Explore the vast range of titles available.

Epithelial ovarian cancer (EOC) is a deadly cancer, and its prognosis has not been changed significantly during several decades. To seek new therapeutic targets for EOC, we performed an in vivo dropout screen in human tumor xenografts using a pooled shRNA library targeting thousands of druggable genes. Then, in follow-up studies, we performed a second screen using a genome-wide CRISPR/Cas9 library. These screens identified 10 high-confidence drug targets that included well-known oncogenes such as ERBB2 and RAF1, and novel oncogenes, notably KPNB1, which we investigated further. Genetic and pharmacological inhibition showed that KPNB1 exerts its antitumor effects through multiphase cell cycle arrest and apoptosis induction. Mechanistically, proteomic studies revealed that KPNB1 acts as a master regulator of cell cycle-related proteins, including p21, p27, and APC/C. Clinically, EOC patients with higher expression levels of KPNB1 showed earlier recurrence and worse prognosis than those with lower expression levels of KPNB1. Interestingly, ivermectin, a Food and Drug Administration-approved antiparasitic drug, showed KPNB1-dependent antitumor effects on EOC, serving as an alternative therapeutic toward EOC patients through drug repositioning. Last, we found that the combination of ivermectin and paclitaxel produces a stronger antitumor effect on EOC both in vitro and in vivo than either drug alone. Our studies have thus identified a combinatorial therapy for EOC, in addition to a plethora of potential drug targets.

N-methyl-D-aspartate receptors (NMDARs emerging from GRIN genes) are tetrameric receptors that form diverse channel compositions in neurons, typically consisting of two GluN1 subunits combined with two GluN2(A-D) subunits. During prenatal stages, the predominant channels are di-heteromers with two GluN1 and two GluN2B subunits due to the high abundance of GluN2B subunits. Postnatally, the expression of GluN2A subunits increases, giving rise to additional subtypes, including GluN2A-containing di-heteromers and tri-heteromers with GluN1, GluN2A, and GluN2B subunits. The latter  emerge as the major receptor subtype at mature synapses in the hippocampus. Despite extensive research on purely di-heteromeric receptors containing two identical GRIN variants, the impact of a single variant on the function of other channel forms, notably tri-heteromers, is lagging. In this study, we systematically investigated the effects of two de novo GRIN2B variants (G689C and G689S) in pure, mixed di- and tri-heteromers. Our findings reveal that incorporating a single variant in mixed di-heteromers or tri-heteromers exerts a dominant negative effect on glutamate potency, although ‘mixed’ channels show improved potency compared to pure variant-containing di-heteromers. We show that a single variant within a receptor complex does not impair the response of all receptor subtypes to the positive allosteric modulator pregnenolone-sulfate (PS), whereas spermine completely fails to potentiate tri-heteromers containing GluN2A and -2B-subunits. We examined PS on primary cultured hippocampal neurons transfected with the variants, and observed a positive impact over current amplitudes and synaptic activity. Together, our study supports previous observations showing that mixed di-heteromers exhibit improved glutamate potency and extend these findings towards the exploration of the effect of Loss-of-Function variants over tri-heteromers. Notably, we provide an initial and crucial demonstration of the beneficial effects of GRIN2B -relevant potentiators on tri-heteromers. Our results underscore the significance of studying how different variants affect distinct receptor subtypes, as these effects cannot be inferred solely from observations made on pure di-heteromers. Overall, this study contributes to ongoing efforts to understand the pathophysiology of GRINopathies and provides insights into potential treatment strategies.

CRISPR-Cas9 gene editing has transformed our ability to rapidly interrogate the functional impact of somatic mutations in human cancers. Droplet-based technology enables the analysis of Cas9-introduced gene edits in thousands of single cells. Using this technology, we analyze Ba/F3 cells engineered to express single or multiplexed loss-of-function mutations recurrent in chronic lymphocytic leukemia. Our approach reliably quantifies mutational co-occurrences, zygosity status, and the occurrence of Cas9 edits at single-cell resolution.

Oligo‐astheno‐teratozoospermia (OAT) is a common cause of male infertility, but the genetic basis of most OAT cases is still unknown. Here, one homozygous loss‐of‐function (LOF) variant in TDRD6, c.G1825T/p.Gly609X, was identified in an infertile patient with severe OAT by whole‐exome sequencing (WES) and Sanger confirmation. Furthermore, Tdrd6‐mutant mice (p.Gly615X; equivalent to p.Gly609X in human TDRD6) were generated. Remarkably, the Tdrd6‐mutated mice mimicked the severe OAT symptoms of the patient. In addition, the architecture of chromatoid bodies (CBs) were disrupted in round spermatids from Tdrd6‐mutant mice, leading to blocked spermatogenesis in the round spermatids. The assembly of PIWIL1, TDRD1, TDRD7 and DDX25 in CBs was disturbed in the Tdrd6‐mutant mice. Applying immunoprecipitation‐mass spectrometry (IP‐MS), we identified some TDRD6‐interacting partners, including CB proteins TDRD7, MAEL and PCBP1. Moreover, we described the assisted reproductive technology (ART) outcomes of the infertile patient and his partner. Altogether, our findings provide necessary evidences to support the idea that the homozygous LOF variant in TDRD6 induces male infertility with severe OAT, suggesting that TDRD6 could be a useful genetic diagnostic target for male infertility.

The JAK/STAT signaling pathway plays a key role in cytokine signaling and is involved in development, immunity, and tumorigenesis for nearly any cell. At first glance, the JAK/STAT signaling pathway appears to be straightforward. However, on closer examination, the factors influencing the JAK/STAT signaling activity, such as cytokine diversity, receptor profile, overlapping JAK and STAT specificity among non-redundant functions of the JAK/STAT complexes, positive regulators (e.g., cooperating transcription factors), and negative regulators (e.g., SOCS, PIAS, PTP), demonstrate the complexity of the pathway’s architecture, which can be quickly disturbed by mutations. The JAK/STAT signaling pathway has been, and still is, subject of basic research and offers an enormous potential for the development of new methods of personalized medicine and thus the translation of basic molecular research into clinical practice beyond the use of JAK inhibitors. Gain-of-function and loss-of-function mutations in the three immunologically particularly relevant signal transducers STAT1, STAT3, and STAT6 as well as JAK1 and JAK3 present themselves through individual phenotypic clinical pictures. The established, traditional paradigm of loss-of-function mutations leading to immunodeficiency and gain-of-function mutation leading to autoimmunity breaks down and a more differentiated picture of disease patterns evolve. This review is intended to provide an overview of these specific syndromes from a clinical perspective and to summarize current findings on pathomechanism, symptoms, immunological features, and therapeutic options of STAT1, STAT3, STAT6, JAK1, and JAK3 loss-of-function and gain-of-function diseases.

Under dehydration in plants, antagonistic activities of histone 3 lysine 4 (H3K4) methyl-transferase and histone demethylase maintain a dynamic and homeostatic state of gene expression by orientating transcriptional reprogramming toward growth or stress tolerance. However, the histone demethylase that specifically controls histone methylation homeostasis under dehydration stress remains unknown. Here, we document that a histone demethylase, JMJ17, belonging to the KDM5/JARID1 family, plays crucial roles in response to dehydration stress and abscisic acid (ABA) in Arabidopsis thaliana. jmj17 loss-of-function mutants displayed dehydration stress tolerance and ABA hypersensitivity in terms of stomatal closure. JMJ17 specifically demethylated H3K4me1/2/3 via conserved iron-binding amino acids in vitro and in vivo. Moreover, H3K4 demethylase activity of JMJ17 was required for dehydration stress response. Systematic combination of genome-wide chromatin immunoprecipitation coupled with massively parallel DNA sequencing (ChIP-seq) and RNA-sequencing (RNA-seq) analyses revealed that a loss-of-function mutation in JMJ17 caused an ectopic increase in genome-wide H3K4me3 levels and activated a plethora of dehydration stress-responsive genes. Importantly, JMJ17 bound directly to the chromatin of OPEN STOMATA 1 (OST1) and demethylated H3K4me3 for the regulation of OST1 mRNA abundance, thereby modulating the dehydration stress response. Our results demonstrate a new function of a histone demethylase under dehydration stress in plants.

AMPA receptors (AMPARs) are tetrameric ligand-gated channels made up of combinations of GluA1-4 subunits encoded by GRIA1-4 genes. GluA2 has an especially important role because, following post-transcriptional editing at the Q607 site, it renders heteromultimeric AMPARs Ca 2+ -impermeable, with a linear relationship between current and trans-membrane voltage. Here, we report heterozygous de novo GRIA2 mutations in 28 unrelated patients with intellectual disability (ID) and neurodevelopmental abnormalities including autism spectrum disorder (ASD), Rett syndrome-like features, and seizures or developmental epileptic encephalopathy (DEE). In functional expression studies, mutations lead to a decrease in agonist-evoked current mediated by mutant subunits compared to wild-type channels. When GluA2 subunits are co-expressed with GluA1, most GRIA2 mutations cause a decreased current amplitude and some also affect voltage rectification. Our results show that de-novo variants in GRIA2 can cause neurodevelopmental disorders, complementing evidence that other genetic causes of ID, ASD and DEE also disrupt glutamatergic synaptic transmission. Genetic variants in ionotropic glutamate receptors have been implicated in neurodevelopmental disorders. Here, the authors report heterozygous de novo mutations in the GRIA2 gene in 28 individuals with intellectual disability and neurodevelopmental abnormalities associated with reduced Ca 2+ transport and AMPAR currents.”

Key messageThe WRKY transcription factor WRKY12 negatively regulates Cd tolerance in Arabidopsis via the glutathione-dependent phytochelatin synthesis pathway by directly targeting GSH1 and indirectly repressing phytochelatin synthesis-related gene expression.Cadmium (Cd) is a widespread pollutant toxic to plants. The glutathione (GSH)-dependent phytochelatin (PC) synthesis pathway plays key roles in Cd detoxification. However, its regulatory mechanism remains largely unknown. Here, we showed a previously unknown function of the WRKY transcription factor WRKY12 in the regulation of Cd tolerance by repressing the expression of PC synthesis-related genes. The expression of WRKY12 was inhibited by Cd stress. Enhanced Cd tolerance was observed in the WRKY12 loss-of-function mutants, whereas increased Cd sensitivity was found in the WRKY12-overexpressing plants. Overexpression and loss-of-function of WRKY12 were associated respectively with increased and decreased Cd accumulation by repressing or releasing the expression of the genes involved in the PC synthesis pathway. Transient expression assay showed that WRKY12 repressed the expression of GSH1, GSH2, PCS1, and PCS2. Further analysis indicated that WRKY12 could directly bind to the W-box of the promoter in GSH1 but not in GSH2, PCS1, and PCS2 in vivo. Together, our results suggest that WRKY12 directly targets GSH1 and indirectly represses PC synthesis-related gene expression to negatively regulate Cd accumulation and tolerance in Arabidopsis.

Male infertility is a major reproductive disorder, which is clinically characterized by highly heterogeneous phenotypes of abnormal sperm count or quality. To date, five male patients with biallelic loss-of-function (LOF) variants of PARN-like ribonuclease domain-containing exonuclease 1 (PNLDC1) have been reported to experience infertility with nonobstructive azoospermia. The aim of this study was to identify the genetic cause of male infertility with oligo-astheno-teratozoospermia (OAT) in a patient from a Chinese Han family. Whole-exome and Sanger sequencing analyses identified a homozygous LOF variant (NM_173516.2, c.142C>T, p.Gln48Ter) in PNLDC1. Hematoxylin and eosin staining revealed that the spermatozoa of the patient with OAT had an irregular head phenotype, including microcephaly, head tapering, and globozoospermia. Consistently, peanut agglutinin staining of the spermatozoa revealed a complete or partial loss of the acrosome. Furthermore, the disomy rate of chromosomes in the patient's spermatozoa was significantly increased compared with that of a fertile control sample. We reported an LOF variant of the PNLDC1 gene responsible for OAT.

Causal loss-of-function (LOF) variants for Mendelian and severe complex diseases are enriched in 'mutation intolerant' genes. We show how such observations can be interpreted in light of a model of mutation-selection balance and use the model to relate the pathogenic consequences of LOF mutations at present to their evolutionary fitness effects. To this end, we first infer posterior distributions for the fitness costs of LOF mutations in 17,318 autosomal and 679 X-linked genes from exome sequences in 56,855 individuals. Estimated fitness costs for the loss of a gene copy are typically above 1%; they tend to be largest for X-linked genes, whether or not they have a Y homolog, followed by autosomal genes and genes in the pseudoautosomal region. We compare inferred fitness effects for all possible de novo LOF mutations to those of de novo mutations identified in individuals diagnosed with one of six severe, complex diseases or developmental disorders. Probands carry an excess of mutations with estimated fitness effects above 10%; as we show by simulation, when sampled in the population, such highly deleterious mutations are typically only a couple of generations old. Moreover, the proportion of highly deleterious mutations carried by probands reflects the typical age of onset of the disease. The study design also has a discernible influence: a greater proportion of highly deleterious mutations is detected in pedigree than case-control studies, and for autism, in simplex than multiplex families and in female versus male probands. Thus, anchoring observations in human genetics to a population genetic model allows us to learn about the fitness effects of mutations identified by different mapping strategies and for different traits.