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α-Neurexins are essential and highly expressed presynaptic cell-adhesion molecules that are frequently linked to neuropsychiatric and neurodevelopmental disorders. Despite their importance, how the elaborate extracellular sequences of α-neurexins contribute to synapse function is poorly understood. We recently characterized the presynaptic gain-of-function phenotype caused by a missense mutation in an evolutionarily conserved extracellular sequence of neurexin-3α (A687T) that we identified in a patient diagnosed with profound intellectual disability and epilepsy. The striking A687T gain-of-function mutation on neurexin-3α prompted us to systematically test using mutants whether the presynaptic gain-of-function phenotype is a consequence of the addition of side-chain bulk (i.e., A687V) or polar/hydrophilic properties (i.e., A687S). We used multidisciplinary approaches in mixed-sex primary hippocampal cultures to assess the impact of the neurexin-3αA687 residue on synapse morphology, function and ligand binding. Unexpectedly, neither A687V nor A687S recapitulated the neurexin-3α A687T phenotype. Instead, distinct from A687T, molecular replacement with A687S significantly enhanced postsynaptic properties exclusively at excitatory synapses and selectively increased binding to neuroligin-1 and neuroligin-3 without changing binding to neuroligin-2 or LRRTM2. Importantly, we provide the first experimental evidence supporting the notion that the position A687 of neurexin-3α and the N-terminal sequences of neuroligins may contribute to the stability of α-neurexin--neuroligin-1 trans-synaptic interactions and that these interactions may specifically regulate the postsynaptic strength of excitatory synapses. [ABSTRACT FROM AUTHOR]

Background and Purpose Experimental Approach Key Results Conclusions and Implications Pathogenic gain‐of‐function mutations in Cav1.3 L‐type voltage‐gated Ca2+‐channels (CACNA1D) cause neurodevelopmental disorders with or without endocrine symptoms. We aimed to confirm a pathogenic gain‐of function phenotype of CACNA1D de novo missense mutations A749T and L271H, and investigated the molecular mechanism causing their enhanced sensitivity for the Ca2+‐channel blocker isradipine, a potential therapeutic for affected patients.Wildtype and mutant channels were expressed in tsA‐201 cells and their gating analysed using whole‐cell and single‐channel patch‐clamp recordings. The voltage‐dependence of isradipine action was quantified using protocols inducing variable fractions of inactivated channels. The molecular basis for altered channel gating in the mutants was investigated using in silico modelling and molecular dynamics simulations.Both mutations were confirmed pathogenic due to characteristic shifts of voltage‐dependent activation and inactivation towards negative potentials (~20 mV). At negative holding potentials both mutations showed significantly higher isradipine sensitivity compared to wildtype. The affinity for wildtype and mutant channels increased with channel inactivation as predicted by the modulated receptor hypothesis (30‐ to 40‐fold). The IC50 was indistinguishable for wildtype and mutants when >50% of channels were inactivated.Mutations A749T and L271H induce pathogenic gating changes. Like wildtype, isradipine inhibition is strongly voltage‐dependent. Our data explains their apparent higher drug sensitivity at a given negative voltage by the availability of more inactivated channels due to their more negative inactivation voltage range. Low nanomolar isradipine concentrations will only inhibit Cav1.3 channels in neurons during prolonged depolarized states without selectivity for mutant channels. [ABSTRACT FROM AUTHOR]

This document is an interview with Dr. Samuel Refetoff, a renowned thyroid researcher. The interview provides a brief background on Dr. Refetoff's upbringing in Bulgaria and his journey to becoming a medical professional. It also discusses his early work with a family in Los Angeles who had suspected thyroid problems, which eventually led to the discovery of resistance to thyroid hormone. The interview highlights the challenges faced by Dr. Refetoff in getting his ideas accepted and the assistance he received from other experts in the field. Dr. Refetoff describes his research on target tissue resistance to thyroid hormone and the discovery of mutations in the thyroid hormone receptor genes. He also discusses other areas of his research, including thyroxine-binding globulin deficiency and TSH resistance. Dr. Refetoff emphasizes the importance of supporting young researchers in the field of thyroidology and expresses concerns about the lack of funding for studies on congenital thyroid diseases. The conversation concludes with a discussion of Dr. Refetoff's personal life and family. [Extracted from the article]

Loss-of-function germline von Hippel-Lindau (VHL) tumor suppressor mutations cause VHL disease, which predisposes individuals to kidney cancer, hemangioblastomas, and paragangliomas. The risk that a given VHL disease family will manifest some or all these tumor types is profoundly influenced by the VHL allele it carries. For example, almost all VHL disease families that develop paraganglioma have missense VHL mutations. VHL families with null VHL alleles develop kidney cancer and hemangioblastomas without a high risk of paraganglioma. The latter is surprising because the VHL gene product, pVHL, suppresses the HIF2 transcription factor and gain-of-function HIF2 mutations are also linked to paraganglioma. Paragangliomas arise from the sympathetic or parasympathetic nervous system. Given the lack of human paraganglioma cell lines, we studied the effects of inactivating VHL in neuroblastoma cell lines, which also arise from the sympathetic nervous system. We found that total loss of pVHL function profoundly impairs the fitness of neuroblastoma cell lines in a HIF2-dependent manner both ex vivo and in vivo. This fitness defect can be rescued by pVHL variants linked to paraganglioma, but not by pVHL variants associated with a low risk of paraganglioma. These findings suggest that HIF2 activity above a critical threshold prevents the development of paraganglioma. [ABSTRACT FROM AUTHOR]

PIEZO2 is a trimeric mechanically-gated ion channel expressed by most sensory neurons in the dorsal root ganglia. Mechanosensitive PIEZO2 channels are also genetically required for normal touch sensation in both mice and humans. We previously showed that PIEZO2 channels are also strongly modulated by membrane voltage. Specifically, it is only at very positive voltages that all channels are available for opening by mechanical force. Conversely, most PIEZO2 channels are blocked at normal negative resting membrane potentials. The physiological function of this unusual biophysical property of PIEZO2 channels, however, remained unknown. We characterized the biophysical properties of three PIEZO2 ion channel mutations at an evolutionarily conserved arginine (R2756). Using genome engineering in mice we generated Piezo2 R2756H/R2756H and Piezo2 R2756K/R2756K knock-in mice to characterize the physiological consequences of altering PIEZO2 voltage sensitivity in vivo. We measured endogenous mechanosensitive currents in sensory neurons isolated from the dorsal root ganglia and characterized mechanoreceptor and nociceptor function using electrophysiology. Mice were also assessed behaviourally and morphologically. Mutations at the conserved Arginine (R2756) dramatically changed the biophysical properties of the channel relieving voltage block and lowering mechanical thresholds for channel activation. Piezo2R2756H/R2756H and Piezo2R2756K/R2756K knock-in mice that were homozygous for gain-of-function mutations were viable and were tested for sensory changes. Surprisingly, mechanosensitive currents in nociceptors, neurons that detect noxious mechanical stimuli, were substantially sensitized in Piezo2 knock-in mice, but mechanosensitive currents in most mechanoreceptors that underlie touch sensation were only mildly affected by the same mutations. Single-unit electrophysiological recordings from sensory neurons innervating the glabrous skin revealed that rapidly-adapting mechanoreceptors that innervate Meissner's corpuscles exhibited slightly decreased mechanical thresholds in Piezo2 knock-in mice. Consistent with measurements of mechanically activated currents in isolated sensory neurons essentially all cutaneous nociceptors, both fast conducting Aδ-mechanonociceptors and unmyelinated C-fibre nociceptors were substantially more sensitive to mechanical stimuli and indeed acquired receptor properties similar to ultrasensitive touch receptors in Piezo2 knock-in mice. Mechanical stimuli also induced enhanced ongoing activity in cutaneous nociceptors in Piezo2 knock-in mice and hyper-sensitive PIEZO2 channels were sufficient alone to drive ongoing activity, even in isolated nociceptive neurons. Consistently, Piezo2 knock-in mice showed substantial behavioural hypersensitivity to noxious mechanical stimuli. Our data indicate that ongoing activity and sensitization of nociceptors, phenomena commonly found in human chronic pain syndromes, can be driven by relieving the voltage-block of PIEZO2 ion channels. Indeed, membrane depolarization caused by multiple noxious stimuli may sensitize nociceptors by relieving voltage-block of PIEZO2 channels. [ABSTRACT FROM AUTHOR]

USP25 encodes ubiquitin-specific protease 25, a key member of the deubiquitinating enzyme family that is involved in neural fate determination. Although abnormal expression in Down's syndrome was reported previously, the specific role of USP25 in human diseases has not been defined. In this study, we performed trio-based whole exome sequencing in a cohort of 319 cases (families) with generalized epilepsy of unknown aetiology. Five heterozygous USP25 variants, including two de novo and three co-segregated variants, were determined in eight individuals affected by generalized seizures and/or febrile seizures from five unrelated families. The frequency of USP25 variants showed a significantly high aggregation in this cohort compared with the East Asian population and all populations in the gnomAD database. The mean age at onset of febrile and afebrile seizures were 10 months (infancy) and 11.8 years (juvenile), respectively. The patients achieved seizure freedom, except that one had occasional nocturnal seizures at the last follow-up. Two patients exhibited intellectual disability. Usp25 was expressed ubiquitously in mouse brain with two peaks, on embryonic Days 14–16 and postnatal Day 21, respectively. In human brain, likewise, USP25 is expressed in the fetus/early childhood stage and with a second peak at ∼12–20 years old, consistent with the seizure onset age in patients during infancy and in juveniles. To investigate the functional impact of USP25 deficiency in vivo , we established Usp25 knockout mice, which showed increased seizure susceptibility compared with wild-type mice in a pentylenetetrazol-induced seizure test. To explore the impact of USP25 variants, we used multiple functional detections. In HEK293 T cells, the variant associated with a severe phenotype (p.Gln889Ter) led to a significant reduction of mRNA and protein expressions but formed stable truncated dimers with an increment of deubiquitinating enzyme activities and abnormal cellular aggregations, indicating a gain-of-function effect. The p.Gln889Ter and p.Leu1045del variants increased neuronal excitability in mouse brain, with a higher firing ability in p.Gln889Ter. These functional impairments align with the severity of the observed phenotypes, suggesting a genotype–phenotype correlation. Hence, a moderate association between USP25 and epilepsy was noted, indicating that USP25 is potentially a predisposing gene for epilepsy. Our results from Usp25 null mice and the patient-derived variants indicated that USP25 would play an epileptogenic role via loss-of-function or gain-of-function effects. The truncated variant p.Gln889Ter would have a profoundly different effect on epilepsy. Together, our results underscore the significance of USP25 heterozygous variants in epilepsy, thereby highlighting the critical role of USP25 in the brain. [ABSTRACT FROM AUTHOR]

KCNMA1 located on chromosome 10q22.3, encodes the pore-forming α subunit of the 'Big K+' (BK) large conductance calcium and voltage-activated K + channel. Numerous evidence suggests the functional BK channel alterations produced by different KCNMA1 alleles may associate with different symptoms, such as paroxysmal non kinesigenic dyskinesia with gain of function and ataxia with loss of function. Functional classifications revealed two major patterns, gain of function and loss of function effects on channel properties in different cell lines. In the literature, two mutations have been shown to confer gain of function properties to BK channels: D434G and N995S. In this study, we report the functional characterization of a variant which was previously reported the whole exome sequencing revealed bi-allelic nonsense variation of the cytoplasmic domain of calcium-activated potassium channel subunit alpha-1 protein. To detect functional consequences of the variation, we parallely conducted two independent approaches. One is immunostaining using and the other one is electrophysiological recording using patch-clamp on wild-type and R458X mutant cells to detect the differences between wild-type and the mutant cells. We detected the gain of function effect for the mutation (NM_001161352.1 (ENST00000286628.8):c.1372C > T;Arg458*) using two parallel approaches. According to the result we found, the reported mutation causes the loss of function in the cell. It should be noted that in future studies, it can be thought that the functions of genes associated with channelopathies may have a dual effect such as loss and gain. [ABSTRACT FROM AUTHOR]

Rapeseed (Brassica napus) is an important oilseed crop worldwide. Plant vascular tissues are responsible for long-distance transport of water and nutrients and for providing mechanical support. The lateral roots absorb water and nutrients. The genetic basis of vascular tissue and lateral root development in rapeseed remains unknown. This study characterized an ethyl methanesulfonate-mutagenized rapeseed mutant, T16 , which showed dwarf stature, reduced lateral roots, and leaf wilting. SEM observations showed that the internode cells were shortened. Observations of tissue sections revealed defects in vascular bundle development in the stems and petioles. Genetic analysis revealed that the phenotypes of T16 were controlled by a single semi-dominant nuclear gene. Map-based cloning and genetic complementarity identified BnaA03.IAA13 as the functional gene; a G-to-A mutation in the second exon changed glycine at position 79 to glutamic acid, disrupting the conserved degron motif VGWPP. Transcriptome analysis in roots and stems showed that auxin and cytokinin signaling pathways were disordered in T16. Evolutionary analysis showed that AUXIN/INDOLE-3-ACETIC ACID is conserved during plant evolution. The heterozygote of T16 showed significantly reduced plant height while maintaining other agronomic traits. Our findings provide novel insights into the regulatory mechanisms of vascular tissue and lateral root development, and offer a new germplasm resource for rapeseed breeding. [ABSTRACT FROM AUTHOR]

Species of Saccharomyces genus have played an irreplaceable role in alcoholic beverage and baking industry for centuries. S. cerevisiae has also become an organism of choice for industrial production of alcohol and other valuable chemicals and a model organism shaping the rise of modern genetics and genomics in the past few decades. Today´s brewing industry faces challenges of decreasing consumption of traditional beer styles and increasing consumer demand for new styles, flavors and aromas. The number of currently used brewer's strains and their genetic diversity is yet limited and implementation of more genetic and phenotypic variation is seen as a solution to cope with the market challenges. This requires modification of current production strains or introduction of novel strains from other settings, e.g. industrial or wild habitats into the brewing industry. Due to legal regulation in many countries and negative customer perception of GMO organisms, the production of food and beverages requires non-GMO production organisms, whose development can be difficult and time-consuming. Here, we apply FIND-IT (Fast Identification of Nucleotide variants by DigITal PCR), an ultrafast genome-mining method, for isolation of novel yeast variants with varying flavor profiles. The FIND-IT method uses combination of random mutagenesis, droplet digital PCR with probes that target a specific desired mutation and a sub-isolation of the mutant clone. Such an approach allows the targeted identification and isolation of specific mutant strains with eliminated production of certain flavor and off-flavors and/or changes in the strain metabolism. We demonstrate that the technology is useful for the identification of loss-of function or gain of function mutations in unrelated industrial and wild strains differing in ploidy. Where no other phenotypic selection exists, this technology serves together with standard breeding techniques as a modern tool facilitating a modification of (brewer's) yeast strains leading to diversification of the product portfolio. • Mutagenesis and ddPCR allow for targeted mutant identification in brewer's yeast. • Modification of flavor profiles has been achieved in various industrial yeasts. • Heterothallic S. eubayanus strain facilitates development of novel lager hybrids. • Combination of the method with other non-GM techniques creates phenotypic diversity. • Method enables non-GMO engineering and improvement of brewing-related properties. [ABSTRACT FROM AUTHOR]

The KRAS protein, a product of the KRAS gene (V-ki-ras2 Kirsten rat sarcoma viral oncogene homolog), functions as a small GTPase that alternates between an active GTP-bound state (KRAS(ON)) and an inactive GDP-bound state (KRAS (OFF)). The KRASG12C mutation results in the accumulation of KRASG12C(OFF), promoting cell cycle survival and proliferation primarily through the canonical MAPK and PI3K pathways. The KRASG12C mutation is found in 13% of lung adenocarcinomas. Previously considered undruggable, sotorasib and adagrasib are the first available OFF-state KRASG12C inhibitors, but treatment resistance is frequent. In this review, after briefly summarizing the KRAS pathway and the mechanism of action of OFF-state KRASG12C inhibitors, we discuss primary and acquired resistance mechanisms. Acquired resistance is the most frequent, with "on-target" mechanisms such as a new KRAS mutation preventing inhibitor binding; and "off-target" mechanisms leading to bypass of KRAS through gain-of-function mutations in other oncogenes such as NRAS, BRAF, and RET; or loss-of-function mutations in tumor suppressor genes such as PTEN. Other "offtarget" mechanisms described include epithelial-to-mesenchymal transition and histological transformation. Multiple co-existing mechanisms can be found in patients, but few cases have been published. We highlight the lack of data on non-genomic resistance and the need for comprehensive clinical studies exploring histological, genomic, and non-genomic changes at resistance. This knowledge could help foster new treatment initiatives in this challenging context. [ABSTRACT FROM AUTHOR]

The OAS-RNase L pathway is one of the oldest innate RNA sensing pathways that leads to interferon (IFN) signaling and cell death. OAS recognizes viral RNA and then activates RNase L, which subsequently cleaves both cellular and viral RNA, creating "processed RNA" as an endogenous ligand that further triggers RIG-I-like receptor signaling. However, the IFN response and antiviral activity of the OAS-RNase L pathway are weak compared to other RNA-sensing pathways. Here, we discover that the SKIV2L RNA exosome limits the antiviral capacity of the OAS-RNase L pathway. SKIV2L-deficient cells exhibit remarkably increased interferon responses to RNase L-processed RNA, resulting in heightened antiviral activity. The helicase activity of SKIV2L is indispensable for this function, acting downstream of RNase L. SKIV2L depletion increases the antiviral capacity of OAS-RNase L against RNA virus infection. Furthermore, SKIV2L loss exacerbates autoinflammation caused by human OAS1 gain-of-function mutations. Taken together, our results identify SKIV2L as a critical barrier to OAS-RNase L-mediated antiviral immunity that could be therapeutically targeted to enhance the activity of a basic antiviral pathway. Synopsis: The antiviral activity of the OAS-RNase L pathway is weaker than that of other RNA-sensing pathways. This study shows a moderating effect of the SKIV2L cytoplasmic RNA exosome, which degrades the immunogenic RNA produced by RNase L. The SKIV2L cytoplasmic RNA exosome limits the dsRNA-induced innate immune response. The RNA helicase activity of SKIV2L is essential for this effect. SKIV2L acts selectively downstream of RNase L in the OAS-RNase L pathway. SKIV2L loss exacerbates autoinflammation caused by human OAS1 gain-of-function mutations. The helicase activity of SKIV2L acts downstream of RNase L to reduce the immunogenic output of the OAS-RNase L pathway. [ABSTRACT FROM AUTHOR]

Nucleostemin (NS) plays a role in liver regeneration, and aging reduces its expression in the baseline and regenerating livers following 70% partial hepatectomy (PHx). Here we interrogate the mechanism controlling NS expression during liver regeneration and aging. The NS promoter was analyzed by TRANSFAC. Functional studies were performed using cell-based luciferase assay, endogenous NS expression in Hep3B cells, mouse livers with a gain-of-function mutation of C/EBPα (S193D), and mouse livers with C/EBPα knockdown. We found a CAAT box with four C/EBPα binding sites (-1216 to -735) and a GC box with consensus binding sites for c-Myc, E2F1, and p300-associated protein complex (-633 to -1). Age-related changes in NS expression correlated positively with the expression of c-Myc, E2F1, and p300, and negatively with that of C/EBPα and C/EBPβ. PHx upregulated NS expression at 1d, coinciding with an increase in E2F1 and a decrease in C/EBPα. C/EBPα bound to the consensus sequences found in the NS promoter in vitro and in vivo, inhibited its transactivational activity in a binding site-dependent manner, and decreased the expression of endogenous NS in Hep3B cells. In vivo activation of C/EBPα by the S193D mutation resulted in a 4th-day post-PHx reduction of NS, a feature shared by 16-m/o livers. Finally, C/EBPα knockdown increased its expression in aged (24-m/o) livers under both baseline and regeneration conditions. This study reports the C/EBPα suppression of NS expression in aged livers, providing a new perspective on the mechanistic orchestration of tissue homeostasis in aging. [ABSTRACT FROM AUTHOR]

Tuberous Sclerosis Complex (TSC) is a lynchpin disorder, as it results in overactive mammalian target of rapamycin (mTOR) signaling, which has been implicated in a multitude of disease states. TSC is an autosomal dominant disease where 90% of affected individuals develop epilepsy. Epilepsy results from aberrant neuronal excitability that leads to recurring seizures. Under neurotypical conditions, the coordinated activity of voltage-gated ion channels keep neurons operating in an optimal range, thus providing network stability. Interestingly, loss or gain of function mutations in voltage-gated potassium, sodium, or calcium channels leads to altered excitability and seizures. To date, little is known about voltage-gated ion channel expression and function in TSC. However, data is beginning to emerge on how mTOR signaling regulates voltage-gated ion channel expression in neurons. Herein, we provide a comprehensive review of the literature describing common seizure types in patients with TSC, and suggest possible parallels between acquired epilepsies with known voltage-gated ion channel dysfunction. Furthermore, we discuss possible links towardmTOR regulation of voltage-gated ion channels expression and channel kinetics and the underlying epileptic manifestations in patients with TSC. [ABSTRACT FROM AUTHOR]

Warts, Hypogammaglobulinemia, Infections, and Myelokathexis (WHIM) syndrome is a rare primary immunodeficiency disease in humans caused by a gain of function in CXCR4, mostly due to inherited heterozygous mutations in CXCR4. One major clinical symptom of WHIM patients is their high susceptibility to human papillomavirus (HPV) induced disease, such as warts. Persistent high risk HPV infections cause 5% of all human cancers, including cervical, anogenital, head and neck and some skin cancers. WHIM mice bearing the same mutation identified in WHIM patients were created to study the underlying causes for the symptoms manifest in patients suffering from the WHIM syndrome. Using murine papillomavirus (MmuPV1) as an infection model in mice for HPV-induced disease, we demonstrate that WHIM mice are more susceptible to MmuPV1-induced warts (papillomas) compared to wild type mice. Namely, the incidence of papillomas is higher in WHIM mice compared to wild type mice when mice are exposed to low doses of MmuPV1. MmuPV1 infection facilitated both myeloid and lymphoid cell mobilization in the blood of wild type mice but not in WHIM mice. Higher incidence and larger size of papillomas in WHIM mice correlated with lower abundance of infiltrating T cells within the papillomas. Finally, we demonstrate that transplantation of bone marrow from wild type mice into WHIM mice normalized the incidence and size of papillomas, consistent with the WHIM mutation in hematopoietic cells contributing to higher susceptibility of WHIM mice to MmuPV1-induced disease. Our results provide evidence that MmuPV1 infection in WHIM mice is a powerful preclinical infectious model to investigate treatment options for alleviating papillomavirus infections in WHIM syndrome. Author summary: Mice carrying the same gain-of-function mutation in the gene CXCR4 that is present in human patients suffering from the Warts, Hypogammaglobulinemia, Infections, and Myelokathexis (WHIM) syndrome were previously created to understand the biology underlying this syndrome and to develop better means for treating WHIM patients. WHIM mice display neutropenia and lymphopenia symptoms as do WHIM patients. One of the key features of the WHIM syndrome in humans is increased susceptibility to infections by human papillomaviruses (HPV) with the majority of WHIM patients experiencing persistent warts and some developing anogenital cancers, both caused by HPVs. In this study we use a mouse papillomavirus, MmuPV1, which is a model for HPV infection in humans, to ask if the WHIM mice are more susceptible to infection and to understand why. We demonstrate that WHIM mice are more susceptible to MmuPV1-induced disease and that correcting the neutropenia and lymphopenia by bone marrow transplantation was effective at decreasing susceptibility to MmuPV1 induced disease. Our data support WHIM mice as a disease model for WHIM syndrome for future investigations on curative treatment options. [ABSTRACT FROM AUTHOR]

Summary: Systemic mastocytosis (SM) is a rare haematological neoplasm associated with the gain of function mutation KIT D816V in 90% of adult patients. Classically, cytogenetic aberrations are not common except in cases of SM associated with another haematological neoplasm. We highlight here an unusual clinical presentation of SM and demonstrate the utility of advanced cytogenetic analysis (optical genome mapping, OGM) in detecting a novel cytogenetic abnormality resulting in an unusual mechanism of DNMT3A and TET2 loss of function. [ABSTRACT FROM AUTHOR]

Background: Developmental and epileptic encephalopathies (DEE) are a group of disorders often linked to de novo mutations, including those in the ATP6V1A gene. These mutations, particularly dominant gain-of-function (GOF) variants, have been associated with a spectrum of phenotypes, ranging from severe DEE and infantile spasms to milder conditions like autism spectrum disorder and language delays. Methods: We aim to expand ATP6V1A-related disease spectrum by describing a six-year-old boy who presented with a febrile seizure, mild intellectual disability (ID), language delay, acquired microcephaly, and dysmorphic features. Results: Genetic analysis revealed a novel de novo heterozygous pathogenic variant (c.82G>A, p.Val28Met) in the ATP6V1A gene. He did not develop epilepsy, and neuroimaging remained normal over five years of follow-up. Although ATP6V1A mutations have traditionally been linked to severe neurodevelopmental disorders, often with early-onset epilepsy, they may exhibit milder, non-progressive phenotypes, challenging previous assumptions about the severity of ATP6V1A-related conditions. Conclusions: This case expands the known clinical spectrum, illustrating that not all patients with ATP6V1A mutations exhibit severe neurological impairment or epilepsy and underscoring the importance of including this gene in differential diagnoses for developmental delays, especially when febrile seizures or dysmorphic features are present. Broader genotype–phenotype correlations are essential for improving predictive accuracy and guiding clinical management, especially as more cases with mild presentations are identified. [ABSTRACT FROM AUTHOR]

Caffeine is a natural compound that inhibits the major cellular signaling regulator target of rapamycin (TOR), leading to widespread effects including growth inhibition. Saccharomyces cerevisiae yeast can adapt to tolerate high concentrations of caffeine in coffee and cacao fermentations and in experimental systems. While many factors affecting caffeine tolerance and TOR signaling have been identified, further characterization of their interactions and regulation remain to be studied. We used experimental evolution of S. cerevisiae to study the genetic contributions to caffeine tolerance in yeast, through a collaboration between high school students evolving yeast populations coupled with further research exploration in university labs. We identified multiple evolved yeast populations with mutations in PDR1 and PDR5 , which contribute to multidrug resistance, and showed that gain-of-function mutations in multidrug resistance family transcription factors Pdr1 , Pdr3 , and Yrr1 differentially contribute to caffeine tolerance. We also identified loss-of-function mutations in TOR effectors Sit4 , Sky1 , and Tip41 and showed that these mutations contribute to caffeine tolerance. These findings support the importance of both the multidrug resistance family and TOR signaling in caffeine tolerance and can inform future exploration of networks affected by caffeine and other TOR inhibitors in model systems and industrial applications. [ABSTRACT FROM AUTHOR]

Introduction: Alexander disease (AxD) is a rare leukodystrophy caused by dominant gain-of-function mutations in the gene encoding the astrocyte intermediate filament, glial fibrillary acidic protein (GFAP). However, there is an urgent need for biomarkers to assist in monitoring not only the progression of disease but also the response to treatment. GFAP is the obvious candidate for such a biomarker, as it is measurable in body fluids that are readily accessible for biopsy, namely cerebrospinal fluid and blood. However, in the case of ASOs, the treatment that is furthest in development, GFAP is the target of therapy and presumably would go down independent of disease status. Hence, there is a critical need for biomarkers that are not directly affected by the treatment strategy. Methods: We explored the potential utility of biomarkers currently being studied in other neurodegenerative diseases and injuries, specifically neurofilament light protein (NfL), phosphorylated forms of tau, and amyloid-β peptides (Aβ42/40). Results and Conclusions: Here, we report that GFAP is elevated in plasma of all age groups afflicted by AxD, including those with adult onset. NfL and p-tau are also elevated, but to a much lesser extent than GFAP. In contrast, the levels of Aß40 and Aß42 are not altered in AxD. [ABSTRACT FROM AUTHOR]

Lysosomes are dynamic cellular structures that adaptively remodel their membrane in response to stimuli, including membrane damage. Lysosomal dysfunction plays a central role in the pathobiology of Parkinson's disease (PD). Gain-of-function mutations in Leucine-rich repeat kinase 2 (LRRK2) cause familial PD and genetic variations in its locus increase the risk of developing the sporadic form of the disease. We previously uncovered a process we term LYTL (LYsosomal Tubulation/sorting driven by LRRK2), wherein membrane-damaged lysosomes generate tubules sorted into mobile vesicles. Subsequently, these vesicles interact with healthy lysosomes. LYTL is orchestrated by LRRK2 kinase activity, via the recruitment and phosphorylation of a subset of RAB GTPases. Here, we summarize the current understanding of LYTL and its regulation, as well as the unknown aspects of this process. [ABSTRACT FROM AUTHOR]

Activation of G proteins through nucleotide exchange initiates intracellular signaling cascades essential for life processes. Under normal conditions, nucleotide exchange is regulated by the formation of G protein–G protein–coupled receptor complexes. Single point mutations in the Gα subunit of G proteins bypass this interaction, leading to loss of function or constitutive gain of function, which is closely linked with the onset of multiple diseases. Despite the recognized significance of Gα mutations in disease pathology, structural information for most variants is lacking, potentially due to inherent protein dynamics that pose challenges for crystallography. To address this, we leveraged an integrative spectroscopic and computational approach to structurally characterize seven of the most frequently observed and clinically relevant mutations in the stimulatory Gα subunit, GαS. A previously proposed allosteric model of Gα activation linked structural changes in the nucleotide-binding pocket with functionally important changes in interactions between switch regions. We investigated this allosteric connection in GαS by integrating data from variable temperature CD spectroscopy, which measured changes in global protein structure and stability, and molecular dynamics simulations, which observed changes in interaction networks between GαS switch regions. Additionally, saturation-transfer difference NMR spectroscopy was applied to observe changes in nucleotide interactions with residues within the nucleotide binding site. These data have enabled testing of predictions regarding how mutations in GαS result in loss or gain of function and evaluation of proposed structural mechanisms. The integration of experimental and computational data allowed us to propose a more nuanced classification of mechanisms underlying GαS gain-of-function and loss-of-function mutations. [ABSTRACT FROM AUTHOR]

Genetic variations in the genes encoding G protein-coupled receptors (GPCRs) can disrupt receptor structure and function, which can result in human genetic diseases. Disease-causing mutations have been reported in at least 55 GPCRs for more than 66 monogenic diseases in humans. The spectrum of pathogenic and likely pathogenic variants includes loss of function variants that decrease receptor signaling on one extreme and gain of function that may result in biased signaling or constitutive activity, originally modeled on prototypical rhodopsin GPCR variants identified in retinitis pigmentosa, on the other. GPCR variants disrupt ligand binding, G protein coupling, accessory protein function, receptor desensitization and receptor recycling. Next generation sequencing has made it possible to identify variants of uncertain significance (VUS). We discuss variants in receptors known to result in disease and in silico strategies for disambiguation of VUS such as sorting intolerant from tolerant and polymorphism phenotyping. Modeling of variants has contributed to drug development and precision medicine, including drugs that target the melanocortin receptor in obesity and interventions that reverse loss of gonadotropin-releasing hormone receptor from the cell surface in idiopathic hypogonadotropic hypogonadism. Activating and inactivating variants of the calcium sensing receptor (CaSR) gene that are pathogenic in familial hypocalciuric hypercalcemia and autosomal dominant hypocalcemia have enabled the development of calcimimetics and calcilytics. Next generation sequencing has continued to identify variants in GPCR genes, including orphan receptors, that contribute to human phenotypes and may have therapeutic potential. Variants of the CaSR gene, some encoding an arginine-rich region that promotes receptor phosphorylation and intracellular retention, have been linked to an idiopathic epilepsy syndrome. Agnostic strategies have identified variants of the pyroglutamylated RF amide peptide receptor gene in intellectual disability and G protein-coupled receptor 39 identified in psoriatic arthropathy. Coding variants of the G protein-coupled receptor L1 (GPR37L1) orphan receptor gene have been identified in a rare familial progressive myoclonus epilepsy. The study of the role of GPCR variants in monogenic, Mendelian phenotypes has provided the basis of modeling the significance of more common variants of pharmacogenetic significance. [ABSTRACT FROM AUTHOR]

Summary: Osteoporosis-pseudoglioma syndrome (OPPG) and LRP5 high bone mass (LRP5-HBM) are two rare bone diseases with opposite clinical symptoms caused by loss-of-function and gain-of-function mutations in LRP5. Bisphosphonates are an effective treatment for OPPG patients. LRP5-HBM has a benign course, and age-related bone loss is found in one LRP5-HBM patient. Purpose: Low-density lipoprotein receptor-related protein 5 (LRP5) is involved in the canonical Wnt signaling pathway. The gain-of-function mutation leads to high bone mass (LRP5-HBM), while the loss-of-function mutation leads to osteoporosis-pseudoglioma syndrome (OPPG). In this study, the clinical manifestations, disease-causing mutations, treatment, and follow-up were summarized to improve the understanding of these two diseases. Methods: Two OPPG patients and four LRP5-HBM patients were included in this study. The clinical characteristics, biochemical and radiological examinations, pathogenic mutations, and structural analysis were summarized. Furthermore, several patients were followed up to observe the treatment effect and disease progress. Results: Congenital blindness, persistent bone pain, low bone mineral density (BMD), and multiple brittle fractures were the main clinical manifestations of OPPG. Complex heterozygous mutations were detected in two OPPG patients. The c.1455G > T mutation in exon 7 was first reported. During the follow-up, BMD of two patients was significantly improved after bisphosphonate treatment. On the contrary, typical clinical features of LRP5-HBM included extremely high BMD without fractures, torus palatinus and normal vision. X-ray showed diffuse osteosclerosis. Two heterozygous missense mutations were detected in four patients. In addition, age-related bone loss was found in one LRP5-HBM patient after 12-year of follow-up. Conclusion: This study deepened the understanding of the clinical characteristics, treatment, and follow-up of OPPG and LRP5-HBM; expanded the pathogenic gene spectrum of OPPG; and confirmed that bisphosphonates were effective for OPPG. Additionally, it was found that Ala242Thr mutation could not protect LRP5-HBM patients from age-related bone loss. This phenomenon deserves further study. [ABSTRACT FROM AUTHOR]

Proliferative glomerulonephritis is a severe condition that often leads to kidney failure. There is a significant lack of effective treatment for these disorders. Here, following the identification of a somatic PIK3CA gain-of-function mutation in podocytes of a patient, we demonstrate using multiple genetically engineered mouse models, single-cell RNA sequencing, and spatial transcriptomics the crucial role played by this pathway for proliferative glomerulonephritis development by promoting podocyte proliferation, dedifferentiation, and inflammation. Additionally, we show that alpelisib, a PI3Kα inhibitor, improves glomerular lesions and kidney function in different mouse models of proliferative glomerulonephritis and lupus nephritis by targeting podocytes. Surprisingly, we determined that pharmacological inhibition of PI3Kα affects B and T lymphocyte populations in lupus nephritis mouse models, with a decrease in the production of proinflammatory cytokines, autoantibodies, and glomerular complement deposition, which are all characteristic features of PI3Kδ inhibition, the primary PI3K isoform expressed in lymphocytes. Importantly, PI3Kα inhibition does not impact lymphocyte function under normal conditions. These findings were then confirmed in human lymphocytes isolated from patients with active lupus nephritis. In conclusion, we demonstrate the major role played by PI3Kα in proliferative glomerulonephritis and show that in this condition, alpelisib acts on both podocytes and the immune system. [ABSTRACT FROM AUTHOR]

GNAO1 mutated in pediatric encephalopathies encodes the major neuronal G protein Gαo. Of the more than 80 pathogenic mutations, most are single amino acid substitutions spreading across the Gαo sequence. We performed extensive characterization of Gαo mutants, showing abnormal GTP uptake and hydrolysis and deficiencies in binding Gβγ and RGS19. Plasma membrane localization of Gαo was decreased for a subset of mutations that leads to epilepsy; dominant interactions with GPCRs also emerged for the more severe mutants. Pathogenic mutants massively gained interaction with Ric8A and, surprisingly, Ric8B proteins, relocalizing them from cytoplasm to Golgi. Of these 2 mandatory Gα-subunit chaperones, Ric8A is normally responsible for the Gαi/Gαo, Gαq, and Gα12/Gα13 subfamilies, and Ric8B solely responsible for Gαs/Gαolf. Ric8 mediates the disease dominance when engaging in neomorphic interactions with pathogenic Gαo through imbalance of the neuronal G protein signaling networks. As the strength of Gαo-Ric8B interactions correlates with disease severity, our study further identifies an efficient biomarker and predictor for clinical manifestations in GNAO1 encephalopathies. Our work uncovers the neomorphic molecular mechanism of mutations underlying pediatric encephalopathies and offers insights into other maladies caused by G protein malfunctioning and further genetic diseases. [ABSTRACT FROM AUTHOR]

Recent research certified that DOT1L and its mutations represented by R231Q were potential targets for the treatment of lung cancer. Herein, a series of adenosine-containing derivatives were identified with DOT1LR231Q inhibition through antiproliferation assay and Western blot analysis in the H460R231Q cell. The most promising compound 37 significantly reduced DOT1LR231Q mediated H3K79 methylation and effectively inhibited the proliferation, self-renewal, migration, and invasion of lung cancer cell lines at low micromolar concentrations. The cell permeability and cellular target engagement of 37 were verified by both CETSA and DARTS assays. In the H460R231Q OE cell-derived xenograft (CDX) model, 37 displayed pronounced tumor growth inhibition after intraperitoneal administration at 20 mg/kg dose for 3 weeks (TGI = 54.38%), without obvious toxicities. A pharmacokinetic study revealed that 37 possessed tolerable properties (t 1/2 = 1.93 ± 0.91 h, F = 97.2%) after intraperitoneal administration in rats. Mechanism study confirmed that 37 suppressed malignant phenotypes of lung cancer carrying R231Q gain-of-function mutation via the MAPK/ERK signaling pathway. Moreover, analysis of the binding modes between molecules and DOT1LWT/R231Q proteins put forward the "Induced-fit" allosteric model in favor to the discovery of potent DOT1L candidates. A class of novel DOT1LR231Q inhibitors has been discovered, the most promising compound 37 exhibited favorable selectivity, potent in vitro and in vivo anti-lung cancer efficacy, and acceptable pharmacokinetic profile. [Display omitted] [ABSTRACT FROM AUTHOR]

Store-operated Ca2+ entry (SOCE) is a ubiquitous cellular mechanism that cells use to activate extracellular Ca2+ entry when intracellular Ca2+ stores are depleted. In skeletal muscle, SOCE occurs within Ca2+ entry units (CEUs), intracellular junctions between stacks of SR membranes containing STIM1 and transverse tubules (TTs) containing ORAI1. Gain-of-function mutations in STIM1 and ORAI1 are linked to tubular aggregate (TA) myopathy, a disease characterized by the atypical accumulation of tubes of SR origin. Moreover, SOCE and TAs are increased in the muscles of aged male mice. Here, we assessed the longitudinal effects (from 4–6 months to 10–14 months of age) of constitutive, muscle-specific Orai1 knockout (cOrai1 KO) on skeletal muscle structure, function, and the assembly of TAs and CEUs. The results from these studies indicate that cOrai1 KO mice exhibit a shorter lifespan, reduced body weight, exercise intolerance, decreased muscle-specific force and rate of force production, and an increased number of structurally damaged mitochondria. In addition, electron microscopy analyses revealed (i) the absence of TAs with increasing age and (ii) an increased number of SR stacks without adjacent TTs (i.e., incomplete CEUs) in cOrai1 KO mice. The absence of TAs is consistent with TAs being formed as a result of excessive ORAI1-dependent Ca2+ entry. [ABSTRACT FROM AUTHOR]

This article discusses the molecular landscape of FLT3 gene alterations in myeloid neoplasms (MNs), specifically acute myeloid leukemia (AML). FLT3 mutations, including internal tandem duplications (ITD) and mutations in the activation loop (AL), are common in AML and have been associated with poor prognosis. However, the presence and clinical implications of non-canonical FLT3 mutations (FLT3NC) have been less explored. The study analyzed a cohort of MN patients and found that FLT3NC alterations were more prevalent in non-AML MNs compared to AML. Additionally, the study identified three cases of germline FLT3NC, suggesting a potential role of hypomorphic germline FLT3 mutations in somatic gene rescue and leukemogenesis. The findings provide new insights into the complexity of FLT3 mutations in MNs. [Extracted from the article]

KRAS gain-of-function mutations are frequently observed in sporadic arteriovenous malformations. The mechanisms underlying the progression of such KRAS-driven malformations are still incompletely understood, and no treatments for the condition are approved. Here, we show the effectiveness of sotorasib, a specific KRAS G12C inhibitor, in reducing the volume of vascular malformations and improving survival in two mouse models carrying a mosaic Kras G12C mutation. We then administered sotorasib to two adult patients with severe KRAS G12C-related arteriovenous malformations. Both patients had rapid reductions in symptoms and arteriovenous malformation size. Targeting KRAS G12C appears to be a promising therapeutic approach for patients with KRAS G12C-related vascular malformations. [ABSTRACT FROM AUTHOR]

[Display omitted] Targeting oncogenic mutant p53 represents an attractive strategy for cancer treatment due to the high frequency of gain-of-function mutations and ectopic expression in various cancer types. Despite extensive efforts, the absence of a druggable active site for small molecules has rendered these mutants therapeutically non-actionable. Here we develop a selective and effective proteolysis-targeting chimera (PROTAC) for p53-R175H, a common hotspot mutant with dominant-negative and oncogenic activity. Using a novel iterative molecular docking-guided post-SELEX (systematic evolution of ligands by exponential enrichment) approach, we rationally engineer a high-performance DNA aptamer with improved affinity and specificity for p53-R175H. Leveraging this resulting aptamer as a binder for PROTACs, we successfully developed a selective p53-R175H degrader, named dp53m. dp53m induces the ubiquitin–proteasome-dependent degradation of p53-R175H while sparing wildtype p53. Importantly, dp53m demonstrates significant antitumor efficacy in p53-R175H-driven cancer cells both in vitro and in vivo , without toxicity. Moreover, dp53m significantly and synergistically improves the sensitivity of these cells to cisplatin, a commonly used chemotherapy drug. These findings provide evidence of the potential therapeutic value of dp53m in p53-R175H-driven cancers. [ABSTRACT FROM AUTHOR]

Cancer driver genes can undergo positive selection for various types of genetic alterations, including gain-of-function or loss-of-function mutations and copy number alterations (CNA). We investigated the landscape of different types of alterations affecting driver genes in 17,644 cancer exomes and genomes. We find that oncogenes may simultaneously exhibit signatures of positive selection and also negative selection in different gene segments, suggesting a method to identify additional tumor types where an oncogene is a driver or a vulnerability. Next, we characterize the landscape of CNA-dependent selection effects, revealing a general trend of increased positive selection on oncogene mutations not only upon CNA gains but also upon CNA deletions. Similarly, we observe a positive interaction between mutations and CNA gains in tumor suppressor genes. Thus, two-hit events involving point mutations and CNA are universally observed regardless of the type of CNA and may signal new therapeutic opportunities. An analysis with focus on the somatic CNA two-hit events can help identify additional driver genes relevant to a tumor type. By a global inference of point mutation and CNA selection signatures and interactions thereof across genes and tissues, we identify 9 evolutionary archetypes of driver genes, representing different mechanisms of (in)activation by genetic alterations. Inferring the emergence and selection of cancer drivers remains a daunting task. Here, the authors develop MutMatch, a statistical method to analyse somatic mutation rates and estimate conditional selection on cancer driver alterations and genes, which reveals cancer gene archetypes with specific selection pressures for different mutation types. [ABSTRACT FROM AUTHOR]

Gliomas are the most common primary brain tumor and are uniformly lethal. Despite significant advancements in understanding the genetic landscape of gliomas, standard-of-care has remained largely unchanged. Subsets of gliomas are defined by gain-of-function mutations in the metabolic genes encoding isocitrate dehydrogenase (IDH). Efforts to exploit mutant IDH activity and/or directly inhibit it with mutant IDH inhibitors have been the focus of over a decade of research. The recently published INDIGO trial, demonstrating the benefit of the mutant IDH inhibitor vorasidenib in patients with low-grade IDH-mutant gliomas, introduces a new era of precision medicine in brain tumors that is poised to change standard-of-care. In this review, we highlight and contextualize the results of the INDIGO trial and introduce key questions whose answers will guide how mutant IDH inhibitors may be used in the clinic. We discuss possible combination therapies with mutant IDH inhibition and future directions for clinical and translational research. [ABSTRACT FROM AUTHOR]

The IKZF1 gene encodes a transcription factor that belongs to the family of zincfinger DNA-binding proteins associated with chromatin remodeling. The protein product, IKAROS, had been proved to regulate lymphopoiesis. Subsequent mouse model studies have further confirmed its regulating role in lymphopoiesis as well as in hematopoiesis; besides, it associates with immune function, certain immune disorders like common variable immunodeficiency and dysgammaglobulinemia have been proved to be associated with germline IKZF1 mutations. Dysfunction of IKAROS also bears paramount significance in leukemic transformation and alterations of IKZF1 gene predicts a poor prognosis in hematological malignancies. As an independent prognostic marker, IKZF1 has been incorporated in the risk stratification of BCP-ALL and stratification-guided therapy has also been generated. In this review, we provide a concise and comprehensive overview on the multifaceted roles of IKZF1 gene. [ABSTRACT FROM AUTHOR]

Background: Intellectual disability (ID) is a neurodevelopmental condition affecting around 2% of children and young adults worldwide, characterized by deficits in intellectual functioning and adaptive behavior. Genetic factors contribute to the development of ID phenotypes, including mutations and structural changes in chromosomes. Pathogenic variants in the HCFC1 gene cause X-linked mental retardation syndrome, also known as Siderius type X-linked mental retardation. The MN1 gene is necessary for palate development, and mutations in this gene result in a genetic condition called CEBALID syndrome. Methods: Exome sequencing was used to identify the disease-causing variants in two affected families, A and B, from various regions of Pakistan. Affected individuals in these two families presented ID, developmental delay, and behavioral abnormalities. The validation and co-segregation analysis of the filtered variant was carried out using Sanger sequencing. Results: In an X-linked family A, a novel hemizygous missense variant (c.5705G > A; p.Ser1902Asn) in the HCFC1 gene (NM_005334.3) was identified, while in family B exome sequencing revealed a heterozygous nonsense variant (c.3680 G > A; p. Trp1227Ter) in exon-1 of the MN1 gene (NM_032581.4). Sanger sequencing confirmed the segregation of these variants with ID in each family. Conclusions: The investigation of two Pakistani families revealed pathogenic genetic variants in the HCFC1 and MN1 genes, which cause ID and expand the mutational spectrum of these genes. [ABSTRACT FROM AUTHOR]

We report the molecular and hematological identifications of two novel δ-globin gene mutations found in Guangxi Zhuang Autonomous Region, China. Capillary electrophoresis of the proband showed 1.3% Hb A2, accompanied by a minor unknown peak (0.7%) within the Z1 zone. High-performance liquid chromatography also revealed the presence of 1.5% Hb A2 and a 0.6% unknown peak. Routine genetic testing (Gap-PCR and reverse dot-blot hybridization) for common α-thalassemia was performed, and no mutations were observed. Sanger sequencing identified a heterozygous mutation for GAC > AAC at codon 79 (HBD:c.238G > A) and G > A at polyA + 70 (HBD:c.*200G > A) of the δ-globin gene. This variant was named Hb A2-Guangxi [δ79 (EF3) Asp→Asn, HBD:c.238G > A] after the geographic origin of the proband. [ABSTRACT FROM AUTHOR]

Innate immune responses that allow hosts to survive infection depend on the action of multiple conserved signaling pathways. Pathogens and parasites in turn have evolved virulence factors to target these immune signaling pathways in an attempt to overcome host immunity. Consequently, the interactions between host immune molecules and pathogen virulence factors play an important role in determining the outcome of an infection. The immune responses of Drosophila melanogaster provide a valuable model to understand immune signaling and host-pathogen interactions. Flies are commonly infected by parasitoid wasps and mount a coordinated cellular immune response following infection. This response is characterized by the production of specialized blood cells called lamellocytes that form a tight capsule around wasp eggs in the host hemocoel. The conserved JAK-STAT signaling pathway has been implicated in lamellocyte proliferation and is required for successful encapsulation of wasp eggs. Here we show that activity of Stat92E, the D. melanogaster STAT ortholog, is induced in immune tissues following parasitoid infection. Virulent wasp species are able to suppress Stat92E activity during infection, suggesting they target JAK-STAT pathway activation as a virulence strategy. Furthermore, two wasp species (Leptopilina guineaensis and Ganaspis xanthopoda) suppress phenotypes associated with a gain-of-function mutation in hopscotch, the D. melanogaster JAK ortholog, indicating that they inhibit the activity of the core signaling components of the JAK-STAT pathway. Our data suggest that parasitoid wasp virulence factors block JAK-STAT signaling to overcome fly immune defenses. Author summary: Following infection, host immune responses are triggered to provide protection against the invading pathogen. The proper function of these responses depends on the activity of multiple immune signaling pathways. These pathways act in a coordinated manner to orchestrate the immune response, and any disruption can render the host susceptible to infection. Because of this sensitivity, many pathogen species have evolved virulence mechanisms that target host signaling pathways and disrupt their function. We are using the interaction between Drosophila melanogaster and Drosophila-infecting parasitoid wasps to study this relationship between host signaling and pathogen virulence. Parasitoids infect fly larvae, and during infection, transfer virulence protein containing venom into the host, providing a mechanism to alter host signaling. This study is focused on one key immune signaling pathway, the highly conserved JAK-STAT pathway. We find that many virulent parasitoid species restrict the activity of the host JAK-STAT pathway, providing a clue as to their virulence strategy. JAK-STAT signaling plays many additional roles in health and disease, and so this system provides a good model to further understand this important pathway. [ABSTRACT FROM AUTHOR]

The GSK3/SHAGGY‐like kinase plays critical roles in plant development and response to stress, but its specific function remains largely unknown in wheat (Triticum aestivum L.). In this study, we investigated the function of TaGSK3, a GSK3/SHAGGY‐like kinase, in wheat development and response to stress. Our findings demonstrated that TaGSK3 mutants had significant effects on wheat seedling development and brassinosteroid (BR) signalling. Quadruple and quintuple mutants showed amplified BR signalling, promoting seedling development, while a sextuple mutant displayed severe developmental defects but still responded to exogenous BR signals, indicating redundancy and non‐BR‐related functions of TaGSK3. A gain‐of‐function mutation in TaGSK3‐3D disrupted BR signalling, resulting in compact and dwarf plant architecture. Notably, this mutation conferred significant drought and heat stress resistance of wheat, and enhanced heat tolerance independent of BR signalling, unlike knock‐down mutants. Further research revealed that this mutation maintains a higher relative water content by regulating stomatal‐mediated water loss and maintains a lower ROS level to reduces cell damage, enabling better growth under stress. Our study provides comprehensive insights into the role of TaGSK3 in wheat development, stress response, and BR signal transduction, offering potential for modifying TaGSK3 to improve agronomic traits and enhance stress resistance in wheat. Summary statement: Our findings elucidated the redundant role of TaGSK3 in wheat brassinosteroid (BR) signalling and developmental phenotypes, as well as its unique ability to enhance heat resistance independently of the BR signal. [ABSTRACT FROM AUTHOR]

Alterations in the tumor suppressor TP53 gene are the most common mutations in high grade serous ovarian carcinoma. The impact of TP53 mutations on clinical outcomes and platinum resistance is controversial. We sought to evaluate the genomic profile of high grade serous ovarian carcinoma and explore the association of TP53 mutations with platinum resistance. Next generation sequencing data was obtained from our institutional database for patients with high grade serous ovarian carcinoma undergoing primary treatment. Sequencing data, demographic, and clinical information was reviewed. The primary outcome analyzed was time to recurrence or refractory diagnosis. Associations between the primary outcome and different classification schemes for TP53 mutations (structural, functional, hot spot, pathogenicity scores, immunohistochemical staining patterns) were performed. 209 patients met inclusion criteria. TP53 mutations were the most common mutation. There were no differences in platinum response with TP53 hotspot mutations or high pathogenicity scores. Presence of TP53 gain-of-function mutations or measure of TP53 gain-of function activity were not associated with platinum resistance. Immunohistochemical staining patterns correlated with expected TP53 protein function and were not associated with platinum resistance. TP53 hotspot mutations or high pathogenicity scores were not associated with platinum resistance or refractory disease. Contrary to prior studies, TP53 gain-of-function mutations were not associated with platinum resistance. Estimation of TP53 gain-of-function effect using missense mutation phenotype scores was not associated with platinum resistance. The polymorphic nature of TP53 mutations may be too complex to demonstrate effect using simple models, or response to platinum therapy may be independent of initiating TP53 mutation. • The impact of TP53 mutations on platinum response in high grade serous ovarian cancer is unclear. • High TP53 pathogenicity scores were not associated with differential responses to platinum therapy. • TP53 gain-of-function mutations were not associated with platinum resistance, contrary to prior studies. • TP53 gain-of-function effect using missense mutation phenotype scores was not associated with platinum resistance. • Immunohistochemical staining patterns correlated withTP53 function and were not associated with platinum resistance. [ABSTRACT FROM AUTHOR]

Background and purpose: Cryopyrin‐associated periodic syndrome is a rare autoinflammatory disease caused by gain‐of‐function mutations or variants in the NLRP3 gene. Clinically, patients suffer from a broad spectrum of both systemic and neurological symptoms. The aim of this study was to determine whether systemic inflammation demonstrated by serum amyloid A (SAA) elevation is associated with neuroinflammation assessed by optical coherence tomography (OCT). Methods: Thirty eyes of 15 patients with NLRP3 low penetrance mutations (PwNLRP3) and 20 eyes of 10 age‐ and sex‐matched healthy controls were examined by spectral‐domain OCT as part of routine clinical care. All retinal layers and clinical features were evaluated. Results: At baseline no significant retinal neuroaxonal inflammation or degeneration was observed in all measured retinal layers amongst PwNLRP3 compared with healthy controls. In a pooled analysis of all individual OCT time points a significant difference regarding the macular retinal nerve fibre layer was detected. Increased levels of SAA showed a positive association with averaged combined outer plexiform layer and outer nuclear layer volumes (ρ < 0.0001, r2 = 0.35). Conclusion: In cryopyrin‐associated periodic syndrome increased combined outer plexiform layer and outer nuclear layer volumes are mirrored by SAA increase, an acute phase reactant indicating systemic inflammation. Our findings identify OCT as a candidate biomarker to monitor subclinical neuroinflammation and to assess disease activity in PwNLRP3. [ABSTRACT FROM AUTHOR]

Purpose: Gain-of-function mutations in CTNNB1, gene encoding for β-catenin, are observed in 25–30% of hepatocellular carcinomas (HCCs). Recent studies have shown β-catenin activation to have distinct roles in HCC susceptibility to mTOR inhibitors and resistance to immunotherapy. Our goal was to develop and test a computational imaging-based model to non-invasively assess β-catenin activation in HCC, since liver biopsies are often not done due to risk of complications. Methods: This IRB-approved retrospective study included 134 subjects with pathologically proven HCC and available β-catenin activation status, who also had either CT or MR imaging of the liver performed within 1 year of histological assessment. For qualitative descriptors, experienced radiologists assessed the presence of imaging features listed in LI-RADS v2018. For quantitative analysis, a single biopsy proven tumor underwent a 3D segmentation and radiomics features were extracted. We developed prediction models to assess the β-catenin activation in HCC using both qualitative and quantitative descriptors. Results: There were 41 cases (31%) with β-catenin mutation and 93 cases (69%) without. The model's AUC was 0.70 (95% CI 0.60, 0.79) using radiomics features and 0.64 (0.52, 0.74; p = 0.468) using qualitative descriptors. However, when combined, the AUC increased to 0.88 (0.80, 0.92; p = 0.009). Among the LI-RADS descriptors, the presence of a nodule-in-nodule showed a significant association with β-catenin mutations (p = 0.015). Additionally, 88 radiomics features exhibited a significant association (p < 0.05) with β-catenin mutations. Conclusion: Combination of LI-RADS descriptors and CT/MRI-derived radiomics determine β-catenin activation status in HCC with high confidence, making precision medicine a possibility. [ABSTRACT FROM AUTHOR]

Candidiasis is a common fungal infection caused by Candida species, with Candida albicans being the most prevalent. Resistance to azole drugs, commonly used to treat Candida infections, poses a significant challenge. Transcriptional activator candidate 1 (TAC1) gene has emerged as a key player in regulating drug resistance in C. albicans. This review explores the structure and function of the TAC1 gene and its role in azole resistance. This gene encodes a transcription factor that controls the expression of genes involved in drug resistance, such as efflux pump genes (CDR1, CDR2, and MDR1) and ERG11. Mutations in TAC1 can increase these genes' expression and confer resistance to azoles. Various TAC1 gene mutations, mostly gain-of-function mutations, have been identified, which upregulate CDR1 and CDR2 expression, resulting in azole resistance. Understanding the mechanisms of azole resistance mediated by the TAC1 gene is crucial for the strategies in the effective antifungal development pipeline. [ABSTRACT FROM AUTHOR]

Integration of metabolites into the overall metabolic network of a cell requires careful coordination dependent upon the ultimate usage of the metabolite. Different stoichiometric needs, and thus pathway fluxes, must exist for compounds destined for diverse uses, such as carbon sources, nitrogen sources, or stress-protective agents. Herein, we expand upon our previous work that highlighted the nature of glycine betaine (GB) metabolism in Methylobacteria to examine the utilization of GB-derivative compounds dimethylglycine (DMG) and sarcosine into Methylorubrum extorquens in different metabolic capacities, including as sole nitrogen and/or carbon sources. We isolated gain-of-function mutations that allowed M. extorquens PA1 to utilize dimethylglycine as a carbon source and dimethylglycine and sarcosine as nitrogen source. Characterization of mutants demonstrated selection for variants of the AraC-like regulator Mext_3735 that confer constitutive expression of the GB metabolic gene cluster, allowing direct utilization of the downstream GB derivatives. Finally, among the distinct isolates examined, we found that catabolism of the osmoprotectant used for selection (GB or dimethylglycine) enhanced osmotic stress resistance provided in the presence of that particular osmolyte. Thus, access to the carbon and nitrogen and osmoprotective effects of GB and DMG are made readily accessible through adaptive mutations. In M. extorquens PA1, the limitations to exploiting this group of compounds appear to exist predominantly at the levels of gene regulation and functional activity, rather than being constrained by transport or toxicity. [ABSTRACT FROM AUTHOR]

The genetic predisposition to lymphoma is not fully understood. We identified 13 lymphoma‐cancer families (2011–2021), in which 27 individuals developed lymphomas and 26 individuals had cancers. Notably, male is the predominant gender in lymphoma patients, whereas female is the predominant gender in cancer patients (p =.019; OR = 4.72, 95% CI, 1.30–14.33). We collected samples from 18 lymphoma patients, and detected germline variants through exome sequencing. We found that germline protein truncating variants (PTVs) were enriched in DNA repair and immune genes. Totally, we identified 31 heterozygous germline mutations (including 12 PTVs) of 25 DNA repair genes and 19 heterozygous germline variants (including 7 PTVs) of 14 immune genes. PTVs of ATM and PNKP were found in two families, respectively. We performed whole genome sequencing of diffuse large B cell lymphomas (DLBCLs), translocations at IGH locus and activation of oncogenes (BCL6 and MYC) were verified, and homologous recombination deficiency was detected. In DLBCLs with germline PTVs of ATM, deletion and insertion in CD58 were further revealed. Thus, in lymphoma‐cancer families, we identified germline defects of both DNA repair and immune genes in lymphoma patients. [ABSTRACT FROM AUTHOR]

Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies globally, representing a significant public health problem with a poor prognosis. The development of efficient therapeutic strategies for HNSCC prevention and treatment is urgently needed. The PI3K/AKT/mTOR (PAM) signaling pathway is a highly conserved transduction network in eukaryotic cells that promotes cell survival, growth, and cycle progression. Dysfunction in components of this pathway, such as hyperactivity of PI3K, loss of PTEN function, and gain-of-function mutations in AKT, are well-known drivers of treatment resistance and disease progression in cancer. In this review, we discuss the major mutations and dysregulations in the PAM signaling pathway in HNSCC. We highlight the results of clinical trials involving inhibitors targeting the PAM signaling pathway as a strategy for treating HNSCC. Additionally, we examine the primary mechanisms of resistance to drugs targeting the PAM pathway and potential therapeutic strategies. [ABSTRACT FROM AUTHOR]

Aim Method Results Interpretation To expand the phenotypic spectrum of ADGRL1 and explore the correlation between epilepsy and the ADGRL1 genotype.We performed whole‐exome sequencing in a cohort of 115 families (including 195 males and 150 females) with familial febrile seizure or epilepsy with unexplained aetiology. The damaging effects of variants was predicted using protein modelling and multiple in silico tools. All reported patients with ADGRL1 pathogenic variants were analysed.One new ADGRL1 variant (p.Pro753Leu) was identified in one family with genetic epilepsy with febrile seizures. Further analysis of 12 ADGRL1 variants in 16 patients revealed that six patients had epilepsy. Epilepsy types ranged from early‐onset epileptic encephalopathy to genetic epilepsy with febrile seizures plus (GEFS+). All four variants associated with epilepsy were located in the non‐helix or sheet region of ADGRL1. Three of the four epilepsy‐associated variants were missense variants. Thus, all three variants located in the G‐protein‐coupled receptor autoproteolytic‐inducing domain exhibited epilepsy.We found one new missense variant of ADGRL1 in one family with GEFS+. ADGRL1 may be a potential candidate or susceptibility gene for epilepsy. ADGRL1‐associated epilepsy ranged from benign GEFS+ to severe epileptic encephalopathy; the genotypes and variant locations may help explain the phenotypic heterogeneity of patients with the ADGRL1 variant. [ABSTRACT FROM AUTHOR]

Background: Catenin (Cadherin-Associated Protein), Beta 1 (CTNNB1) genomic alterations are rare in prostate cancer (PCa). Gain-of-function mutations lead to overexpression of β-catenin, with consequent hyperactivation of the Wnt/β-catenin signaling pathway, implicated in PCa progression and treatment resistance. To date, successful targeted treatment options for Wnt/β-catenin - driven PCa are lacking. Methods: We report a rare histologic transformation of a CTNNB1 (β-catenin) mutated metastatic castration resistant prostate cancer (mCRPC), clinically characterized by highly aggressive disease course. We histologically and molecularly characterized the liver metastatic tumor samples, as well as successfully generated patient-derived organoids (PDOs) and patient-derived xenograft (PDX) from a liver metastasis. We used the generated cell models for further molecular characterization and drug response assays. Results: Immunohistochemistry of liver metastatic biopsies and PDX tumor showed lack of expression of typical PCa (e.g., AR, PSA, PSAP, ERG) or neuroendocrine markers (synaptophysin), compatible with double-negative CRPC, but was positive for nuclear β-catenin expression, keratin 7 and 34βE12. ERG rearrangement was confirmed by fluorescent in situ hybridization (FISH). Drug response assays confirmed, in line with the clinical disease course, lack of sensitivity to common drugs used in mCRPC (e.g., enzalutamide, docetaxel). The casein kinase 1 (CK1) inhibitor IC261 and the tankyrase 1/2 inhibitor G700-LK showed modest activity. Moreover, despite harbouring a CTNNB1 mutation, PDOs were largely insensitive to SMARCA2/4- targeting PROTAC degraders and inhibitor. Conclusions: The reported CTNNB1-mutated mCRPC case highlights the potential challenges of double-negative CRPC diagnosis and underlines the relevance of further translational research to enable successful targeted treatment of rare molecular subtypes of mCRPC. [ABSTRACT FROM AUTHOR]

Background: Haemochromatosis is a genetic disease characterized by the excessive deposition of iron in various tissues and organs, eventually results in organ damage including cirrhosis, diabetes, cardiomyopathy, etc. SLC40A1-related haemochromatosis is associated with gain-of-function mutations in the SLC40A1 gene, which encodes ferroportin. While sporadic reports of this condition exist in mainland China, the understanding of the phenotype and genetic pattern associated with the SLC40A1 p.Y333H mutation remains incomplete. Case presentation: We report a pedigree with heterozygous p.Y333H mutation in Chinese Han population. The proband is a 64-year-old man complaining of persistent abnormality of liver enzyme levels for 1 year, with a history of knee joint pain, diabetes and skin pigmentation. He displayed markedly elevated serum ferritin level and transferrin saturation. Magnetic resonance imaging showed iron deposition in the liver, spleen, and pancreas, along with cirrhosis and splenomegaly. Whole exome sequencing identified a heterozygous allelic variant c.997T > C (p.Y333H). Genetic screening of family members identified four first-degree relatives and three second-degree relatives having the same mutation. Additional cases with this mutation from two published studies were included. Among the probands and screened relatives, all eight males aged over 30 y had ferritin level > 1000 µg/L, transferrin saturation > 90%. Four patients with organ damage in the present study received therapeutic phlebotomy, alleviating clinical symptoms and improving in transferrin saturation and serum ferritin. Conclusions: This study reports the largest pedigree with heterozygous SLC40A1 p.Y333H mutation in the Chinese population to date. In Chinese families, males over 30 years old with hemochromatosis due to SLC40A1 p.Y333H mutation exhibit severe iron overload phenotypes. [ABSTRACT FROM AUTHOR]

This article discusses a case study of a 67-year-old woman who initially presented with T-cell large granular lymphocytic leukemia (T-LGLL) with eosinophilia. However, as the disease progressed, it transformed into acute myeloid leukemia (AML) originating from mutated myeloid cells. The study explores the clonal relationships and evolution of different cell populations and highlights the role of the STAT5B mutation in the development of myeloid neoplasms. The research suggests that STAT5B alone can induce myeloid leukemia and that the same mutation in different cell lineages can lead to diverse types of neoplasms. Further research is needed to understand the pathogenesis of STAT5B-mutated neoplasms and develop specific inhibitors. [Extracted from the article]

Background and Purpose: GRIN‐related disorders are neurodevelopmental disorders caused by mutations in N‐methyl‐D‐aspartate receptor (NMDAR) subunit genes. A large fraction of these mutations lead to a 'gain of function' (GoF) of the NMDAR. Patients present with a range of symptoms including epilepsy, intellectual disability, behavioural and motor. Controlling seizures is a significant unmet medical need in most patients with GRIN‐related disorders. Although several hundred GRIN mutations have been identified in humans, until recently none of the mouse models carrying Grin mutations/deletions showed an epileptic phenotype. The two recent exceptions both carry mutations of GluN2A. The aim of this study was to assess the efficacy of radiprodil, a selective negative allosteric modulator of GluN2B‐containing NMDARs, in counteracting audiogenic seizures (AGS) in a murine model carrying the GluN2A(N615S) homozygous mutation (Grin2aS/S mice). Experimental Approach: Grin2aS/S mice were acutely treated with radiprodil at different doses before the presentation of a high‐frequency acoustic stimulus commonly used for AGS induction. Key Results: Radiprodil significantly and dose‐dependently reduced the onset and severity of AGS in Grin2aS/S mice. Surprisingly, the results revealed a sex‐dependent difference in AGS susceptibility and in the dose‐dependent protection of radiprodil in the two genders. Specifically, radiprodil was more effective in female versus male mice. Conclusion and Implications: Overall, our data clearly show that radiprodil, a GluN2B selective negative allosteric modulator, may have the potential to control seizures in patients with GRIN2A GoF mutations. Further studies are warranted to better understand the sex‐dependent effects observed in this study. [ABSTRACT FROM AUTHOR]

Gain-of-function mutations in the LRRK2 gene cause Parkinson's disease (PD), characterized by debilitating motor and nonmotor symptoms. Increased phosphorylation of a subset of RAB GTPases by LRRK2 is implicated in PD pathogenesis. We find that increased phosphorylation of RAB3A, a cardinal synaptic vesicle precursor (SVP) protein, disrupts anterograde axonal transport of SVPs in iPSC-derived human neurons (iNeurons) expressing hyperactive LRRK2-p.R1441H. Knockout of the opposing protein phosphatase 1H (PPM1H) in iNeurons phenocopies this effect. In these models, the compartmental distribution of synaptic proteins is altered; synaptophysin and synaptobrevin-2 become sequestered in the neuronal soma with decreased delivery to presynaptic sites along the axon. We find that RAB3A phosphorylation disrupts binding to the motor adaptor MADD, potentially preventing the formation of the RAB3A-MADD-KIF1A/1Bß complex driving anterograde SVP transport. RAB3A hyperphosphorylation also disrupts interactions with RAB3GAP and RAB-GDI1. Our results reveal a mechanism by which pathogenic hyperactive LRRK2 may contribute to the altered synaptic homeostasis associated with characteristic nonmotor and cognitive manifestations of PD. [ABSTRACT FROM AUTHOR]

Gain-of-function mutations in the KCNT1 gene, which encodes the sodium-activated potassium channel known as SLACK, are associated with the rare but devastating developmental and epileptic encephalopathy known as epilepsy of infancy with migrating focal seizures (EIMFS). The design of small molecule inhibitors of SLACK channels represents a potential therapeutic approach to the treatment of EIMFS, other childhood epilepsies, and developmental disorders. Herein, we describe a hit optimization effort centered on a xanthine SLACK inhibitor (8) discovered via a high-throughput screen. Across three distinct regions of the chemotype, we synthesized 58 new analogs and tested each one in a whole-cell automated patch-clamp assay to develop structure–activity relationships for inhibition of SLACK channels. We further evaluated selected analogs for their selectivity versus a variety of other ion channels and for their activity versus clinically relevant SLACK mutants. Selectivity within the series was quite good, including versus hERG. Analog 80 (VU0948578) was a potent inhibitor of WT, A934T, and G288S SLACK, with IC50 values between 0.59 and 0.71 µM across these variants. VU0948578 represents a useful in vitro tool compound from a chemotype that is distinct from previously reported small molecule inhibitors of SLACK channels. [ABSTRACT FROM AUTHOR]