RNA toxicity underlies the pathogenesis of disorders such as myotonic dystrophy type 1 (DM1). Muscular dystrophy is a key element of the pathology of DM1. The means by which RNA toxicity causes muscular dystrophy in DM1 is unclear. Here, we have used the DM200 mouse model of RNA toxicity due to the expression of a mutant DMPK 3'UTR mRNA to model the effects of RNA toxicity on muscle regeneration. Using a BaCl2-induced damage model, we find that RNA toxicity leads to decreased expression of PAX7, and decreased numbers of satellite cells, the stem cells of adult skeletal muscle (also known as MuSCs). This is associated with a delay in regenerative response, a lack of muscle fiber maturation and an inability to maintain a normal number of satellite cells. Repeated muscle damage also elicited key aspects of muscular dystrophy, including fat droplet deposition and increased fibrosis, and the results represent one of the first times to model these classic markers of dystrophic changes in the skeletal muscles of a mouse model of RNA toxicity. Using a ligand-conjugated antisense (LICA) oligonucleotide ASO targeting DMPK sequences for the first time in a mouse model of RNA toxicity in DM1, we find that treatment with IONIS 877864, which targets the DMPK 3'UTR mRNA, is efficacious in correcting the defects in regenerative response and the reductions in satellite cell numbers caused by RNA toxicity. These results demonstrate the possibilities for therapeutic interventions to mitigate the muscular dystrophy associated with RNA toxicity in DM1.

Periodontitis is a common inflammatory disease characterized by a complex etiology, which is the result of a combination of genetic and environmental factors. Genetic variants linked to the periodontitis disease were already investigated, however, little was known regarding the severity of this disease. Recently, long runs of homozygosity (ROH) were associated with several multifactorial diseases. Therefore, in our work, we tried to assess the role of ROH and periodontitis status. We found an association between the excess of homozygosity owing to ROH and staging of periodontitis. More in detail, the total amount of homozygosity owing to ROH is positively associated with an increased severity of periodontitis (P = 0.0001). Regression tree analysis showed the impact of ROH burden in discriminating individuals with mild periodontitis stages I and II and periodontitis stages III and IV (P < 0.001). Furthermore, ROH mapping highlights several regions associated with a severe status of periodontitis (odds ratio > 1). Among them, we found a total of 33 genes. Interestingly, some of these genes were previously associated with granulocyte or platelet measures, both linked to the onset and the progression of periodontal disease. Our results suggest the not only single variants association test could help to risk assessment but even individual genomic features; furthermore, our ROH mapping highlighted the possible role of multiple genes in periodontal development.

Inherited genetic risk factors play a role in multiple myeloma (MM), yet considerable missing heritability exists. Rare risk variants at genome-wide association study (GWAS) loci are a new avenue to explore. Pleiotropy between lymphoid neoplasms (LNs) has been suggested in family history and genetic studies, but no studies have interrogated sequencing for pleiotropic genes or rare risk variants. Sequencing genetically enriched cases can help discover rarer variants. We analyzed exome sequencing in familial or early-onset MM cases to identify rare, functionally relevant variants near GWAS loci for a range of LNs. A total of 149 distinct and significant LN GWAS loci have been published. We identified six recurrent, rare, potentially deleterious variants within 5 kb of significant GWAS single nucleotide polymorphisms in 75 MM cases. Mutations were observed in BTNL2, EOMES, TNFRSF13B, IRF8, ACOXL and TSPAN32. All six genes replicated in an independent set of 255 early-onset MM or familial MM or precursor cases. Expansion of our analyses to the full length of these six genes resulted in a list of 39 rare and deleterious variants, seven of which segregated in MM families. Three genes also had significant rare variant burden in 733 sporadic MM cases compared with 935 control individuals: IRF8 (P = 1.0 × 10-6), EOMES (P = 6.0 × 10-6) and BTNL2 (P = 2.1 × 10-3). Together, our results implicate six genes in MM risk, provide support for genetic pleiotropy between LN subtypes and demonstrate the utility of sequencing genetically enriched cases to identify functionally relevant variants near GWAS loci.

RNA-binding proteins (RBPs) play essential roles in diverse cellular processes through post-transcriptional regulation of RNAs. The subcellular localization of RBPs is thus under tight control, the breakdown of which is associated with aberrant cytoplasmic accumulation of nuclear RBPs such as TDP-43 and FUS, well-known pathological markers for amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD). Here, we report in Drosophila model for ALS/FTD that nuclear accumulation of a cytoplasmic RBP Staufen may be a new pathological feature. We found that in Drosophila C4da neurons expressing PR36, one of the arginine-rich dipeptide repeat proteins (DPRs), Staufen accumulated in the nucleus in Importin- and RNA-dependent manner. Notably, expressing Staufen with exogenous NLS-but not with mutated endogenous NLS-potentiated PR-induced dendritic defect, suggesting that nuclear-accumulated Staufen can enhance PR toxicity. PR36 expression increased Fibrillarin staining in the nucleolus, which was enhanced by heterozygous mutation of stau (stau+/-), a gene that codes Staufen. Furthermore, knockdown of fib, which codes Fibrillarin, exacerbated retinal degeneration mediated by PR toxicity, suggesting that increased amount of Fibrillarin by stau+/- is protective. stau+/- also reduced the amount of PR-induced nuclear-accumulated Staufen and mitigated retinal degeneration and rescued viability of flies expressing PR36. Taken together, our data show that nuclear accumulation of Staufen in neurons may be an important pathological feature contributing to the pathogenesis of ALS/FTD.

At least 5% of children present unexpected difficulties in expressing and understanding spoken language. This condition is highly heritable and often co-occurs with other neurodevelopmental disorders such as dyslexia and ADHD. Through an exome sequencing analysis, we identified a rare missense variant (chr16:84405221, GRCh38.p12) in the ATP2C2 gene. ATP2C2 was implicated in language disorders by linkage and association studies, and exactly the same variant was reported previously in a different exome sequencing study for language impairment (LI). We followed up this finding by genotyping the mutation in cohorts selected for LI and comorbid disorders. We found that the variant had a higher frequency in LI cases (1.8%, N = 360) compared with cohorts selected for dyslexia (0.8%, N = 520) and ADHD (0.7%, N = 150), which presented frequencies comparable to reference databases (0.9%, N = 24 046 gnomAD controls). Additionally, we observed that carriers of the rare variant identified from a general population cohort (N = 42, ALSPAC cohort) presented, as a group, lower scores on a range of reading and language-related measures compared to controls (N = 1825; minimum P = 0.002 for non-word reading). ATP2C2 encodes for an ATPase (SPCA2) that transports calcium and manganese ions into the Golgi lumen. Our functional characterization suggested that the rare variant influences the ATPase activity of SPCA2. Thus, our results further support the role of ATP2C2 locus in language-related phenotypes and pinpoint the possible effects of a specific rare variant at molecular level.

Angelman syndrome (AS) is a rare neurodevelopmental disorder characterized by impaired communication skills, ataxia, motor and balance deficits, intellectual disabilities, and seizures. The genetic cause of AS is the neuronal loss of UBE3A expression in the brain. A novel approach, described here, is a stem cell gene therapy which uses lentivector-transduced hematopoietic stem and progenitor cells to deliver functional UBE3A to affected cells. We have demonstrated both the prevention and reversal of AS phenotypes upon transplantation and engraftment of human CD34+ cells transduced with a Ube3a lentivector in a novel immunodeficient Ube3amat-/pat+ IL2rg-/y mouse model of AS. A significant improvement in motor and cognitive behavioral assays as well as normalized delta power measured by electroencephalogram was observed in neonates and adults transplanted with the gene modified cells. Human hematopoietic profiles observed in the lymphoid organs by detection of human immune cells were normal. Expression of UBE3A was detected in the brains of the adult treatment group following immunohistochemical staining illustrating engraftment of the gene-modified cells expressing UBE3A in the brain. As demonstrated with our data, this stem cell gene therapy approach offers a promising treatment strategy for AS, not requiring a critical treatment window.

The smallest genomic region causing Prader-Willi Syndrome (PWS) deletes the non-coding RNA SNORD116 cluster; however, the function of SNORD116 remains a mystery. Previous work in the field revealed the tantalizing possibility that expression of NHLH2, a gene previously implicated in both obesity and hypogonadism, was downregulated in PWS patients and differentiated stem cells. In silico RNA: RNA modeling identified several potential interaction domains between SNORD116 and NHLH2 mRNA. One of these interaction domains was highly conserved in most vertebrate NHLH2 mRNAs examined. A construct containing the Nhlh2 mRNA, including its 3'-UTR, linked to a c-myc tag was transfected into a hypothalamic neuron cell line in the presence and absence of exogenously-expressed Snord116. Nhlh2 mRNA expression was upregulated in the presence of Snord116 dependent on the length and type of 3'UTR used on the construct. Furthermore, use of actinomycin D to stop new transcription in N29/2 cells demonstrated that the upregulation occurred through increased stability of the Nhlh2 mRNA in the 45 minutes immediately following transcription. In silico modeling also revealed that a single nucleotide variant (SNV) in the NHLH2 mRNA could reduce the predicted interaction strength of the NHLH2:SNORD116 diad. Indeed, use of an Nhlh2 mRNA construct containing this SNV significantly reduces the ability of Snord116 to increase Nhlh2 mRNA levels. For the first time, these data identify a motif and mechanism for SNORD116-mediated regulation of NHLH2, clarifying the mechanism by which deletion of the SNORD116 snoRNAs locus leads to PWS phenotypes.

Obscurin is a giant muscle protein that connects the sarcomere with the sarcoplasmic reticulum, and has poorly understood structural and signalling functions. Increasingly, obscurin variants are implicated in the pathophysiology of cardiovascular diseases. The Arg4344Gln variant (R4344Q) in obscurin domain Ig58, initially discovered in a patient with hypertrophic cardiomyopathy, has been reported to reduce binding to titin domains Z8-Z9, impairing obscurin's Z-disc localization. An R4344Q knock-in mouse developed a cardiomyopathy-like phenotype with abnormal Ca2+-handling and arrhythmias, which were attributed to an enhanced affinity of a putative interaction between obscurin Ig58 and phospholamban (PLN) due to the R4344Q variant. However, the R4344Q variant is found in 15% of African Americans, arguing against its pathogenicity. To resolve this apparent paradox, we quantified the influence of the R4344Q variant (alongside another potentially pathogenic variant: Arg4444Trp (R4444W)) on binding to titin Z8-Z9, novex-3 and PLN using pull-down assays and microscale thermophoresis and characterized the influence on domain stability using differential scanning fluorimetry. We found no changes in titin binding and thermostability for both variants and modestly increased affinities of PLN for R4344Q and R4444W. While we could not confirm the novex-3/obscurin interaction, the PLN/obscurin interaction relies on the transmembrane region of PLN and is not reproducible in mammalian cells, suggesting it is an in vitro artefact. Without clear clinical evidence for disease involvement, we advise against classifying these obscurin variants as pathogenic.

Duchenne muscular dystrophy (DMD) is a severe X-linked inherited muscular disorder characterized by the loss of dystrophin. We have previously shown that monogene therapy using the mini-dystrophin gene improves muscle function in DMD. However, chronic inflammation plays an important role in progressive muscle degeneration in DMD as well. Vascular endothelial growth factor (VEGF) has been used to enhance muscle vasculature, reduce local inflammation and improve DMD muscle function. Temporalis muscles are the key skeletal muscles for mastication and loss of their function negatively affects DMD patient quality of life by reducing nutritional intake, but little is known about the pathology and treatment of the temporalis muscle in DMD. In this work, we tested the hypothesis that the combined delivery of the human mini-dystrophin and human VEGF genes to the temporalis muscles using separate recombinant adeno-associated viral (rAAV) vectors will synergistically improve muscle function and pathology in adult male dystrophin/utrophin double-knockout (mdx/utrn+/-) mice. The experimental mice were divided into four groups including: dystrophin + VEGF combined, dystrophin only, VEGF only and PBS control. After 2 months, gene expression and histological analysis of the temporalis muscles showed a synergistic improvement in temporalis muscle pathology and function coincident with increased restoration of dystrophin-associated protein complexes and nNOS in the dystrophin + VEGF combined group. We also observed significantly reduced inflammatory cell infiltration, central nucleation, and fibrosis in the dystrophin + VEGF combined group. We have demonstrated the efficacy of combined rAAV-mediated dystrophin and VEGF treatment of temporalis muscles in a DMD mouse model.

Syntaxin-binding protein 1 (STXBP1; also called MUNC18-1), encoded by STXBP1, is an essential component of the molecular machinery that controls synaptic vesicle docking and fusion. De novo pathogenic variants of STXBP1 cause a complex set of neurological disturbances, namely STXBP1 encephalopathy (STXBP1-E) that includes epilepsy, neurodevelopmental disorders and neurodegeneration. Several animal studies have suggested the contribution of GABAergic dysfunction in STXBP1-E pathogenesis. However, the pathophysiological changes in GABAergic neurons of these patients are still poorly understood. Here, we exclusively generated GABAergic neurons from STXBP1-E patient-derived induced pluripotent stem cells (iPSCs) by transient expression of the transcription factors ASCL1 and DLX2. We also generated CRISPR/Cas9-edited isogenic iPSC-derived GABAergic (iPSC GABA) neurons as controls. We demonstrated that the reduction in STXBP1 protein levels in patient-derived iPSC GABA neurons was slight (approximately 20%) compared to the control neurons, despite a 50% reduction in STXBP1 mRNA levels. Using a microelectrode array-based assay, we found that patient-derived iPSC GABA neurons exhibited dysfunctional maturation with reduced numbers of spontaneous spikes and bursts. These findings reinforce the idea that GABAergic dysfunction is a crucial contributor to STXBP1-E pathogenesis. Moreover, gene expression analysis revealed specific dysregulation of genes previously implicated in epilepsy, neurodevelopment and neurodegeneration in patient-derived iPSC GABA neurons, namely KCNH1, KCNH5, CNN3, RASGRF1, SEMA3A, SIAH3 and INPP5F. Thus, our study provides new insights for understanding the biological processes underlying the widespread neuropathological features of STXBP1-E.

Over 1200 variants in the ABCA4 gene cause a wide variety of retinal disease phenotypes, the best known of which is autosomal recessive Stargardt disease (STGD1). Disease-causing variation encompasses all mutation categories, from large copy number variants to very mild, hypomorphic missense variants. The most prevalent disease-causing ABCA4 variant, present in ~ 20% of cases of European descent, c.5882G > A p.(Gly1961Glu), has been a subject of controversy since its minor allele frequency (MAF) is as high as ~ 0.1 in certain populations, questioning its pathogenicity, especially in homozygous individuals. We sequenced the entire ~140Kb ABCA4 genomic locus in an extensive cohort of 644 bi-allelic, i.e. genetically confirmed, patients with ABCA4 disease and analyzed all variants in 140 compound heterozygous and 10 homozygous cases for the p.(Gly1961Glu) variant. A total of 23 patients in this cohort additionally harbored the deep intronic c.769-784C > T variant on the p.(Gly1961Glu) allele, which appears on a specific haplotype in ~ 15% of p.(Gly1961Glu) alleles. This haplotype was present in 5/7 of homozygous cases, where the p.(Gly1961Glu) was the only known pathogenic variant. Three cases had an exonic variant on the same allele with the p.(Gly1961Glu). Patients with the c.[769-784C > T;5882G > A] complex allele exhibit a more severe clinical phenotype, as seen in compound heterozygotes with some more frequent ABCA4 mutations, e.g. p.(Pro1380Leu). Our findings indicate that the c.769-784C > T variant is major cis-acting modifier of the p.(Gly1961Glu) allele. The absence of such additional allelic variation on most p.(Gly1961Glu) alleles largely explains the observed paucity of affected homozygotes in the population.

The Polycomb group (PcG) gene RNF2 (RING2) encodes a catalytic subunit of the Polycomb repressive complex 1 (PRC1), an evolutionarily conserved machinery that post-translationally modifies chromatin to maintain epigenetic transcriptional repressive states of target genes including Hox genes. Here, we describe two individuals, each with rare de novo missense variants in RNF2. Their phenotypes include intrauterine growth retardation, severe intellectual disabilities, behavioral problems, seizures, feeding difficulties and dysmorphic features. Population genomics data suggest that RNF2 is highly constrained for loss-of-function (LoF) and missense variants, and both p.R70H and p.S82R variants have not been reported to date. Structural analyses of the two alleles indicate that these changes likely impact the interaction between RNF2 and BMI1, another PRC1 subunit or its substrate Histone H2A, respectively. Finally, we provide functional data in Drosophila that these two missense variants behave as LoF alleles in vivo. The evidence provide support for deleterious alleles in RNF2 being associated with a new and recognizable genetic disorder. This tentative gene-disease association in addition to the 12 previously identified disorders caused by PcG genes attests to the importance of these chromatin regulators in Mendelian disorders.

ΔR4-R23/ΔCT micro-dystrophin (μDys) is a miniaturized version of dystrophin currently evaluated in a Duchenne muscular dystrophy (DMD) gene therapy trial to treat skeletal and cardiac muscle disease. In pre-clinical studies, μDys efficiently rescues cardiac histopathology, but only partially normalizes cardiac function. To gain insights into factors that may impact the cardiac therapeutic efficacy of μDys, we compared by mass spectrometry the composition of purified dystrophin and μDys protein complexes in the mouse heart. We report that compared to dystrophin, μDys has altered associations with α1- and β2-syntrophins, as well as cavins, a group of caveolae-associated signaling proteins. In particular, we found that membrane localization of cavin-1 and cavin-4 in cardiomyocytes requires dystrophin and is profoundly disrupted in the heart of mdx5cv mice, a model of DMD. Following cardiac stress/damage, membrane-associated cavin-4 recruits the signaling molecule ERK to caveolae, which activates key cardio-protective responses. Evaluation of ERK signaling revealed a profound inhibition, below physiological baseline, in the mdx5cv mouse heart. Expression of μDys in mdx5cv mice prevented the development of cardiac histopathology but did not rescue membrane localization of cavins nor did it normalize ERK signaling. Our study provides the first comparative analysis of purified protein complexes assembled in vivo by full-length dystrophin and a therapeutic micro-dystrophin construct. This has revealed disruptions in cavins and ERK signaling that may contribute to DMD cardiomyopathy. This new knowledge is important for ongoing efforts to prevent and treat heart disease in DMD patients.

Nemaline myopathy, a disease of the actin-based thin filament, is one of the most frequent congenital myopathies. To date, no specific therapy is available to treat muscle weakness in nemaline myopathy. We tested the ability of tirasemtiv, a fast skeletal troponin activator that targets the thin filament, to augment muscle force-both in vivo and in vitro-in a nemaline myopathy mouse model with a mutation (H40Y) in Acta1. In Acta1H40Y mice, treatment with tirasemtiv increased the force response of muscles to submaximal stimulation frequencies. This resulted in a reduced energetic cost of force generation, which increases the force production during a fatigue protocol. The inotropic effects of tirasemtiv were present in locomotor muscles and, albeit to a lesser extent, in respiratory muscles, and they persisted during chronic treatment, an important finding as respiratory failure is the main cause of death in patients with congenital myopathy. Finally, translational studies on permeabilized muscle fibers isolated from a biopsy of a patient with the ACTA1H40Y mutation revealed that at physiological Ca2+ concentrations, tirasemtiv increased force generation to values that were close to those generated in muscle fibers of healthy subjects. These findings indicate the therapeutic potential of fast skeletal muscle troponin activators to improve muscle function in nemaline myopathy due to the ACTA1H40Y mutation, and future studies should assess their merit for other forms of nemaline myopathy and for other congenital myopathies.

Several studies have reported association between leukocyte telomere length (LTL) and neuropsychiatric disorders. Although telomere length is affected by environmental factors, genetic variants in certain loci are strongly associated with LTL. Thus, we aimed to identify the genomic relationship between genetic variants of LTL with brain-based regulatory changes and brain volume. We tested genetic colocalization of seven and nine LTL loci in two ancestry groups, European (EUR) and East-Asian (EAS), respectively, with brain morphology measures for 101 T1-magnetic resonance imaging-based region of interests (n = 21 821). The posterior probability (>90%) was observed for 'fourth ventricle', 'gray matter' and 'cerebellar vermal lobules I-IV' volumes. We then tested causal relationship using LTL loci for gene and methylation expression. We found causal pleiotropy for gene (EAS = four genes; EUR = five genes) and methylation expression (EUR = 17 probes; EAS = 4 probes) of brain tissues (P ≤ 2.47 × 10-6). Integrating chromatin profiles with LTL-single nucleotide polymorphisms identified 45 genes (EUR) and 79 genes (EAS) (P ≤ 9.78×10-7). We found additional 38 LTL-genes using chromatin-based gene mapping for EUR ancestry population. Gene variants in three LTL-genes-GPR37, OBFC1 and RTEL1/RTEL1-TNFRSF6B-show convergent evidence of pleiotropy with brain morphology, gene and methylation expression and chromatin association. Mapping gene functions to drug-gene interactions, we identified process 'transmission across chemical synapses' (P < 2.78 × 10-4). This study provides evidence that genetic variants of LTL have pleiotropic roles with brain-based effects that could explain the phenotypic association of LTL with several neuropsychiatric traits.

Ultra-rare genetic disorders can provide proof of concept for efficacy of targeted therapeutics and reveal pathogenic mechanisms relevant to more common conditions. Juvenile polyposis of infancy (JPI) is caused by microdeletions in chromosome 10 that result in haploinsufficiency of two tumor suppressor genes: phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and bone morphogenetic protein receptor type IA (BMPR1A). Loss of PTEN and BMPR1A results in a much more severe phenotype than deletion of either gene alone, with infantile onset pan-enteric polyposis and a high mortality rate. No effective pharmacological therapy exists. A multi-center cohort analysis was performed to characterize phenotype and investigate the therapeutic effect of mammalian target of rapamycin (mTOR) inhibition (adverse events, disease progression, time to colectomy and mortality) in patients with JPI. Among 25 JPI patients identified (mean age of onset 13 months), seven received mTOR inhibitors (everolimus, n = 2; or sirolimus, n = 5). Treatment with an mTOR inhibitor reduced the risk of colectomy (hazard ratio = 0.27, 95% confidence interval = 0.07-0.954, P = 0.042) and resulted in significant improvements in the serum albumin level (mean increase = 16.3 g/l, P = 0.0003) and hemoglobin (mean increase = 2.68 g/dl, P = 0.0077). Long-term mTOR inhibitor treatment was well tolerated over an accumulated follow-up time of 29.8 patient years. No serious adverse events were reported. Early therapy with mTOR inhibitors offers effective, pathway-specific and personalized treatment for patients with JPI. Inhibition of the phosphoinositol-3-kinase-AKT-mTOR pathway mitigates the detrimental synergistic effects of combined PTEN-BMPR1A deletion. This is the first effective pharmacological treatment identified for a hamartomatous polyposis syndrome.

UBQLN2 mutations cause amyotrophic lateral sclerosis (ALS) with frontotemporal dementia (FTD), but the pathogenic mechanisms by which they cause disease remain unclear. Proteomic profiling identified 'mitochondrial proteins' as comprising the largest category of protein changes in the spinal cord (SC) of the P497S UBQLN2 mouse model of ALS/FTD. Immunoblots confirmed P497S animals have global changes in proteins predictive of a severe decline in mitochondrial health, including oxidative phosphorylation (OXPHOS), mitochondrial protein import and network dynamics. Functional studies confirmed mitochondria purified from the SC of P497S animals have age-dependent decline in nearly all steps of OXPHOS. Mitochondria cristae deformities were evident in spinal motor neurons of aged P497S animals. Knockout (KO) of UBQLN2 in HeLa cells resulted in changes in mitochondrial proteins and OXPHOS activity similar to those seen in the SC. KO of UBQLN2 also compromised targeting and processing of the mitochondrial import factor, TIMM44, resulting in accumulation in abnormal foci. The functional OXPHOS deficits and TIMM44-targeting defects were rescued by reexpression of WT UBQLN2 but not by ALS/FTD mutant UBQLN2 proteins. In vitro binding assays revealed ALS/FTD mutant UBQLN2 proteins bind weaker with TIMM44 than WT UBQLN2 protein, suggesting that the loss of UBQLN2 binding may underlie the import and/or delivery defect of TIMM44 to mitochondria. Our studies indicate a potential key pathogenic disturbance in mitochondrial health caused by UBQLN2 mutations.

The N-glycosylation of immunoglobulin G (IgG) affects its structure and function. It has been demonstrated that IgG N-glycosylation patterns are inherited as complex quantitative traits. Genome-wide association studies identified loci harboring genes encoding enzymes directly involved in protein glycosylation as well as loci likely to be involved in regulation of glycosylation biochemical pathways. Many of these loci could be linked to immune functions and risk of inflammatory and autoimmune diseases. The aim of the present study was to discover and replicate new loci associated with IgG N-glycosylation and to investigate possible pleiotropic effects of these loci onto immune function and the risk of inflammatory and autoimmune diseases. We conducted a multivariate genome-wide association analysis of 23 IgG N-glycosylation traits measured in 8090 individuals of European ancestry. The discovery stage was followed up by replication in 3147 people and in silico functional analysis. Our study increased the total number of replicated loci from 22 to 29. For the discovered loci, we suggest a number of genes potentially involved in the control of IgG N-glycosylation. Among the new loci, two (near RNF168 and TNFRSF13B) were previously implicated in rare immune deficiencies and were associated with levels of circulating immunoglobulins. For one new locus (near AP5B1/OVOL1), we demonstrated a potential pleiotropic effect on the risk of asthma. Our findings underline an important link between IgG N-glycosylation and immune function and provide new clues to understanding their interplay.

Synaptic dysfunction and cognitive decline in Huntington's disease (HD) involve hyperactive A disintegrin and metalloproteinase domain-containing protein 10 (ADAM10). To identify the molecular mechanisms through which ADAM10 is associated with synaptic dysfunction in HD, we performed an immunoaffinity purification-mass spectrometry (IP-MS) study of endogenous ADAM10 in the brains of wild-type and HD mice. We found that proteins implicated in synapse organization, synaptic plasticity, and vesicle and organelles trafficking interact with ADAM10, suggesting that it may act as hub protein at the excitatory synapse. Importantly, the ADAM10 interactome is enriched in presynaptic proteins and ADAM10 co-immunoprecipitates with piccolo (PCLO), a key player in the recycling and maintenance of synaptic vesicles. In contrast, reduced ADAM10/PCLO immunoprecipitation occurs in the HD brain, with decreased density of synaptic vesicles in the reserve and docked pools at the HD presynaptic terminal. Conditional heterozygous deletion of ADAM10 in the forebrain of HD mice reduces active ADAM10 to wild-type level and normalizes ADAM10/PCLO complex formation and synaptic vesicle density and distribution. The results indicate that presynaptic ADAM10 and PCLO are a relevant component of HD pathogenesis.

The inverted triangle shape of South America places Argentina territory as a geographical crossroads between the two principal peopling streams that followed either the Pacific or the Atlantic coasts, which could have then merged in Central Argentina (CA). Although the genetic diversity from this region is therefore crucial to decipher past population movements in South America, its characterization has been overlooked so far. We report 92 modern and 22 ancient mitogenomes spanning a temporal range of 5000 years, which were compared with a large set of previously reported data. Leveraging this dataset representative of the mitochondrial diversity of the subcontinent, we investigate the maternal history of CA populations within a wider geographical context. We describe a large number of novel clades within the mitochondrial DNA tree, thus providing new phylogenetic interpretations for South America. We also identify several local clades of great temporal depth with continuity until the present time, which stem directly from the founder haplotypes, suggesting that they originated in the region and expanded from there. Moreover, the presence of lineages characteristic of other South American regions reveals the existence of gene flow to CA. Finally, we report some lineages with discontinuous distribution across the Americas, which suggest the persistence of relic lineages likely linked to the first population arrivals. The present study represents to date the most exhaustive attempt to elaborate a Native American genetic map from modern and ancient complete mitochondrial genomes in Argentina and provides relevant information about the general process of settlement in South America.

Cone dystrophies are a rare subgroup of inherited retinal dystrophies and hallmarked by color vision defects, low or decreasing visual acuity and central vision loss, nystagmus and photophobia. Applying genome-wide linkage analysis and array comparative genome hybridization, we identified a locus for autosomal dominant cone dystrophy on chromosome 16q12 in four independent multigeneration families. The locus is defined by duplications of variable size with a smallest region of overlap of 608 kb affecting the IRXB gene cluster and encompasses the genes IRX5 and IRX6. IRX5 and IRX6 belong to the Iroquois (Iro) protein family of homeodomain-containing transcription factors involved in patterning and regionalization of embryonic tissue in vertebrates, including the eye and the retina. All patients presented with a unique progressive cone dystrophy phenotype hallmarked by early tritanopic color vision defects. We propose that the disease underlies a misregulation of the IRXB gene cluster on chromosome 16q12 and demonstrate that overexpression of Irx5a and Irx6a, the two orthologous genes in zebrafish, results in visual impairment in 5-day-old zebrafish larvae.

The systematic identification of host genetic risk factors is essential for the understanding and treatment of coronavirus disease 2019 (COVID-19). By performing a meta-analysis of two independent genome-wide association summary datasets (N = 680 128), a novel locus at 21q22.11 was identified to be associated with COVID-19 infection (rs9976829 in IFNAR2-IL10RB, odds ratio = 1.16, 95% confidence interval = 1.09-1.23, P = 2.57 × 10-6). The rs9976829 represents a strong splicing quantitative trait locus for both IFNAR2 and IL10RB genes, especially in lung tissue (P = 1.8 × 10-24). Integrative genomics analysis of combining genome-wide association study with expression quantitative trait locus data showed the expression variations of IFNAR2 and IL10RB have prominent effects on COVID-19 in various types of tissues, especially in lung tissue. The majority of IFNAR2-expressing cells were dendritic cells (40%) and plasmacytoid dendritic cells (38.5%), and IL10RB-expressing cells were mainly nonclassical monocytes (29.6%). IFNAR2 and IL10RB are targeted by several interferons-related drugs. Together, our results uncover 21q22.11 as a novel susceptibility locus for COVID-19, in which individuals with G alleles of rs9976829 have a higher probability of COVID-19 susceptibility than those with non-G alleles.

Age-related macular degeneration (AMD) is a complex neurodegenerative eye disease with behavioral and genetic etiology and is the leading cause of irreversible vision loss among elderly Caucasians. Functionally significant genetic variants in the alternative pathway of complement have been strongly linked to disease. More recently, a rare variant in the terminal pathway of complement has been associated with increased risk, Complement component 9 (C9) P167S. To assess the functional consequence of this variant, C9 levels were measured in two independent cohorts of AMD patients. In both cohorts, it was demonstrated that the P167S variant was associated with low C9 plasma levels. Further analysis showed that patients with advanced AMD had elevated sC5b-9 compared to those with non-advanced AMD, although this was not associated with the P167S polymorphism. Electron microscopy of membrane attack complexes (MACs) generated using recombinantly produced wild type or P167S C9 demonstrated identical MAC ring structures. In functional assays, the P167S variant displayed a higher propensity to polymerize and a small increase in its ability to induce hemolysis of sheep erythrocytes when added to C9-depleted serum. The demonstration that this C9 P167S AMD risk polymorphism displays increased polymerization and functional activity provides a rationale for the gene therapy trials of sCD59 to inhibit the terminal pathway of complement in AMD that are underway.