical in rendering a plant desiccation sensitive.

Divergent structural leaf trait spectra in succulent versus non-succulent plant taxa.

BACKGROUND AND SCOPE: Plant functional traits are the result of natural selection to optimize carbon gain, leading to a broad spectrum of traits across environmental gradients. Among plant traits, leaf water storage capacity is paramount for plant drought resistance. We explored whether leaf-succulent taxa follow trait correlations similar to those of non-leaf-succulent taxa to evaluate whether both are similarly constrained by relationships between leaf water storage and climate. METHODS: We tested the relationships among three leaf traits related to water storage capacity and resource use strategies in 132 species comprising three primary leaf types: succulent, sclerophyllous, and leaves with rapid returns on water investment, referred to as fast return. Correlation coefficients among specific leaf area (SLA), water mass per unit of area (WMA), and saturated water content (SWC) were tested, along with relationships between leaf trait spectra and aridity determined from species occurrence records. RESULTS: Both SWC and WMA at a given SLA were ~10-fold higher in succulent leaves than in non-succulent leaves. While SWC actually increased with SLA in non-succulent leaves, no relationship was detected between SWC and SLA in succulent leaves, although WMA decreased with SLA in all leaf types. A principal component analysis (PCA) revealed that succulent taxa occupied a widely different mean trait space than either fast-return (P < 0.0001) or sclerophyllous (P < 0.0001) taxa along the first PCA axis, which explained 63 % of mean trait expression among species. However, aridity only explained 12 % of the variation in PCA1 values. This study is among the first to establish a structural leaf trait spectrum in succulent leaf taxa and quantify contrasts in leaf water storage among leaf types relative to specific leaf area. CONCLUSIONS: Trait coordination in succulent leaf taxa may not follow patterns similar to those of widely studied non-succulent taxa.

Global change impacts on cacti (Cactaceae): current threats, challenges and conservation solutions.

BACKGROUND: The plant family Cactaceae provides some of the most striking examples of adaptive evolution, expressing undeniably the most spectacular New World radiation of succulent plants distributed across arid and semi-arid regions of the Americas. Cacti are widely regarded for their cultural, economic and ecological value, yet they are also recognized as one of the most threatened and endangered taxonomic groups on the planet. SCOPE: This paper reviews current threats to species of cacti that have distributions in arid to semi-arid subtropical regions. Our review focuses primarily on four global change forces: (1) increases in atmospheric CO2 concentrations; (2) increases in mean annual temperatures and heat waves; (3) increases in the duration, frequency and intensity of droughts; and (4) and increases in competition and wildfire frequency from invasion by non-native species. We provide a broad range of potential priorities and solutions for stemming the extinction risk of cacti species and populations. CONCLUSIONS: Mitigating ongoing and emerging threats to cacti will require not only strong policy initiatives and international cooperation, but also new and creative approaches to conservation. These approaches include determining species at risk from climate extremes, enhancing habitat quality after disturbance, approaches and opportunities for ex situ conservation and restoration, and the potential use of forensic tools for identifying plants that have been removed illegally from the wild and sold on open markets.

Measurement of Cutting Temperature in Interrupted Machining Using Optical Spectrometry.

This research presents an experimental study focused on measuring temperature at the tool flank during the up-milling process at high cutting speed. The proposed system deals with emissivity compensation through a two-photodetector system and during calibration. A ratio pyrometer composed of two photodetectors and a multimode fiber-optic coupler is employed to capture the radiation emitted by the cutting insert. The pyrometer is calibrated using an innovative calibration system that addresses theoretical discrepancies arising from various factors affecting the measurement of cutting temperature. This calibration system replicates the milling process to generate a calibration curve. Experimentally, AISI 4140 steel is machined with coated tungsten carbide inserts, using cutting speeds of 300 and 400 m/min, and feed rates of 0.08 and 0.16 mm/tooth. The results reveal a maximum recorded cutting temperature of 518 °C and a minimum of 304 °C. The cutting temperature tends to increase with higher cutting speeds and feed rates, with cutting speed being the more influential factor in this increase. Both the pyrometer calibration and experimental outcomes yield satisfactory results. Finally, the results showed that the process and the device prove to be a convenient, effective, and precise method of measuring cutting temperature in machine processes.

Impact of the COVID-19 pandemic on the incidence, etiology, and antimicrobial resistance of healthcare-associated infections in a critical care unit in Western Qatar.

BACKGROUND: Healthcare-associated infections (HAIs) in critical patients affect the quality and safety of patient care and increase patient morbidity and mortality. During the COVID-19 pandemic, an increase in the incidence of HAIs, particularly device-associated infections (DAIs), was reported worldwide. In this study, we aimed to estimate the incidence of HAIs in an intensive care unit (ICU) during a 10-year period and compare HAI incidence during the preCOVID-19 and COVID-19 periods. METHODS: A retrospective, observational study of HAIs in the medical-surgical ICU at The Cuban Hospital was conducted. DAIs included central line-associated bloodstream infections (CLABSI), catheter-associated urinary tract infections (CAUTI), and ventilator-associated pneumonia (VAP). Data included the annual incidence of HAIs, etiology, and antimicrobial resistance, using definitions provided by the Centers for Disease Control and Prevention, except for other respiratory tract infections (RTIs). RESULTS: 155 patients with HAI infections were reported, from which 130 (85.5%) were identified during the COVID-19 period. The frequencies of DAIs and non-DAIs were higher during the COVID-19 period, except for Clostridium difficile infections. Species under Enterobacter, Klebsiella, and Pseudomonas dominated in both periods, and higher frequencies of Acinetobacter, Enterococcus, Candida, Escherichia coli, Serratia marcescens, and Stenotrophoma maltophila were noted during COVID-19 period. Device utilization ratio increased to 10.7% for central lines and 12.9% for ventilators, while a reduction of 15% in urinary catheter utilization ratio was observed. DAI incidence was higher during the COVID-19 pandemic, with risks for CLABSI, VAP, and CAUTI increased by 2.79 (95% confidence interval, 0.93-11.21; p < 0.0050), 15.31 (2.53-625.48), and 3.25 (0.68-31.08), respectively. CONCLUSION: The incidence of DAIs increased during the pandemic period, with limited evidence of antimicrobial resistance observed. The infection control program should evaluate strategies to minimize the impact of the pandemic on HAIs.

Ten-year incidence and impact of coronavirus infections on incidence, etiology, and antimicrobial resistance of healthcare-associated infections in a critical care unit in Western Qatar.

BACKGROUND: Healthcare-associated infections (HAI) in critical patients affect the quality and safety of patient care as they impact morbidity and mortality. During the COVID-19 pandemic, an increase in the incidence rate was reported worldwide. We aim to describe the incidence of HAI in the intensive care unit (ICU) during a 10-year follow-up period and compare the incidence during the pre-COVID-19 and COVID-19 periods. METHODS: A retrospective observational study of HAI in the medical-surgical ICU at The Cuban Hospital was conducted. The data collected include the annual incidence of HAI, its etiology, and antimicrobial resistance, using the Centers for Disease Control and Prevention definitions, except for other respiratory tract infections (RTIs). RESULTS: A total of 155 patients had HAI, of which 130 (85.5%) were identified during COVID-19. The frequency of device-associated infections (DAI) and non-DAI was higher during COVID-19, except for Clostridium difficile infections. Etiology was frequently related to species of Enterobacter, Klebsiella, and Pseudomonas in both periods, and a higher frequency of Acinetobacter, Enterococcus, Candida, Escherichia coli, Serratia marcescens, and Stenotrophomonas maltophilia was noted during the COVID-19 period. Device utilization ratio increased by 10.7% for central lines and 12.9% for ventilators, while a reduction of 15% in urinary catheter utilization ratio was observed. DAI incidence was higher during the COVID-19, with a 2.79 higher risk of infection (95% CI: 0.93-11.21; p < 0.0050), 15.31 (2.53-625.48), and 3.25 (0.68-31.08) for CLABSI, VAP, and CAUTI, respectively. CONCLUSION: The incidence of DAI increased during the pandemic period as compared to the pre-pandemic period, and limited evidence of the impact on antimicrobial resistance was observed. The infection control program should evaluate strategies to minimize the impact of pandemics on HAI.

Catheter-associated urinary tract infection and urinary catheter utilization ratio over 9 years, and the impact of the COVID-19 pandemic on the incidence of infection in medical and surgical wards in a single facility in Western Qatar.

INTRODUCTION: Catheter-associated urinary tract infection (CAUTI) is a frequently reported healthcare-associated infection in critical and non-critical patients. Limited data are available about CAUTI incidence in non-critical patients. We aim to describe the incidence of CAUTI over 9 years and evaluate the impact of the pandemic on the incidence in non-critical acute care patients. METHODS: A retrospective observational study of CAUTI in medical-surgical and maternity wards was carried out at a public hospital in the west of the State of Qatar. Data collected included the annual CAUTI incidence (per 1,000 device days), urinary catheter utilization ratio (UC-UR), etiology, and antimicrobial resistance. RESULTS: 115,238 patient days and 6,681 urinary catheters (UC) days were recorded over the study period, and 9 and 4 CAUTI were confirmed in medical-surgical and maternity wards, respectively. The infection rate was 1.9 per 1,000 UC days, and the UC-UR was 0.06. The CAUTI rate was higher in medical-surgical wards over the COVID-19 period (2.4 × 1,000 UC days) in comparison with the non-COVID-19 period (1.7 × 1,000 UC days) (RR 1.46; 1.12-1.80). However, in the maternity ward, the result was 0 and 2.5 × 1,000 UC days during these periods, respectively. No differences were observed in the infection rate among periods for all patients (RR 1.06; 0.81-1.31). Multidrug-resistant organisms were identified in 7 patients, and non-multidrug-resistant in 6 cases. CONCLUSION: The study findings describe a lower CAUTI risk over 9 years in non-critical acute care patients. The impact of COVID-19 on the CAUTI risk is mainly related to medical patients who had previously been admitted to critical care. The infection control program should consider these data as a benchmark for quality improvement.

A Recurrent Missense Variant in AP2M1 Impairs Clathrin-Mediated Endocytosis and Causes Developmental and Epileptic Encephalopathy.

The developmental and epileptic encephalopathies (DEEs) are heterogeneous disorders with a strong genetic contribution, but the underlying genetic etiology remains unknown in a significant proportion of individuals. To explore whether statistical support for genetic etiologies can be generated on the basis of phenotypic features, we analyzed whole-exome sequencing data and phenotypic similarities by using Human Phenotype Ontology (HPO) in 314 individuals with DEEs. We identified a de novo c.508C>T (p.Arg170Trp) variant in AP2M1 in two individuals with a phenotypic similarity that was higher than expected by chance (p = 0.003) and a phenotype related to epilepsy with myoclonic-atonic seizures. We subsequently found the same de novo variant in two individuals with neurodevelopmental disorders and generalized epilepsy in a cohort of 2,310 individuals who underwent diagnostic whole-exome sequencing. AP2M1 encodes the µ-subunit of the adaptor protein complex 2 (AP-2), which is involved in clathrin-mediated endocytosis (CME) and synaptic vesicle recycling. Modeling of protein dynamics indicated that the p.Arg170Trp variant impairs the conformational activation and thermodynamic entropy of the AP-2 complex. Functional complementation of both the µ-subunit carrying the p.Arg170Trp variant in human cells and astrocytes derived from AP-2µ conditional knockout mice revealed a significant impairment of CME of transferrin. In contrast, stability, expression levels, membrane recruitment, and localization were not impaired, suggesting a functional alteration of the AP-2 complex as the underlying disease mechanism. We establish a recurrent pathogenic variant in AP2M1 as a cause of DEEs with distinct phenotypic features, and we implicate dysfunction of the early steps of endocytosis as a disease mechanism in epilepsy.

Cut-off points for serum ferritin to identify low iron stores during the first year of life in a cohort of Mexican infants.

The aim of this study was to identify serum ferritin (SF) cut-off points (COPs) in a cohort of healthy full-term normal birth weight infants who had repeated measurements of SF and haemoglobin every 3 months during the first year of life. The study included 746 full-term infants with birth weight ≥2,500 g, having uncomplicated gestations and births. Participants received prophylactic iron supplementation (1 mg/day of iron element) from the first to the 12th month of life and did not develop anaemia during the first year of life. Two statistical methods were considered to identify COPs for low iron stores at 3, 6, 9 and 12 months of age: deviation from mean and cluster analysis. According to the K-means cluster analysis results by age and sex, COPs at 3 and 6 months for girls were 39 and 21 µg/L and for boys 23 and 11 µg/L, respectively. A single COP of 10 µg/L was identified, for girls and boys, at both 9 and 12 months. Given the physiological changes in SF concentration during the first year of life, our study identified dynamic COPs, which differed by sex in the first semester. Adequate SF COPs are necessary to identify low iron stores at an early stage of iron deficiency, which represents one of the most widespread public health problems around the world, particularly in low- and middle-income countries.

Why Are There So Many Flowering Plants? A Multiscale Analysis of Plant Diversification.

The causes of the rapid diversification and extraordinary richness of flowering plants (angiosperms) relative to other plant clades is a long-standing mystery. Angiosperms are only one among 10 major land plant clades (phyla) but include ∼90% of land plant species. However, most studies that have tried to identify which traits might explain the remarkable diversification of angiosperms have focused only on richness patterns within angiosperms and tested only one or a few traits at a single hierarchical scale. Here, we assemble a database of 31 diverse traits among 678 families and analyze relationships between traits and diversification rates across all land plants at three hierarchical levels (phylum, order, and family) using phylogenetic multiple regression. We find that most variation (∼85%) in diversification rates among major clades (phyla) is explained by biotically mediated fertilization (e.g., insect pollination) and clade-level geographic range size. Different sets of traits explain diversification at different hierarchical levels, with geographic range size dominating among families. Surprisingly, we find that traits related to local-scale species interactions (i.e., biotic fertilization) are particularly important for explaining diversification patterns at the deepest timescales, whereas large-scale geographic factors (i.e., clade-level range size) are more important at shallower timescales. This dichotomy might apply broadly across organisms.

Depolarization causes the formation of a ternary complex between GlialCAM, MLC1 and ClC-2 in astrocytes: implications in megalencephalic leukoencephalopathy.

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a rare type of leukodystrophy caused by mutations in either MLC1 or GLIALCAM. GlialCAM is necessary for the correct targeting of MLC1, but also for the targeting of the Cl- channel ClC-2. Furthermore, GlialCAM modifies ClC-2 functional properties in vitro. However, in vivo studies in GlialCAM-/- mice have shown that the modification of ClC-2 activity only occurs in oligodendrocytes, despite GlialCAM and ClC-2 being expressed in astrocytes. Thus, the relationship between GlialCAM, MLC1 and ClC-2 in astrocytes is unknown. Here, we show that GlialCAM, ClC-2 and MLC1 can form a ternary complex in cultured astrocytes, but only under depolarizing conditions. We also provide biochemical evidences that this ternary complex exists in vivo. The formation of this complex changes ClC-2 localization in the membrane and its functional properties. ClC-2 association with GlialCAM/MLC1 depends on calcium flux through L-type calcium channels and activation of calcium-dependent calpain proteases. Based on these studies, we propose that the chloride influx mediated by GlialCAM/MLC1/ClC-2 in astrocytes may be needed to compensate an excess of potassium, as occurs in conditions of high neuronal activity. We suggest that a defect in this compensation may contribute to the pathogenesis of MLC disease.

Phylogenetic Relationships and Evolutionary Trends in the Cactus Family.

Members of the cactus family are keystone species of arid and semiarid biomes in the Americas, as they provide shelter and resources to support other members of ecosystems. Extraordinary examples are the several species of flies of the genus Drosophila that lay eggs and feed in their rotting stems, which provide a model system for studying evolutionary processes. Although there is significant progress in understanding the evolution of Drosophila species, there are gaps in our knowledge about the cactus lineages hosting them. Here, we review the current knowledge about the evolution of Cactaceae, focusing on phylogenetic relationships and trends revealed by the study of DNA sequence data. During the last several decades, the availability of molecular phylogenies has considerably increased our understanding of the relationships, biogeography, and evolution of traits in the family. Remarkably, although succulent cacti have very low growth rates and long generation times, they underwent some of the fastest diversifications observed in the plant kingdom, possibly fostered by strong ecological interactions. We have a better understanding of the reproductive biology, population structure and speciation mechanisms in different clades. The recent publication of complete genomes for some species has revealed the importance of phenomena such as incomplete lineage sorting. Hybridization and polyploidization are common in the family, and have been studied using a variety of phylogenetic methods. We discuss potential future avenues for research in Cactaceae, emphasizing the need of a concerted effort among scientists in the Americas, together with the analyses of data from novel sequencing techniques.

Streptococcal Immunity Is Constrained by Lack of Immunological Memory following a Single Episode of Pyoderma.

The immunobiology underlying the slow acquisition of skin immunity to group A streptococci (GAS), is not understood, but attributed to specific virulence factors impeding innate immunity and significant antigenic diversity of the type-specific M-protein, hindering acquired immunity. We used a number of epidemiologically distinct GAS strains to model the development of acquired immunity. We show that infection leads to antibody responses to the serotype-specific determinants on the M-protein and profound protective immunity; however, memory B cells do not develop and immunity is rapidly lost. Furthermore, antibodies do not develop to a conserved M-protein epitope that is able to induce immunity following vaccination. However, if re-infected with the same strain within three weeks, enduring immunity and memory B-cells (MBCs) to type-specific epitopes do develop. Such MBCs can adoptively transfer protection to naïve recipients. Thus, highly protective M-protein-specific MBCs may never develop following a single episode of pyoderma, contributing to the slow acquisition of immunity and to streptococcal endemicity in at-risk populations.

Reversal of Ethanol-induced Intoxication by a Novel Modulator of Gßγ Protein Potentiation of the Glycine Receptor.

The acute intoxicating effects of ethanol in the central nervous system result from the modulation of several molecular targets. It is widely accepted that ethanol enhances the activity of the glycine receptor (GlyR), thus enhancing inhibitory neurotransmission, leading to motor effects, sedation, and respiratory depression. We previously reported that small peptides interfered with the binding of Gßγ to the GlyR and consequently inhibited the ethanol-induced potentiation of the receptor. Now, using virtual screening, we identified a subset of small molecules capable of interacting with the binding site of Gßγ. One of these compounds, M554, inhibited the ethanol potentiation of the GlyR in both evoked currents and synaptic transmission in vitro When this compound was tested in vivo in mice treated with ethanol (1-3.5 g/kg), it was found to induce a faster recovery of motor incoordination in rotarod experiments and a shorter sedative effect in loss of righting reflex assays. This study describes a novel molecule that might be relevant for the design of useful therapeutic compounds in the treatment of acute alcohol intoxication.

Lysosomal Dysfunction Caused by Cellular Accumulation of Silica Nanoparticles.

Nanoparticles (NPs) are widely used as components of drugs or cosmetics and hold great promise for biomedicine, yet their effects on cell physiology remain poorly understood. Here we demonstrate that clathrin-independent dynamin 2-mediated caveolar uptake of surface-functionalized silica nanoparticles (SiNPs) impairs cell viability due to lysosomal dysfunction. We show that internalized SiNPs accumulate in lysosomes resulting in inhibition of autophagy-mediated protein turnover and impaired degradation of internalized epidermal growth factor, whereas endosomal recycling proceeds unperturbed. This phenotype is caused by perturbed delivery of cargo via autophagosomes and late endosomes to SiNP-filled cathepsin B/L-containing lysosomes rather than elevated lysosomal pH or altered mTOR activity. Given the importance of autophagy and lysosomal protein degradation for cellular proteostasis and clearance of aggregated proteins, these results raise the question of beneficial use of NPs in biomedicine and beyond.

Megalencephalic leukoencephalopathy with subcortical cysts protein 1 regulates glial surface localization of GLIALCAM from fish to humans.

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a leukodystrophy characterized by myelin vacuolization and caused by mutations in MLC1 or GLIALCAM. Patients with recessive mutations in either MLC1 or GLIALCAM show the same clinical phenotype. It has been shown that GLIALCAM is necessary for the correct targeting of MLC1 to the membrane at cell junctions, but its own localization was independent of MLC1 in vitro. However, recent studies in Mlc1(-/-) mice have shown that GlialCAM is mislocalized in glial cells. In order to investigate whether the relationship between Mlc1 and GlialCAM is species-specific, we first identified MLC-related genes in zebrafish and generated an mlc1(-/-) zebrafish. We have characterized mlc1(-/-) zebrafish both functionally and histologically and compared the phenotype with that of the Mlc1(-/-) mice. In mlc1(-/-) zebrafish, as in Mlc1(-/-) mice, Glialcam is mislocalized. Re-examination of a brain biopsy from an MLC patient indicates that GLIALCAM is also mislocalized in Bergmann glia in the cerebellum. In vitro, impaired localization of GlialCAM was observed in astrocyte cultures from Mlc1(-/-) mouse only in the presence of elevated potassium levels, which mimics neuronal activity. In summary, here we demonstrate an evolutionary conserved role for MLC1 in regulating glial surface levels of GLIALCAM, and this interrelationship explains why patients with mutations in either gene (MLC1 or GLIALCAM) share the same clinical phenotype.

Streptococcal toxins: role in pathogenesis and disease.

Group A Streptococcus (Streptococcus pyogenes), group B Streptococcus (Streptococcus agalactiae) and Streptococcus pneumoniae (pneumococcus) are host-adapted bacterial pathogens among the leading infectious causes of human morbidity and mortality. These microbes and related members of the genus Streptococcus produce an array of toxins that act against human cells or tissues, resulting in impaired immune responses and subversion of host physiological processes to benefit the invading microorganism. This toxin repertoire includes haemolysins, proteases, superantigens and other agents that ultimately enhance colonization and survival within the host and promote dissemination of the pathogen.

Insights into MLC pathogenesis: GlialCAM is an MLC1 chaperone required for proper activation of volume-regulated anion currents.

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a rare type of leukodystrophy caused by mutations in either MLC1 or GLIALCAM genes and is associated with myelin and astrocyte vacuolation. It has been suggested that MLC is caused by impaired cell volume regulation as a result of defective activation of astrocytic volume-regulated anion currents (VRAC). GlialCAM brings MLC1 and the ClC-2 Cl(-) channel to cell-cell junctions, even though the role of ClC-2 in MLC disease remains incompletely understood. To gain insights into the biological role of GlialCAM in the pathogenesis of MLC disease, here we analyzed the gain- and loss-of-function phenotypes of GlialCAM in Hela cells and primary astrocytes, focusing on its interaction with the MLC1 protein. Unexpectedly, GlialCAM ablation provoked intracellular accumulation and reduced expression of MLC1 at the plasma membrane. Conversely, over-expression of GlialCAM increased the cellular stability of mutant MLC1 variants. Reduction in GlialCAM expression resulted in defective activation of VRAC and augmented vacuolation, phenocopying MLC1 mutations. Importantly, over-expression of GlialCAM together with MLC1 containing MLC-related mutations was able to reactivate VRAC currents and to reverse the vacuolation caused in the presence of mutant MLC1. These results indicate a previously unrecognized role of GlialCAM in facilitating the biosynthetic maturation and cell surface expression of MLC1, and suggest that pharmacological strategies aimed to increase surface expression of MLC1 and/or VRAC activity may be beneficial for MLC patients.

A metacalibrated time-tree documents the early rise of flowering plant phylogenetic diversity.

The establishment of modern terrestrial life is indissociable from angiosperm evolution. While available molecular clock estimates of angiosperm age range from the Paleozoic to the Late Cretaceous, the fossil record is consistent with angiosperm diversification in the Early Cretaceous. The time-frame of angiosperm evolution is here estimated using a sample representing 87% of families and sequences of five plastid and nuclear markers, implementing penalized likelihood and Bayesian relaxed clocks. A literature-based review of the palaeontological record yielded calibrations for 137 phylogenetic nodes. The angiosperm crown age was bound within a confidence interval calculated with a method that considers the fossil record of the group. An Early Cretaceous crown angiosperm age was estimated with high confidence. Magnoliidae, Monocotyledoneae and Eudicotyledoneae diversified synchronously 135-130 million yr ago (Ma); Pentapetalae is 126-121 Ma; and Rosidae (123-115 Ma) preceded Asteridae (119-110 Ma). Family stem ages are continuously distributed between c. 140 and 20 Ma. This time-frame documents an early phylogenetic proliferation that led to the establishment of major angiosperm lineages, and the origin of over half of extant families, in the Cretaceous. While substantial amounts of angiosperm morphological and functional diversity have deep evolutionary roots, extant species richness was probably acquired later.

GlialCAM, a CLC-2 Cl(-) channel subunit, activates the slow gate of CLC chloride channels.

GlialCAM, a glial cell adhesion molecule mutated in megalencephalic leukoencephalopathy with subcortical cysts, targets the CLC-2 Cl(-) channel to cell contacts in glia and activates CLC-2 currents in vitro and in vivo. We found that GlialCAM clusters all CLC channels at cell contacts in vitro and thus studied GlialCAM interaction with CLC channels to investigate the mechanism of functional activation. GlialCAM slowed deactivation kinetics of CLC-Ka/barttin channels and increased CLC-0 currents opening the common gate and slowing its deactivation. No functional effect was seen for common gate deficient CLC-0 mutants. Similarly, GlialCAM targets the common gate deficient CLC-2 mutant E211V/H816A to cell contacts, without altering its function. Thus, GlialCAM is able to interact with all CLC channels tested, targeting them to cell junctions and activating them by stabilizing the open configuration of the common gate. These results are important to better understand the physiological role of GlialCAM/CLC-2 interaction.