The causal effect of lower plasma sclerostin on cardiovascular disease (CVD) risk has previously been examined with the aim of investigating potential side effects of pharmacological sclerostin inhibition for treatment of osteoporosis. We explored the relationship between plasma sclerostin levels and CVDs and bone phenotypes using Mendelian randomization (MR) and correlation between plasma sclerostin levels and these outcomes. We used variants identified in genome-wide association studies of plasma sclerostin levels in large proteomic datasets from the UK Biobank (Olink) and Iceland (SomaScan) as instruments in two separate MR analyses. These analyses did not provide evidence of association between the effects of sequence variants on plasma sclerostin levels and their effects on CVDs and CVD risk factors (P > 0.05). Several of the instruments had heterogenic effects on bone phenotypes and causal estimates in MR were non-significant (P > 0.05/8). Plasma sclerostin levels correlated positively with coronary artery disease, myocardial infarction and CVD risk factors. Our results do not provide evidence supporting the hypothesis that lower plasma sclerostin levels increase CVD risk and suggest that plasma sclerostin levels are not a good surrogate for pharmacological inhibition.

Spinal muscular atrophy (SMA) is a neurodegenerative disease caused by reduced levels of the survival motor neuron (SMN) protein, an essential component of the RNA splicing machinery. Although disruption of alternative splicing is a well-established hallmark of SMA, the specific splicing events that contribute to disease pathogenesis remain poorly understood. We utilised an established Caenorhabditis elegans SMA model to investigate global splicing changes using poly(A)+ RNA-seq and custom transcriptome assembly. Zygotic loss of smn-1 led to extensive transcriptomic changes, including over 1000 alternative splicing events, many of which were functionally tied to larval development. Exon skipping and intron retention were the most prevalent splicing alterations, and sequence motif analysis indicated a general shift from strong to weak splice site usage; however, no single motif accounted for the majority of observed splicing changes. Notably, we identified an overlap between smn-1 dependent splicing and those regulated by U6 snRNA m6A methylation. Our findings reinforce the conserved, broad role of SMN in maintaining splicing fidelity and reveal specific sequence biases associated with splicing errors in SMA.

Orofacial clefts (OFCs) are one of the most common structural birth defects, with the prevalence of OFC varying across populations, and studies on the causes of OFCs in diverse populations are necessary, but still limited. We analyzed whole genome sequencing data on 419 parent-child trios from the Philippines, a population with a particularly high rate of OFC. To identify novel genes for OFCs, we studied both common variation and de novo variants (DNVs). We identified a significant enrichment in both loss-of-function (N = 62; P = 8.34 × 10-5) and protein-altering DNVs (N = 394; P = 1.49 × 10-7) among OFC probands. Among the genes individually enriched for DNVs was GRHL2 (P = 6.60 × 10-6), where there were two DNVs, a stop-gain and a frameshift deletion. We then queried OFC trios from other cohorts in the Gabriella Miller Kids First program (total N = 1254) and GeneMatcher and identified an 89 kb de novo deletion in GRHL2 and a de novo 8q22.3 microdeletion with one breakpoint in GRHL2. Additionally, within the common variant analyses we found significant gene x gene interactions with GRHL2. GRHL2 is a conserved transcription factor involved in embryonic development, with truncating mutations causing autosomal dominant progressive hearing loss and missense variants causing autosomal recessive ectodermal dysplasia. Heterozygous variation in its homolog, GRHL3, causes Van der Woude syndrome and isolated cleft palate. Additionally, mice deficient for either Grhl2 or Grhl3 have craniofacial anomalies, including facial and palatal clefts, strongly supporting GRHL2 as a risk locus for OFCs.

Fetal hemoglobin (HbF) modulates the clinical severity of sickle cell disease (SCD) by inhibiting the polymerization of sickle hemoglobin. Elevated HbF levels are associated with milder disease phenotypes, fewer Vaso-occlusive crises, and reduced organ damage. Understanding the molecular regulation of HbF expression is critical for the development of new therapeutic strategies, including pharmacologic agents and gene-based interventions aimed at ameliorating the course of SCD. We investigated transcriptomic expression in erythroid cells during the transition from the neonatal period to early childhood to identify genes associated with HbF regulation. Reticulocyte transcriptomes were compared between samples obtained at birth (cord blood), when HbF levels ranged from 72.6% to 90%, and at 18 months of age (whole blood), when HbF levels declined to 5.9%-10.3%. Reticulocytes were enriched, RNA extracted, and high-throughput RNA sequencing was performed, followed by differential gene expression and network analyses. Analysis of 20 346 genes revealed 1245 differentially expressed genes, of which 631 genes were upregulated in cord blood reticulocytes. The differentially expressed genes were significantly enriched in pathways related to cell signaling, proliferation, differentiation, metabolism, immune functionality, and erythropoiesis. Developmental shifts in the erythroid transcriptome uncover key biological processes that may regulate HbF expression. These findings offer a valuable panel of candidate genes for future functional studies and highlight new potential molecular targets for therapeutic modulation of HbF in sickle cell disease.

Thalassemia is a genetic blood disorder caused by disrupted hemoglobin synthesis, posing significant public health challenges. This study aims to expand the identification of novel thalassemia variants in a large cohort from pre-marriage screenings in Ganzhou, Southern Jiangxi, China. Data from 229 246 individuals screened through Next-Generation Sequencing (NGS) from 2019 to 2022 led to the identification of 180 participants with novel variants, marking the first large-scale documentation of such variations in the population. Among them, 51.1% were male with a mean age of 27 years, and the frequency of novel thalassemia variants was 0.079%. We uncovered 180 novel variants, including 68 α-thalassemia variants across 33 types (0.0297% frequency) and 112 β-thalassemia variants belonging to 40 unique types (0.0489% frequency). The most common α-thalassemia genotype was HBA1:c.95 + 9C > T at 17.65%, while HBB:c.-180G > C was most prevalent among β-thalassemia variants at 23.21%. Ten novel α-thalassemia variants were linked to mild α-thalassemia, and clinical phenotypes were documented for 21 complex genotypes. This study catalogues 73 novel variants and highlights the genetic diversity of thalassemia, informing future preventive strategies.

Alpha-fetoprotein (AFP), a fetal plasma protein, serves as a diagnostic marker for hepatocellular carcinoma (HCC) and germ cell tumors, with prior genome-wide association studies (GWAS) identifying AFP and PPIP5K1 as associated with its levels. The aim of this study was to identify novel genetic loci associated with serum AFP levels in the Taiwanese population and to elucidate their potential regulatory mechanisms, particularly in liver tissue, by integrating GWAS with expression quantitative trait loci (eQTL) analyses. We conducted a two-stage GWAS of serum AFP levels using participants from the Taiwan Biobank. The discovery cohort included 18 267 individuals, and findings were replicated in an independent sample of 21 994 individuals. Linear mixed models were used to assess genome-wide associations, adjusting for age, sex, and population structure via principal components. Quality control measures were applied to both genotyped and imputed SNPs. To explore functional implications, eQTL analyses were performed using publicly available liver tissue data, focusing on liver-specific regulatory effects. We identified 57 candidate genes across 10 genomic regions on chromosomes 2, 3, 4, 15, 17, and 22. For instance, SNPs in genes like SMC6, SENP7, and TP53BP1 demonstrated significant associations with AFP levels, contributing previously unreported genetic variations. eQTL analysis linked 55 of these genes in regulatory functions, especially within liver tissues, supporting their involvement in AFP expression. Our findings, integrating GWAS and eQTL approaches, enhance understanding of AFP heritability and suggest diagnostic and therapeutic potential for HCC, pending further validation in personalized medicine contexts.

Bi-allelic mutations in GBA1, a gene that encodes the lysosomal enzyme β-glucocerebrosidase (GCase), cause Gaucher disease (GD). Although GD carriers do not exhibit clinical manifestations, GBA1 mutations are the highest risk factor for Parkinson's disease (PD) in GD patients and carriers of the disease [1-5]. GCase breaks down glucosylceramide (GluCer), a sphingolipid that accumulates in GD. GluCer is deacylated by the lysosomal enzyme acid ceramidase (ACDase) to glucosylsphingosine (GluSph) [6-8]. GluSph is neurotoxic and accumulates to high levels in neuronopathic GD brains [9, 10]. However, whether this metabolic pathway involving ACDase plays a role in GBA1-associated PD (GBA1/PD) is not known. In this report we used induced pluripotent stem cells (hiPSCs) from PD patients harboring heterozygote GBA1 mutations to examine the role of ACDase in promoting α-synuclein accumulation and aggregation, a hallmark of PD. Compared to isogenic controls, hiPSC-derived PD dopamine (DA) neurons had elevated levels of pathogenic α-synuclein species. There was also reduced nuclear localization of transcription factor EB (TFEB), impaired autophagy, and decreased levels of cathepsin D (CathD), a lysosomal protease involved in α-synuclein degradation [11]. Treatment of the mutant DA neurons with a number of different ACDase inhibitors, or CRISPR/Cas9 knockdown (KD) of the ASAH1 gene, reversed all the phenotypic abnormalities of the mutant DA neurons. We conclude that in GBA1/PD-DA neurons, ACDase contributes to deregulation of key nodes of the autophagy/lysosomal pathway (ALP) involved in α-synuclein clearance. Our results suggest that ACDase is a potential therapeutic target for treating GBA1-associated PD.

Brody myopathy is an ultra-rare autosomal recessive inherited disorder that impairs skeletal muscle function in humans. It is caused by deficiency of the Sarco(Endo)plasmic reticulum Ca2+-ATPase isoform1 (SERCA1), arising from defects, mainly missense mutations, in the ATP2A1 gene. At present, neither specific therapy, nor mouse model exists for Brody myopathy. Bovine pseudomyotonia (PMT) is a very rare skeletal muscle disorder. As Brody myopathy, it is an autosomal recessive inherited disorder caused by missense variants in the atp2a1 gene. Most mutations generate proteins corrupted in proper folding that although catalytically active, were ubiquitinated and prematurely degraded by the ubiquitin-proteasome system, thus sharing with Cystic Fibrosis the same pathogenetic mechanism. Bovine PMT, despite unconventional, is currently the unique mammalian model of Brody disease. In this study, we show that CFTR correctors, particularly C17, successfully rescue SERCA1 mutants both in vitro and in vivo models. Our findings suggest that CFTR correctors may be a potential innovative pharmacological approach addressing Brody patients in which mutated SERCA1 retains its activity.

FOXP1 (Forkhead-box protein P1) is a crucial transcription factor in neural development and is associated with schizophrenia (SCZ). FOXP1-regulated genes may contribute to genetic risk of SCZ and this may vary across different stages of neurodevelopment. We analyzed RNA-seq transcriptomic data from mouse and human models of FOXP1 loss-of-function across prenatal and postnatal developmental stages, including neural stem cells from embryonic mice (E14.5) and human brain organoids (equivalent to second trimester), and cortical tissues from different mouse postnatal stages P0, P7, and P47. P0 in mice corresponds to the third trimester in humans, while P7 and P47 represent early childhood and adolescence, respectively. Linkage disequilibrium score regression assessed if FOXP1-regulated genes were enriched for SCZ heritability. Gene-set enrichment analysis investigated if FOXP1-regulated genes were enriched for SCZ-associated genes reported as differentially expressed in single cortical cell studies. SynGO analysis mapped FOXP1-regulated genes to synaptic locations and functions. FOXP1-regulated genes were enriched for SCZ heritability, with significant results for E14.5, P7 and P47 but not P0. The P7 gene-set showed the strongest enrichment for SCZ-associated genes from single cortical cell studies. FOXP1-regulated genes at both P7 and P47 were involved in multiple synaptic functions and were mainly enriched within glutamatergic excitatory neurons, with P47 also showing enrichment within GABAergic inhibitory neurons. Prenatal FOXP1-regulated genes were enriched in progenitor cells and also mapped to the synapse. Genetic risk for SCZ within FOXP1-regulated genes follows a dynamic trajectory across developmental stages, showing strongest effects at a timepoint that maps to early childhood.

To describe the genetic, phenotypic, and pathologic manifestations of patients presenting with inherited kidney cancer and germline variants of the Tuberous Sclerosis Complex (TSC) genes.

American Indians and Hispanics/Latinos have a high burden of chronic kidney disease (CKD) and they may share disease associated genetic variants. This study aims to identify loci for CKD and albuminuria that are shared between these populations.

A vascular anomaly could be a vascular tumor or a vascular malformation. Vascular malformation is subclassified into fast-flow, including arteriovenous malformation and portwine stain, and slow-flow group comprising venous malformation, lymphatic malformation, and venolymphatic malformation. Recent data have shown that somatic mutations of genes in PIK3/AKT/mTOR and RAS/MAPK/ERK pathways are a major cause of this disorder. We conducted a gene panel testing (129 genes) with high-depth next-generation sequencing (NGS), which can detect very low-level mosaicism (~ 1%), on the tissue obtained from 26 patients in a cohort of mixed types of vascular malformation, comprising 2 fast-flow and 24 slow-flow malformations. Pathogenic/likely pathogenic (P/LP) variants were identified in 21 of 26 patients, yielding the overall diagnostic rate of 80.8%. The leading causes identified were PIK3CA (57.1%) and TEK (33.3%), especially in the slow-flow group, whereas HRAS and GNAQ were found positive in patients with fast-flow malformations. Three of 11 P/LP variants were previously unreported in vascular malformation, including those from HRAS, PIK3CA, and TEK. Most variants were detected as a solo, except for double mutations of TEK in patients with blue rubber bleb nevus syndrome (BRBNS) and a non-syndromic venous malformation. The level of mosaicism in the tissue ranged from 0.93% to 16.53%, with 60% (15/25) of the variants having ≥ 5% mosaicism. Three variant of uncertain significance of IDH1 and NACC1 were found and deserve further investigation for their pathogenic role. Data from the present study suggest the potential benefit of targeted therapy, in particular drugs in the mTOR pathway, for these patients.

The DNA mismatch repair protein MSH3 reversibly shifts from nucleus to the cytosol upon IL-6 signaling, abrogating the repair function of the MSH3-MSH2 heterodimer in the nucleus and increases aggressiveness and metastasis potential of colorectal cancers. A polymorphism proximate to MSH3's nuclear localization signal (NLS), Δ27bpMSH3, alters NLS function such that IL-6 triggers Δ27bpMSH3 accumulation in the cytosol. Public databases indicate Δ27bpMSH3 is rare in the germline yet we previously identified its presence in half of colon cancer cell lines tested and 19% of ulcerative colitis (UC) tissue samples. Here in examining ~ 200 each of UC, early-onset (eo)CRC, and late-onset (lo)CRC patients, biallelic MSH3 NLS germline polymorphisms were exclusively present in 15% of controls but in 18% of UC and 17% of eoCRC patients and were higher among CRC stage 3/4 patients compared to stage 2 patients; these marginal increases could potentiate inflammation-to-cancer transformation and/or metastatic disease. Using cell models we demonstrate IL-6-induced binding of wild type and Δ27bpMSH3 to the NFκB activating complex NEMO/IKKγ which stabilizes MSH3 after disengaging from its nuclear partner MSH2, linking inflammation with DNA repair protein stability. Additional NLS modifications using MSH3-FLAG mimics cytosolic Δ27bpMSH3 retention to cause loss-of-function after inflammation.

The Meta-Analysis of Glucose and Insulin-related traits Consortium (MAGIC) identified 242 loci associated with glycaemic traits fasting insulin (FI), fasting glucose (FG), 2 h-Glucose (2hGlu), and glycated haemoglobin (HbA1c). However, for the majority, the causal variant(s) remain(s) unknown. Modelling multiple traits and integrating functional annotations have each been shown to improve fine-mapping resolution. Here, we aimed to determine whether combining these techniques would further improve fine-mapping resolution. Using single-trait fine-mapping results from FINEMAP as input, we performed multi-trait fine-mapping with flashfm at 50 loci significantly associated with more than one glycaemic trait. We used fGWAS to build models of enriched annotations by considering 32 cell-type specific and 28 static annotations. We used these models to define prior probabilities to perform annotation informed fine-mapping with both FINEMAP (single-trait) and flashfm (multi-trait). Multi-trait fine-mapping of 106 locus-trait associations significantly (P = 1.23 × 10-17) reduced the median size of the credible sets accounting for 99% of the posterior probability of being causal (99CS) to 21.5 variants compared to the 60.5 variants in single-trait fine-mapping. Annotation informed single-trait fine-mapping of 211 locus-trait associations reduced (P = 4.24 × 10-12) the median 99CS size from 72 in agnostic single-trait fine-mapping to 52 variants. Annotation informed multi-trait fine-mapping of 110 locus-trait associations led to a further significant (P = 2.69 × 10-18) decrease in median 99CS size to 14.5 variants compared to 51.0 in annotation informed single-trait fine-mapping. In conclusion, by applying combined multi-trait and annotation informed fine-mapping to 50 loci, we refined the number of potential causal variants by 71.1% compared to single-trait agnostic fine-mapping.

STXBP1 (Syntaxin-binding protein 1) is a presynaptic SNARE complex regulator essential for neurotransmitter release. De novo heterozygous mutations in Stxbp1 represent one of the most common genetic causes of early onset epileptic encephalopathies (STXBP1 related disorders, STXBP1-RD). STXBP1 protein is ubiquitously expressed across all neuronal populations. While impaired synaptic E/I balance is established, neuronal subtype-specific mechanisms of STXBP1-RD remain poorly defined. Here, we deployed multi-level genetic models to delineate the neuronal and behavioral consequences of STXBP1 insufficiency. In C. elegans, systemic evaluation of STXBP1 (UNC-18) deficiency revealed deficits in serotonergic neurons, manifested as progressive dendritic atrophy. In a new established Stxbp1 haploinsufficient mouse model, we confirmed serotonergic system dysfunction, characterized by reduced serotonergic neuron numbers, decreased 5-HT levels, and compensatory upregulation of serotonin receptors. Finally, mice with serotonergic neuron-specific Stxbp1 haploinsufficiency recapitulated a subset of neurological phenotypes. Together, this study reveals the underestimated serotonergic dysfunction as a pathological component of STXBP1-RD.

ADSS1 myopathy is an ultrarare congenital myopathy characterized by progressive cardiac and skeletal muscle degeneration with childhood to adolescent onset. This autosomal recessive disease is caused by mutations in the ADSS1 gene, encoding the enzyme adenylosuccinate synthetase (AdSS1). AdSS1 plays a critical role in the adenine nucleotide cycle, which is important for energy metabolism in muscle cells. Enzymatic defects, engendered by loss-of-function mutations in ADSS1, lead to a bottleneck in the adenine nucleotide cycle, causing metabolic dysfunction that ultimately results in progressive muscle weakness, mobility impairment, and respiratory and cardiac dysfunction, often requiring the use of a ventilator. Despite its debilitating nature, there are currently no cures or targeted treatments available, and little research into possible therapeutic strategies has been done. With a limited patient profile encompassing fewer than 200 known patients worldwide, establishing a mouse model for ADSS1 myopathy is critical to understanding its pathogenesis and for developing future therapies. Here, we present and characterize the first mouse model of ADSS1 myopathy-a constitutive Adss1 knockout model-by (1) defining its natural history, (2) exploring its metabolic pathomechanisms, and (3) characterizing its histopathological features. We find that Adss1KO/KO mice have subtle motor deficits and present with histopathological features consistent with patient phenotypes. Overall, we show that despite a relatively mild phenotype, this novel mouse model has quantifiable pathological features that can be used to develop therapies for, and further probe pathophysiology of, ADSS1 myopathy.

The small heat shock protein HSPB6 (a.k.a. Hsp20) is highly expressed in striated and smooth muscles. It modulates the oligomerization of its paralogs HSPB1 and CRYAB (HSPB5) and is involved e.g. in cytoskeletal regulation and autophagy. While HSPB6 variants have been implicated in cardiomyopathy, they have not been previously linked to neuromuscular disease. We report here a patient with late-onset myopathy and cataract, carrying in cis the novel HSPB6 variant c.464delC and the common polymorphism c.488G > C, together resulting in the extended protein p.Pro155Argfs*25;p.Gly163Arg. The family history was consistent with dominant inheritance. The mutant protein showed decreased solubility due to phase separation propensity, and caused mislocalization of CRYAB and BAG3, and a decrease of HSPB1 in transfected cells. The patient's muscle biopsy showed rimmed vacuoles and, in line with the functional studies, accumulation of HSPB6 and its interaction partners. The identified HSPB6 variants are most likely the cause of the muscle disease in this family, thus identifying HSPB6 mutations as a novel cause of vacuolar myopathy. Other reported HSPB6 variants causing a late frameshift or extension may cause disease in a similar fashion.

Hypertension is a prevalent age-related condition with varying life risk factors. This study aimed to develop age-stratified prediction models integrating genetic risk scores (GRS) with clinical factors to improve hypertension risk stratification.

Spinal Muscular Atrophy (SMA) is characterized by a reduction of survival of motoneuron (SMN) protein, resulting in proximal muscle atrophy. SMA is a multi-system disease involving patients with alterations in multiple organs and metabolic pathways. Approved therapies focus on increasing SMN protein level either in the central nervous system or systemically. However, none of these therapies result in a cure. Patients show perturbations in several organs, including altered lipid metabolism such as leptin proteohormone levels, dicarboxylic aciduria and altered β-oxidation. In this study, we describe alterations along the neuroendocrine axis of leptin homeostasis in white adipose tissue (WAT) and hypothalamus of the severe Taiwanese SMA mouse model. Body weight was significantly decreased in SMA mice accompanied by significantly changed leptin protein levels in WAT of pre-symptomatic (P3) mice. Additionally, transcriptome and proteome analyses of WAT and hypothalamus revealed alterations in lipid and glucose metabolic pathways. We also identified several altered targets associated with appetite regulation. Our findings emphasize dysregulations in lipid and glucose metabolism and reinforce the need for research on metabolism in a disease with a predominant neuromuscular phenotype.

Heterozygous missense variants in CDK19 have been found in patients diagnosed with Developmental and Epileptic Encephalopathy-87 (DEE87) who present with global developmental delay, intellectual disability and other neuromuscular deficiencies. Two missense variants in CDK19, Y32H and T196A, were first proposed to be loss of function based on experiments in Drosophila models of DEE87. Subsequently, it was proposed that Y32H is a gain of function with elevated kinase activity. We present a detailed functional evaluation of these dominant missense variants in several contexts. We use fly models of DEE87 in which endogenous cdk8, the fly ortholog to human CDK8 and CDK19, is depleted through RNA interference (RNAi) while expressing the human genes. Depletion of Drosophila cdk8 causes thicker muscle myofibrils, fused mitochondria, and climbing defects. The expression of wild-type human CDK19 in a fly cdk8 knockdown background rescues these defects, highlighting functional conservation. In our assays, we used a cdk8 depleted background and individually expressed either the variant or wildtype CDK19 to compare the function of the variants relative to wild-type. We demonstrate that Y32H can rescue defects caused by cdk8 depletion, while T196A is unable to due to possible loss of function. Further, we find that supplementation of the fly diet with an antioxidant improves T196A phenotypes. Our Drosophila studies allowed us to assay these variants for further insight into their functional nature and to obtain translational knowledge that may be applied back to human health.

The occurrence of preterm premature rupture of membranes (PPROM) significantly impacts maternal and fetal health due to its association with the cellular composition and genetic changes of the fetal membrane. However, the specific cell type responsible for triggering PPROM and the underlying mechanisms are still largely unexplored. We employed single-cell RNA sequencing (scRNA-seq) analysis on fetal membrane along with adjacent placental tissues about two centimeters from the umbilical cord obtained from women who delivered full-term in labor (FTIL), preterm premature without rupture of membrane (PPWROM), as well as PPROM，immunofluorescence were used to verify the findings. Our result s highlighted notable differences in cell type composition and interactions among these three groups. Of particular significance, we have identified a previously unrecognized subtype of trophoblast cells known as FABP7+Tb, a transitional state cell between cytotrophoblasts (CTB) and extravillous trophoblasts (EVT) cells, which appears to have some impact on PPWROM. Additionally, up-regulated expression of MMP11 in EVT-1 may serve as a promising biomarker for PPROM diagnosis. Furthermore, our study unveiled distinct interaction patterns among different trophoblast subtypes under varying pathological conditions, as well as significant variations in the interactions of trophoblast cells with other cell types, especially the pathways that are orchestrated by cell-cell cross-talk. Our study offers a comprehensive cell type and interaction map for the human fetal membrane along with adjacent placental tissues about two centimeters from the umbilical cord, providing insights into the molecular mechanisms that drive PPROM and uncovering potential targets for the early prediction of this condition.