Compensatory growth and recovery of cartilage cytoarchitecture after transient cell death in fetal mouse limbs.

A major question in developmental and regenerative biology is how organ size and architecture are controlled by progenitor cells. While limb bones exhibit catch-up growth (recovery of a normal growth trajectory after transient developmental perturbation), it is unclear how this emerges from the behaviour of chondroprogenitors, the cells sustaining the cartilage anlagen that are progressively replaced by bone. Here we show that transient sparse cell death in the mouse fetal cartilage is repaired postnatally, via a two-step process. During injury, progression of chondroprogenitors towards more differentiated states is delayed, leading to altered cartilage cytoarchitecture and impaired bone growth. Then, once cell death is over, chondroprogenitor differentiation is accelerated and cartilage structure recovered, including partial rescue of bone growth. At the molecular level, ectopic activation of mTORC1 correlates with, and is necessary for, part of the recovery, revealing a specific candidate to be explored during normal growth and in future therapies.

Nuclear factor Y is a pervasive regulator of neuronal gene expression.

Nervous system function relies on the establishment of complex gene expression programs that provide neuron-type-specific and core pan-neuronal features. These complementary regulatory paradigms are controlled by terminal selector and parallel-acting transcription factors (TFs), respectively. Here, we identify the nuclear factor Y (NF-Y) TF as a pervasive direct and indirect regulator of both neuron-type-specific and pan-neuronal gene expression. Mapping global NF-Y targets reveals direct binding to the cis-regulatory regions of pan-neuronal genes and terminal selector TFs. We show that NFYA-1 controls pan-neuronal gene expression directly through binding to CCAAT boxes in target gene promoters and indirectly by regulating the expression of terminal selector TFs. Further, we find that NFYA-1 regulation of neuronal gene expression is important for neuronal activity and motor function. Thus, our research sheds light on how global neuronal gene expression programs are buffered through direct and indirect regulatory mechanisms.

A genomic survey of Clostridioides difficile isolates from hospitalized patients in Melbourne, Australia.

IMPORTANCE: There has been a decrease in healthcare-associated Clostridioides difficile infection in Australia, but an increase in the genetic diversity of infecting strains, and an increase in community-associated cases. Here, we studied the genetic relatedness of C. difficile isolated from patients at a major hospital in Melbourne, Australia. Diverse ribotypes were detected, including those associated with community and environmental sources. Some types of isolates were more likely to carry antimicrobial resistance determinants, and many of these were associated with mobile genetic elements. These results correlate with those of other recent investigations, supporting the observed increase in genetic diversity and prevalence of community-associated C. difficile, and consequently the importance of sources of transmission other than symptomatic patients. Thus, they reinforce the importance of surveillance for in both hospital and community settings, including asymptomatic carriage, food, animals, and other environmental sources to identify and circumvent important sources of C. difficile transmission.

Extensive genome analysis identifies novel plasmid families in Clostridium perfringens.

Globally, the anaerobic bacterium Clostridium perfringens causes severe disease in a wide array of hosts; however, C. perfringens strains are also carried asymptomatically. Accessory genes are responsible for much of the observed phenotypic variation and virulence within this species, with toxins frequently encoded on conjugative plasmids and many isolates carrying up to 10 plasmids. Despite this unusual biology, current genomic analyses have largely excluded isolates from healthy hosts or environmental sources. Accessory genomes, including plasmids, also have often been excluded from broader scale phylogenetic investigations. Here we interrogate a comprehensive collection of 464 C. perfringens genomes and identify the first putative non-conjugative enterotoxin (CPE)-encoding plasmids and a putative novel conjugative locus (Bcp) with sequence similarity to a locus reported from Clostridium botulinum. We sequenced and archived 102 new C. perfringens genomes, including those from rarely sequenced toxinotype B, C, D and E isolates. Long-read sequencing of 11 C. perfringens strains representing all toxinotypes (A-G) identified 55 plasmids from nine distinct plasmid groups. Interrogation of the 464 genomes in this collection identified 1045 plasmid-like contigs from the nine plasmid families, with a wide distribution across the C. perfringens isolates. Plasmids and plasmid diversity play an essential role in C. perfringens pathogenicity and broader biology. We have expanded the C. perfringens genome collection to include temporal, spatial and phenotypically diverse isolates including those carried asymptomatically in the gastrointestinal microbiome. This analysis has resulted in the identification of novel C. perfringens plasmids whilst providing a comprehensive understanding of species diversity.

The proteasome regulator Rpn4 controls antifungal drug tolerance by coupling protein homeostasis with metabolic responses to drug stress.

Fungal pathogens overcome antifungal drug therapy by classic resistance mechanisms, such as increased efflux or changes to the drug target. However, even when a fungal strain is susceptible, trailing or persistent microbial growth in the presence of an antifungal drug can contribute to therapeutic failure. This trailing growth is caused by adaptive physiological changes that enable the growth of a subpopulation of fungal cells in high drug concentrations, in what is described as drug tolerance. Mechanistically, antifungal drug tolerance is incompletely understood. Here we report that the transcriptional activator Rpn4 is important for drug tolerance in the human fungal pathogen Candida albicans. Deletion of RPN4 eliminates tolerance to the commonly used antifungal drug fluconazole. We defined the mechanism and show that Rpn4 controls fluconazole tolerance via two target pathways. First, Rpn4 activates proteasome gene expression, which enables sufficient proteasome capacity to overcome fluconazole-induced proteotoxicity and the accumulation of ubiquitinated proteins targeted for degradation. Consistently, inhibition of the proteasome with MG132 eliminates fluconazole tolerance and resistance, and phenocopies the rpn4Δ/Δ mutant for loss of tolerance. Second, Rpn4 is required for wild type expression of the genes required for the synthesis of the membrane lipid ergosterol. Our data indicates that this function of Rpn4 is required for mitigating the inhibition of ergosterol biosynthesis by fluconazole. Based on our findings, we propose that Rpn4 is a central hub for fluconazole tolerance in C. albicans by coupling the regulation of protein homeostasis (proteostasis) and lipid metabolism to overcome drug-induced proteotoxicity and membrane stress.

CRISPR screens identify gene targets at breast cancer risk loci.

BACKGROUND: Genome-wide association studies (GWAS) have identified > 200 loci associated with breast cancer risk. The majority of candidate causal variants are in non-coding regions and likely modulate cancer risk by regulating gene expression. However, pinpointing the exact target of the association, and identifying the phenotype it mediates, is a major challenge in the interpretation and translation of GWAS. RESULTS: Here, we show that pooled CRISPR screens are highly effective at identifying GWAS target genes and defining the cancer phenotypes they mediate. Following CRISPR mediated gene activation or suppression, we measure proliferation in 2D, 3D, and in immune-deficient mice, as well as the effect on DNA repair. We perform 60 CRISPR screens and identify 20 genes predicted with high confidence to be GWAS targets that promote cancer by driving proliferation or modulating the DNA damage response in breast cells. We validate the regulation of a subset of these genes by breast cancer risk variants. CONCLUSIONS: We demonstrate that phenotypic CRISPR screens can accurately pinpoint the gene target of a risk locus. In addition to defining gene targets of risk loci associated with increased breast cancer risk, we provide a platform for identifying gene targets and phenotypes mediated by risk variants.

Infection induces tissue-resident memory NK cells that safeguard tissue health.

Tissue health is dictated by the capacity to respond to perturbations and then return to homeostasis. Mechanisms that initiate, maintain, and regulate immune responses in tissues are therefore essential. Adaptive immunity plays a key role in these responses, with memory and tissue residency being cardinal features. A corresponding role for innate cells is unknown. Here, we have identified a population of innate lymphocytes that we term tissue-resident memory-like natural killer (NKRM) cells. In response to murine cytomegalovirus infection, we show that circulating NK cells were recruited in a CX3CR1-dependent manner to the salivary glands where they formed NKRM cells, a long-lived, tissue-resident population that prevented autoimmunity via TRAIL-dependent elimination of CD4+ T cells. Thus, NK cells develop adaptive-like features, including long-term residency in non-lymphoid tissues, to modulate inflammation, restore immune equilibrium, and preserve tissue health. Modulating the functions of NKRM cells may provide additional strategies to treat inflammatory and autoimmune diseases.

Efficacy and safety of sotagliflozin in patients with type 2 diabetes and stage 3 chronic kidney disease.

AIM: To assess the efficacy and safety of sotagliflozin, a dual inhibitor of sodium-glucose co-transporters 1 and 2, in adults with type 2 diabetes (T2D) and stage 3 chronic kidney disease (CKD3). MATERIALS AND METHODS: This phase 3, randomized, placebo-controlled trial evaluated sotagliflozin 200 and 400 mg in 787 patients with T2D and an estimated glomerular filtration rate of 30-59 ml/min/1.73m2 . The primary objective was superiority of week 26 HbA1c reductions with sotagliflozin versus placebo. Secondary endpoints included changes in other glycaemic and renal endpoints overall and in CKD3 subgroups. RESULTS: At 26 weeks, the placebo-adjusted mean change in HbA1c (from a baseline of 8.3% ± 1.0%) was -0.1% (95% CI: -0.2% to 0.05%; P = .2095) and -0.2% (-0.4% to -0.09%; P = .0021) in the sotagliflozin 200 and 400 mg groups, respectively. Significant reductions in fasting plasma glucose and body weight, but not systolic blood pressure, were observed. Among patients with at least A2 albuminuria at week 26, the urine albumin-creatinine ratio (UACR) was reduced with both sotagliflozin doses relative to placebo. At week 52, UACR was reduced with sotagliflozin 200 mg in the CKD3B group. Adverse events (AEs), including serious AEs, were similar between the treatment groups. CONCLUSIONS: After 26 weeks, HbA1c was significantly reduced with sotagliflozin 400 but not 200 mg compared with placebo in this CKD3 cohort. UACR in patients with at least A2 albuminuria was reduced with each of the two doses at 26 weeks, but changes were not sustained at week 52. The safety findings were consistent with previous reports (NCT03242252).

Development of a Head Acceleration Event Classification Algorithm for Female Rugby Union.

Instrumented mouthguards have been used to detect head accelerations and record kinematic data in numerous sports. Each recording requires validation through time-consuming video verification. Classification algorithms have been posed to automatically categorise head acceleration events and spurious events. However, classification algorithms must be designed and/or validated for each combination of sport, sex and mouthguard system. This study provides the first algorithm to classify head acceleration data from exclusively female rugby union players. Mouthguards instrumented with kinematic sensors were given to 25 participants for six competitive rugby union matches in an inter-university league. Across all instrumented players, 214 impacts were recorded from 460 match-minutes. Matches were video recorded to enable retrospective labelling of genuine and spurious events. Four machine learning algorithms were trained on five matches to predict these labels, then tested on the sixth match. Of the four classifiers, the support vector machine achieved the best results, with area under the receiver operator curve (AUROC) and area under the precision recall curve (AUPRC) scores of 0.92 and 0.85 respectively, on the test data. These findings represent an important development for head impact telemetry in female sport, contributing to the safer participation and improving the reliability of head impact data collection within female contact sport.

Skeletal phenotypes in secreted frizzled-related protein 4 gene knockout mice mimic skeletal architectural abnormalities in subjects with Pyle's disease from SFRP4 mutations.

Mutations in SFRP4 cause Pyle's bone disease with wide metaphyses and increased skeletal fragility. The WNT signaling pathway plays important roles in determining skeletal architecture and SFRP4 is a secreted Frizzled decoy receptor that inhibits WNT signaling. Seven cohorts of male and female Sfrp4 gene knockout mice, examined through 2 years of age, had a normal lifespan but showed cortical and trabecular bone phenotypes. Mimicking human Erlenmeyer flask deformities, bone cross-sectional areas were elevated 2-fold in the distal femur and proximal tibia but only 30% in femur and tibia shafts. Reduced cortical bone thickness was observed in the vertebral body, midshaft femur and distal tibia. Elevated trabecular bone mass and numbers were observed in the vertebral body, distal femur metaphysis and proximal tibia metaphysis. Midshaft femurs retained extensive trabecular bone through 2 years of age. Vertebral bodies had increased compressive strength, but femur shafts had reduced bending strength. Trabecular, but not cortical, bone parameters in heterozygous Sfrp4 mice were modestly affected. Ovariectomy resulted in similar declines in both cortical and trabecular bone mass in wild-type and Sfrp4 KO mice. SFRP4 is critical for metaphyseal bone modeling involved in determining bone width. Sfrp4 KO mice show similar skeletal architecture and bone fragility deficits observed in patients with Pyle's disease with SFRP4 mutations.

Contrasting effects of Ksr2, an obesity gene, on trabecular bone volume and bone marrow adiposity.

Pathological obesity and its complications are associated with an increased propensity for bone fractures. Humans with certain genetic polymorphisms at the kinase suppressor of ras2 (KSR2) locus develop severe early-onset obesity and type 2 diabetes. Both conditions are phenocopied in mice with Ksr2 deleted, but whether this affects bone health remains unknown. Here we studied the bones of global Ksr2 null mice and found that Ksr2 negatively regulates femoral, but not vertebral, bone mass in two genetic backgrounds, while the paralogous gene, Ksr1, was dispensable for bone homeostasis. Mechanistically, KSR2 regulates bone formation by influencing adipocyte differentiation at the expense of osteoblasts in the bone marrow. Compared with Ksr2's known role as a regulator of feeding by its function in the hypothalamus, pair-feeding and osteoblast-specific conditional deletion of Ksr2 reveals that Ksr2 can regulate bone formation autonomously. Despite the gains in appendicular bone mass observed in the absence of Ksr2, bone strength, as well as fracture healing response, remains compromised in these mice. This study highlights the interrelationship between adiposity and bone health and provides mechanistic insights into how Ksr2, an adiposity and diabetic gene, regulates bone metabolism.

Our bones are living tissues which constantly reshape and renew themselves. This ability relies on stem cells present in the marrow cavity, which can mature into the various types of cells needed to produce new bone material, marrow fat, or other components. Obesity and associated conditions such as type 2 diabetes are often linked to harmful changes in the skeleton. In particular, these metabolic conditions are associated with weight-bearing bones becoming more prone to facture and healing poorly. Mice genetically modified to model obesity and diabetes could help researchers to study exactly how these conditions ­ and the genetic changes that underlie them ­ impact bone health. Gomez et al. aimed to address this question by focusing on KSR2, a gene involved in energy consumption and feeding behavior. Children who carry certain KSR2 mutations are prone to obesity and type 2 diabetes; mice lacking the gene also develop these conditions due to uncontrolled eating. Closely examining mutant mice in which Ksr2 had been deactivated in every cell revealed that the weight-bearing bones of these animals were also more likely to break, and the fractures then healed more slowly. This was the case even though these bones had higher mass and less marrow fat compared to healthy mice. Non-weight bearing bones (such as the spine) did not exhibit these changes. Further experiments revealed that, when expressed normally in the skeleton, Ksr2 skews the stem cell maturation process towards marrow fat cells instead of bone-creating cells. This suggests a new role for Ksr2, which therefore seems to independently regulate both feeding behavior and bone health. In addition, the work by Gomez et al. demonstrate that Ksr2 mutant mice could be a useful model to better understand how obesity and diabetes affect human bones, and to potentially develop new therapies.

Metabolic, Intestinal, and Cardiovascular Effects of Sotagliflozin Compared With Empagliflozin in Patients With Type 2 Diabetes: A Randomized, Double-Blind Study.

OBJECTIVE: Inhibiting sodium-glucose cotransporters (SGLTs) improves glycemic and cardiovascular outcomes in patients with type 2 diabetes (T2D). We investigated the differential impact of selective SGLT2 inhibition and dual inhibition of SGLT1 and SGLT2 on multiple parameters. RESEARCH DESIGN AND METHODS: Using a double-blind, parallel-group design, we randomized 40 patients with T2D and hypertension to receive the dual SGLT1 and SGLT2 inhibitor sotagliflozin 400 mg or the selective SGLT2 inhibitor empagliflozin 25 mg, with preexisting antihypertensive treatment, for 8 weeks. In an in-house testing site, mixed-meal tolerance tests (MMTTs) and other laboratory and clinical evaluations were used to study metabolic, intestinal, cardiovascular, and urinary parameters over 24 h. RESULTS: Changes from baseline in glycemic and blood pressure control; intestinal, urine, and metabolic parameters; and cardiovascular biomarkers were generally similar with sotagliflozin and empagliflozin. During the breakfast MMTT, sotagliflozin significantly reduced incremental area under the curve (AUC) values for postprandial glucose, insulin, and glucose-dependent insulinotropic polypeptide (GIP) and significantly increased incremental AUCs for postprandial glucagon-like peptide 1 (GLP-1) relative to empagliflozin, consistent with sotagliflozin-mediated inhibition of intestinal SGLT1. These changes waned during lunch and dinner MMTTs. Both treatments significantly lowered GIP incremental AUCs relative to baseline over the 14 h MMTT interval; the most vigorous effect was seen with sotagliflozin soon after start of the first meal of the day. No serious or severe adverse events were observed. CONCLUSIONS: Changes from baseline in glycemic and blood pressure control, cardiovascular biomarkers, and other parameters were comparable between sotagliflozin and empagliflozin. However, sotagliflozin but not empagliflozin inhibited intestinal SGLT1 after breakfast as shown by larger changes in postprandial glucose, insulin, GIP, and GLP-1 AUCs, particularly after breakfast. Additional study is warranted to assess the clinical relevance of transient SGLT1 inhibition and differences in incretin responses (NCT03462069).

Transcriptomic analysis of preovipositional embryonic arrest in a nonsquamate reptile (Chelonia mydas).

After gastrulation, oviductal hypoxia maintains turtle embryos in an arrested state prior to oviposition. Subsequent exposure to atmospheric oxygen upon oviposition initiates recommencement of embryonic development. Arrest can be artificially extended for several days after oviposition by incubation of the egg under hypoxic conditions, with development recommencing in an apparently normal fashion after subsequent exposure to normoxia. To examine the transcriptomic events associated with embryonic arrest in green sea turtles (Chelonia mydas), RNA-sequencing analysis was performed on embryos from freshly laid eggs and eggs incubated in either normoxia (oxygen tension ~159 mmHg) or hypoxia (<8 mmHg) for 36 h after oviposition (n = 5 per group). The patterns of gene expression differed markedly among the three experimental groups. Normal embryonic development in normoxia was associated with upregulation of genes involved in DNA replication, the cell cycle, and mitosis, but these genes were commonly downregulated after incubation in hypoxia. Many target genes of hypoxia inducible factors, including the gene encoding insulin-like growth factor binding protein 1 (igfbp1), were downregulated by normoxic incubation but upregulated by incubation in hypoxia. Notably, some of the transcriptomic effects of hypoxia in green turtle embryos resembled those reported to be associated with hypoxia-induced embryonic arrest in diverse taxa, including the nematode Caenorhabditis elegans and zebrafish (Danio rerio). Hypoxia-induced preovipositional embryonic arrest appears to be a unique adaptation of turtles. However, our findings accord with the proposition that the mechanisms underlying hypoxia-induced embryonic arrest per se are highly conserved across diverse taxa.

The Role and Targets of the RNA-Binding Protein ProQ in the Gram-Negative Bacterial Pathogen Pasteurella multocida.

The Gram-negative pathogen Pasteurella multocida is the causative agent of many important animal diseases. While a number of P. multocida virulence factors have been identified, very little is known about how gene expression and protein production is regulated in this organism. One mechanism by which bacteria regulate transcript abundance and protein production is riboregulation, which involves the interaction of a small RNA (sRNA) with a target mRNA to alter transcript stability and/or translational efficiency. This interaction often requires stabilization by an RNA-binding protein such as ProQ or Hfq. In Escherichia coli and a small number of other species, ProQ has been shown to play a critical role in stabilizing sRNA-mRNA interactions and preferentially binds to the 3' stem-loop regions of the mRNA transcripts, characteristic of intrinsic transcriptional terminators. The aim of this study was to determine the role of ProQ in regulating P. multocida transcript abundance and identify the RNA targets to which it binds. We assessed differentially expressed transcripts in a proQ mutant and identified sites of direct ProQ-RNA interaction using in vivo UV-cross-linking and analysis of cDNA (CRAC). These analyses demonstrated that ProQ binds to, and stabilizes, ProQ-dependent sRNAs and transfer RNAs in P. multocida via adenosine-enriched, highly structured sequences. The binding of ProQ to two RNA molecules was characterized, and these analyses showed that ProQ bound within the coding sequence of the transcript PmVP161_1121, encoding an uncharacterized protein, and within the 3' region of the putative sRNA Prrc13. IMPORTANCE Regulation in P. multocida involving the RNA-binding protein Hfq is required for hyaluronic acid capsule production and virulence. This study further expands our understanding of riboregulation by examining the role of a second RNA-binding protein, ProQ, in transcript regulation and abundance in P. multocida.

Mice Lacking Gpr75 are Hypophagic and Thin.

PURPOSE: Humans with haploinsufficiency of GPR75, an orphan GPCR, are thin. Gpr75 knockout (KO) mice are also thin with improved glucose homeostasis. We wanted to confirm these findings in Gpr75 KO mice and determine whether decreased energy intake and/or increased energy expenditure contributed to the thin phenotype. METHODS: Gpr75 KO mice were generated by homologous recombination. All studies compared female and male Gpr75 KO mice to their wild type (WT) littermates. Body composition was measured by DXA and QMR technologies. Glucose homeostasis was evaluated by measuring glucose and insulin levels during oral glucose tolerance tests (OGTTs). Food intake was measured in group-housed mice. In singly housed mice, energy expenditure was measured in Oxymax indirect calorimetry chambers, and locomotor activity was measured in Oxymax and Photobeam Activity System chambers. RESULTS: In all 12 cohorts of adult female or male mice, Gpr75 KO mice had less body fat; pooled data showed that, compared to WT littermates (n = 103), Gpr75 KO mice (n = 118) had 49% less body fat and 4% less LBM (P < 0.001 for each). KO mice also had 8% less body fat at weaning (P < 0.05), and during the month after weaning as the thin phenotype became more exaggerated, Gpr75 KO mice ate significantly less than, but had energy expenditure and activity levels comparable to, their WT littermates. During OGTTs, Gpr75 KO mice showed improved glucose tolerance (glucose AUC 23% lower in females, P < 0.05, and 26% lower in males, P < 0.001), accompanied by significantly decreased insulin levels and significantly increased insulin sensitivity indices. CONCLUSION: Gpr75 KO mice are thin at weaning, are hypophagic as the thin phenotype becomes more exaggerated, and exhibit improved glucose tolerance and insulin sensitivity as healthy-appearing adults. These results suggest that inhibiting GPR75 in obese humans may safely decrease energy intake and body fat while improving glucose tolerance and insulin sensitivity.

3D-cardiomics: A spatial transcriptional atlas of the mammalian heart.

Understanding the spatial gene expression and regulation in the heart is key to uncovering its developmental and physiological processes, during homeostasis and disease. Numerous techniques exist to gain gene expression and regulation information in organs such as the heart, but few utilize intuitive true-to-life three-dimensional representations to analyze and visualise results. Here we combined transcriptomics with 3D-modelling to interrogate spatial gene expression in the mammalian heart. For this, we microdissected and sequenced transcriptome-wide 18 anatomical sections of the adult mouse heart. Our study has unveiled known and novel genes that display complex spatial expression in the heart sub-compartments. We have also created 3D-cardiomics, an interface for spatial transcriptome analysis and visualization that allows the easy exploration of these data in a 3D model of the heart. 3D-cardiomics is accessible from http://3d-cardiomics.erc.monash.edu/.

Sex differences in neck strength and head impact kinematics in university rugby union players.

Globally, over three million women participate in rugby union, yet injury prevention and training strategies are predominantly based on androcentric data. These strategies may have limited generalisability to females, given the cervical spine is more susceptible to whiplash and less adept at resisting inertial loading. A total of 53 university rugby union players (25 female, 28 male, 20.7 ± 1.8 years) had their isometric neck strength measured. Bespoke instrumented mouthguards were used to record the magnitude of head impact events in six female and seven male competitive matches. Mean female maximal isometric neck strength was 47% lower than male. Independent samples Mann-Whitney U tests showed no significant differences for peak linear head acceleration (female: median 11.7 g, IQR 7.9 g; male: median 12.5 g, IQR 7.0 g p=.23) or peak rotational head acceleration (female: median 800.2 rad·s-2, IQR 677.7 rad·s-2; male: median 849.4 rad·s-2, IQR 479.8 rad·s-2; p=.76), despite the mean male body mass being 24% greater than female. Coded video analysis revealed substantial differences in head-impact mechanisms; uncontrolled whiplash dominated >50% of all recorded female impact events and <0.5% in males. Direct head-to-ground impacts comprised 26.1% of female and 9.7% of male impacts, with whiplash occurring in 78.0% and 0.5%, respectively. Overall, the data provided in this study do not support the generalisation of male-derived training and injury-prevention data to female rugby athletes. These results suggest a considerable research effort is required to identify specific weakness of female rugby players and derive appropriate training, injury prevention and return to play protocols.Highlights Video analysis revealed substantial differences in head-impact mechanisms, with uncontrolled whiplash dominating >50% of all recorded female impact events but rarely in males.Isometric neck strength was 47% lower in female players than males.Direct head-to-ground impacts accounted for 26.1% and 9.7% of female and male impacts, with whiplash occurring in 78.0% and 0.5%, respectively.

PI(3,4)P2-mediated cytokinetic abscission prevents early senescence and cataract formation.

Cytokinetic membrane abscission is a spatially and temporally regulated process that requires ESCRT (endosomal sorting complexes required for transport)­dependent control of membrane remodeling at the midbody, a subcellular organelle that defines the cleavage site. Alteration of ESCRT function can lead to cataract, but the underlying mechanism and its relation to cytokinesis are unclear. We found a lens-specific cytokinetic process that required PI3K-C2α (phosphatidylinositol-4-phosphate 3-kinase catalytic subunit type 2α), its lipid product PI(3,4)P2 (phosphatidylinositol 3,4-bisphosphate), and the PI(3,4)P2­binding ESCRT-II subunit VPS36 (vacuolar protein-sorting-associated protein 36). Loss of each of these components led to impaired cytokinesis, triggering premature senescence in the lens of fish, mice, and humans. Thus, an evolutionarily conserved pathway underlies the cell type­specific control of cytokinesis that helps to prevent early onset cataract by protecting from senescence.

High-Throughput Screening of Mouse Gene Knockouts Identifies Established and Novel High Body Fat Phenotypes.

PURPOSE: Obesity is a major public health problem. Understanding which genes contribute to obesity may better predict individual risk and allow development of new therapies. Because obesity of a mouse gene knockout (KO) line predicts an association of the orthologous human gene with obesity, we reviewed data from the Lexicon Genome5000TM high throughput phenotypic screen (HTS) of mouse gene KOs to identify KO lines with high body fat. MATERIALS AND METHODS: KO lines were generated using homologous recombination or gene trapping technologies. HTS body composition analyses were performed on adult wild-type and homozygous KO littermate mice from 3758 druggable mouse genes having a human ortholog. Body composition was measured by either DXA or QMR on chow-fed cohorts from all 3758 KO lines and was measured by QMR on independent high fat diet-fed cohorts from 2488 of these KO lines. Where possible, comparisons were made to HTS data from the International Mouse Phenotyping Consortium (IMPC). RESULTS: Body fat data are presented for 75 KO lines. Of 46 KO lines where independent external published and/or IMPC KO lines are reported as obese, 43 had increased body fat. For the remaining 29 novel high body fat KO lines, Ksr2 and G2e3 are supported by data from additional independent KO cohorts, 6 (Asnsd1, Srpk2, Dpp8, Cxxc4, Tenm3 and Kiss1) are supported by data from additional internal cohorts, and the remaining 21 including Tle4, Ak5, Ntm, Tusc3, Ankk1, Mfap3l, Prok2 and Prokr2 were studied with HTS cohorts only. CONCLUSION: These data support the finding of high body fat in 43 independent external published and/or IMPC KO lines. A novel obese phenotype was identified in 29 additional KO lines, with 27 still lacking the external confirmation now provided for Ksr2 and G2e3 KO mice. Undoubtedly, many mammalian obesity genes remain to be identified and characterized.