OBJECTIVE:Dystonia is one of the most prevalent movement disorders, characterized by significant clinical and etiological heterogeneity. Despite considerable heritability (~25%), the etiology in most patients remains elusive. Moreover, understanding correlations between clinical manifestations and genetic variants has become increasingly complex. METHODS:Exome sequencing was conducted on 1924 genetically unsolved, mainly late-onset isolated dystonia patients, recruited primarily from two dystonia registries (DysTract and the Dystonia Coalition). Rare variants in genes previously linked to dystonia (n = 406) were examined, confirmed via Sanger sequencing, and analyzed for segregation when possible. RESULTS:We identified 137 distinct likely pathogenic/pathogenic variants (according to ACMG criteria) across 51 genes in 163/1924 patients, including 153/1895 index patients (diagnostic yield 8.1%). The strongest predictors of a genetic diagnosis were generalized dystonia (28.6% yield) and age at onset (20.4% yield in patients with onset < 30 years). Notably, 56.2% of these variants were novel, with recurrent variants in EIF2AK2, VPS16, KCNMA1, and SLC2A1. Additionally, 321 index patients (16.9%) harbored variants of uncertain significance in 102 genes. The most frequently implicated genes included VPS16, THAP1, GCH1, SGCE, GNAL, and KMT2B. Presumably pathogenic variants in less well-established dystonia genes were also found, including KCNMA1, KIF1A, and ZMYND11. At least six variants (in ADCY5, GNB1, IR2BPL, KCNN2, KMT2B, and VPS16) occurred de novo, supporting pathogenicity. INTERPRETATION:This study provides valuable insights into the genetic landscape of dystonia, underscores the utility of exome sequencing for diagnosis, substantiates several candidate genes, and expands the phenotypic spectrum of some genes to include prominent, sometimes isolated dystonia.

Parkinson’s disease (PD) is a neurodegenerative disorder with a high variability of age at onset, disease severity, and progression. This suggests that other factors, including genetic, environmental, or biological factors, are at play in PD. The loss of PINK1 causes a recessive form of PD and is typically fully penetrant; however, it features a wide range in disease onset, further supporting the existence of protective factors, endogenous or exogenous, to play a role. The loss of Pink1 in Drosophila melanogaster results in locomotion deficits, also observed in PINK1-related PD in humans. In flies, Pink1 deficiency induces defects in the ability to fly; nonetheless, around ten percent of the mutant flies are still capable of flying, indicating that advantageous factors affecting penetrance also exist in flies. Here, we aimed to identify the mechanisms underlying this reduced penetrance in Pink1-deficient flies. We performed genetic screening in pink1-mutant flies to identify RNA expression alterations affecting the flying ability. The most important biological processes involved were transcriptional and translational activities, endoplasmic reticulum (ER) regulation, and flagellated movement and microtubule organization. We validated two ER-related proteins, zonda and windbeutel, to positively affect the flying ability of Pink1-deficient flies. Thus, our data suggest that these processes are involved in the reduced penetrance and that influencing them may be beneficial for Pink1 deficiency.

Post-COVID syndrome (PCS) currently affects approximately 3-17% of people following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and has the potential to become a significant global health burden. PCS presents with various symptoms, and methods for improved PCS assessment are presently developed to guide therapy. Nevertheless, there are few mechanistic insights and treatment options. Here, we performed single-cell RNA transcriptomics on nasal biopsies from 33 patients suffering from PCS with mild, moderate, or severe symptoms. We identified 17 different cell clusters representing 12 unique cell populations, including all major epithelial cell types of the conducting airways and basal, secretory, and ciliated cells. Severe PCS was associated with decreased numbers of ciliated cells and the presence of immune cells. Ensuing inflammatory signaling upregulated TGFβ and induced an epithelial-mesenchymal transition, which led to the high abundance of basal cells and a mis-stratified epithelium. We confirmed the results in vitro using an air-liquid interface culture and validated TNFα as the causal inflammatory cytokine. In summary, our results show that one mechanism for sustained PCS is not through continued viral load, but through the presence of immune cells in nasal tissue leading to impaired mucosal barrier function and repeated infections. These findings could be further explored as a therapeutic option akin to other chronic inflammatory diseases by inhibiting the TNFα-TGFβ axis, restoring the nasal epithelium, and reducing respiratory tract-related infections.### Competing Interest StatementThe authors have declared no competing interest.

Dystonia due to pathogenic variants in the THAP1 gene (DYT-THAP1) shows variable expressivity and reduced penetrance of 50

Hintergrund Asthma is characterized by chronic inflammation, airway hyperresponsiveness, and episodic shortness of breath. Exacerbations of asthma symptoms are commonly induced by viruses, which promote acute pro-inflammatory responses. The nasal mucosa is the first barrier to such viral insults and comprises a heterogeneous epithelial cell mixture. Disturbances to normal mucosa formation are associated with worse asthma outcomes and more exacerbations. The pro-inflammatory cytokine tumor necrosis factor-alpha (TNFα) is increased during asthma exacerbations and has been associated with altered cellular differentiation processes. This study aimed to explore whether prolonged TNFα exposure affects the correctness of the airway mucosal layer.

Chemosensory especially olfactory dysfunction (OD) is common after COVID-19.The exact mechanism of this impairment is connected with molecular changes, which can be examined using single-cell RNA sequencing (SCS).The objective of this study was to evaluate cell populations,receptor expressions and analyze differential gene expressions leading to the identification of signaling pathways,in post-COVID patients (PCPs).

BackgroundPathogenic variants in several genes have been linked to genetic forms of isolated or combined dystonia. The phenotypic and genetic spectrum and the frequency of pathogenic variants in these genes have not yet been fully elucidated, neither in patients with dystonia nor with other, sometimes co-occurring movement disorders such as Parkinson's disease (PD).ObjectivesTo screen >2000 patients with dystonia or PD for rare variants in known dystonia-causing genes.MethodsWe screened 1207 dystonia patients from Germany (DysTract consortium), Spain, and South Korea, and 1036 PD patients from Germany for pathogenic variants using a next-generation sequencing gene panel. The impact on DNA methylation of KMT2B variants was evaluated by analyzing the gene's characteristic episignature.ResultsWe identified 171 carriers (109 with dystonia [9.0%]; 62 with PD [6.0%]) of 131 rare variants (minor allele frequency <0.005). A total of 52 patients (48 dystonia [4.0%]; four PD [0.4%, all with GCH1 variants]) carried 33 different (likely) pathogenic variants, of which 17 were not previously reported. Pathogenic biallelic variants in PRKRA were not found. Episignature analysis of 48 KMT2B variants revealed that only two of these should be considered (likely) pathogenic.ConclusionThis study confirms pathogenic variants in GCH1, GNAL, KMT2B, SGCE, THAP1, and TOR1A as relevant causes in dystonia and expands the mutational spectrum. Of note, likely pathogenic variants only in GCH1 were also found among PD patients. For DYT-KMT2B, the recently described episignature served as a reliable readout to determine the functional effect of newly identified variants. (c) 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Tenosynovial giant cell tumor (TGCT) is a rare type of tumor that originates from the synovium of joints and tendon sheaths. It is characterized by recurring genetic abnormalities, often involving the CSF1 gene. Common symptoms include pain and swelling, which are not specific to TGCT, so MRI and a pathological biopsy are needed for an accurate diagnosis. We report the case of a 45-year-old man who experienced painful swelling in his right hip for six months. Initially, this was diagnosed as Erdheim-Chester disease. However, whole exome sequencing (WES) and RNA-Sequencing revealed a CSF1::GAPDHP64 fusion, leading to a revised diagnosis of TGCT. The patient was treated with pegylated interferon and imatinib, which resulted in stable disease after three months. Single-cell transcriptome analysis identified seven distinct cell clusters, revealing that neoplastic cells expressing CSF1 attract macrophages. Analysis of ligand-receptor interactions showed significant communication between neoplastic cells and macrophages mediated by CSF1 and CSF1R. Our findings emphasize the importance of comprehensive molecular analysis in diagnosing and treating rare malignancies like TGCT.

IntroductionPrimarily driven by autoreactive B cells, pemphigus foliaceus (PF) is an uncommon autoimmune blistering skin disease of sporadic occurrence worldwide. However, PF reaches a prevalence of 3% in the endemic areas of Brazil, the highest ever registered for any autoimmune disease, which indicates environmental factors influencing the immune response in susceptible individuals. We aimed to provide insights into the immune repertoire of patients with PF living in the endemic region of the disease, compared to healthy individuals from the endemic region and a non-endemic area.MethodsWe characterized the B-cell repertoire in i) nontreated patients (n=5); ii) patients under immunosuppressive treatment (n=5); iii) patients in remission without treatment (n=6); and two control groups iv) from the endemic (n=6) and v) non-endemic areas in Brazil (n=4). We used total RNA extracted from peripheral blood mononuclear cells and performed a comprehensive characterization of the variable region of immunoglobulin heavy chain (IGH) in IgG and IgM using next-generation sequencing.ResultsCompared to individuals from a different area, we observed remarkably lower clonotype diversity in the B-cell immune repertoire of patients and controls from the endemic area (p < 0.02), suggesting that the immune repertoire in the endemic area is under geographically specific and intense environmental pressure. Moreover, we observed longer CDR3 sequences in patients, and we identified differential disease-specific usage of IGHV segments, including increased IGHV3-30 and decreased IGHV3-23 in patients with active disease (p < 0.04). Finally, our robust network analysis discovered clusters of CDR3 sequences uniquely observed in patients with PF.DiscussionOur results indicate that environmental factors, in addition to disease state, impact the characteristics of the repertoire. Our findings can be applied to further investigation of the environmental factors that trigger pemphigus and expand the knowledge for identifying new targeted and more effective therapies.

The Post-COVID Syndrome (PCS) currently affects approximately 3-17% of people after infection with the Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has the potential to become a significant global health burden. The Post-COVID Syndrome has various symptoms, and currently, methods for improved PCS scoring are developed to guide therapy. Yet mechanistic insights and treatment options are scarce. We thought to investigate individual nasal epithelial cells from patients with various levels of PCS intensity to gain mechanistic insights. We performed single-cell RNA sequencing (Signleron, Germany) from nasal biopsies samples (n=30) of clinically well-characterized patients with mild, moderate or severe PCS symptoms based on a recently published PCS score (Bahmer et al). Transcriptional profiles of 35.000 cells identified 18 different cell types, such as all major epithelial cell types of the conducting airways, including basal, secretory, and ciliated cells. In addition, we used enrichment analysis to link key molecules and biological pathways to disease severity. High PCS scores are associated with decreased levels of ciliated cells and an increase in the total number of immune cells. Significantly associated signaling pathways included NFkB and WNT signalling, and signal-response Reactome interactions between immune cells and epithelial cells were detected. An immune/epithelial cross-talk may decrease ciliated epithelial cells, which might interfere with a proper mucosal barrier function. These findings could be further investigated as a therapeutic option for preventing severe outcomes in symptomatic post-Covid patients.

BackgroundProtein synthesis is a tightly controlled process, involving a host of translation-initiation factors and microRNA-associated repressors. Variants in the translational regulator EIF2AK2 were first linked to neurodevelopmental-delay phenotypes, followed by their implication in dystonia. Recently, de novo variants in EIF4A2, encoding eukaryotic translation initiation factor 4A isoform 2 (eIF4A2), have been described in pediatric cases with developmental delay and intellectual disability. ObjectiveWe sought to characterize the role of EIF4A2 variants in dystonic conditions. MethodsWe undertook an unbiased search for likely deleterious variants in mutation-constrained genes among 1100 families studied with dystonia. Independent cohorts were screened for EIF4A2 variants. Western blotting and immunocytochemical studies were performed in patient-derived fibroblasts. ResultsWe report the discovery of a novel heterozygous EIF4A2 frameshift deletion (c.896_897del) in seven patients from two unrelated families. The disease was characterized by adolescence- to adulthood-onset dystonia with tremor. In patient-derived fibroblasts, eIF4A2 production amounted to only 50% of the normal quantity. Reduction of eIF4A2 was associated with abnormally increased levels of IMP1, a target of Ccr4-Not, the complex that interacts with eIF4A2 to mediate microRNA-dependent translational repression. By complementing the analyses with fibroblasts bearing EIF4A2 biallelic mutations, we established a correlation between IMP1 expression alterations and eIF4A2 functional dosage. Moreover, eIF4A2 and Ccr4-Not displayed significantly diminished colocalization in dystonia patient cells. Review of international databases identified EIF4A2 deletion variants (c.470_472del, c.1144_1145del) in another two dystonia-affected pedigrees. ConclusionsOur findings demonstrate that EIF4A2 haploinsufficiency underlies a previously unrecognized dominant dystonia-tremor syndrome. The data imply that translational deregulation is more broadly linked to both early neurodevelopmental phenotypes and later-onset dystonic conditions. & COPY; 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Pathogenic variants in THAP1 can cause dystonia with a penetrance of about 50 %. The underlying mechanisms are unknown and can be considered as means of endogenous disease protection. Since THAP1 encodes a transcription factor, drivers of this variability putatively act at the transcriptome level. Several transcriptome studies tried to elucidate THAP1 function in diverse cellular and mouse models, including mutation carrier-derived cells and iPSC-derived neurons, unveiling various differentially expressed genes and affected pathways. These include nervous system development, dopamine signalling, myelination, or cell-cell adhesion. A network diffusion analysis revealed mRNA splicing, mitochondria, DNA repair, and metabolism as significant pathways that may represent potential targets for therapeutic interventions.

De novo variants in the myelin regulatory factor (MYRF), a transcription factor involved in the differentiation of oligodendrocytes, have been linked recently to the cardiac and urogenital syndrome, while familiar variants are associated with nanophthalmos. Here, we report for the first time on a patient with a de novo stop-gain variant in MYRF (p.Q838*) associated with Scimitar syndrome, 46,XY partial gonadal dysgenesis (GD) and severe hyperopia. Since variants in MYRF have been described in both 46,XX and 46,XY GD, we assumed a role of MYRF in the early development of the bipotential gonad. We used publicly available single cell sequencing data of human testis and ovary from different developmental stages and analysed them for MYRF expression. We identified MYRF expression in the subset of coelomic epithelial cells at stages of gonadal ridge development in 46,XX and 46,XY individuals. Differential gene expression analysis revealed significantly upregulated genes. Within these, we identified CITED2 as a gene containing a MYRF binding site. It has been shown that Cited2−/− mice have gonadal defects in both testis and ovary differentiation, as well as defects in heart development and establishment of the left–right axis. This makes MYRF a potential candidate as an early regulator of gonadal and heart development via upregulation of the transcriptional cofactor CITED2.

Mutated and unmutated IgE and IgG play different and partly opposing roles in allergy development, but the mechanisms controlling their relative production are incompletely understood. Here, we analyzed the IgE-response in murine food allergy. Deep sequencing of the complementary-determining region (CDR) repertoires indicated that an ongoing unmutated extrafollicular IgE response coexists with a germinal center response, even after long-lasting allergen challenges. Despite overall IgG1-dominance, a significant proportion of clonotypes contained several-fold more IgE than IgG1. Clonotypes with differential bias to either IgE or IgG1 showed distinct hypermutation and clonal expansion. Hypermutation rates were associated with different physiochemical binding properties of individual B-cell receptors (BCR). Increasing BCR signaling strength inhibited class switching from IgG1 to IgE in vitro, preferentially constraining IgE formation. These data indicate that antigen-binding properties of individual BCRs determine differential IgE hypermutation and IgE versus IgG1 production on the level of single B-cell clones.

The THAP1 gene encodes a transcription factor, and pathogenic variants cause a form of autosomal dominant, isolated dystonia (DYT‐THAP1) with reduced penetrance. Factors underlying both reduced penetrance and the disease mechanism of DYT‐THAP1 are largely unknown.

Coronavirus disease 2019 (COVID-19) can damage cerebral small vessels and cause neurological symptoms. Here we describe structural changes in cerebral small vessels of patients with COVID-19 and elucidate potential mechanisms underlying the vascular pathology. In brains of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals and animal models, we found an increased number of empty basement membrane tubes, so-called string vessels representing remnants of lost capillaries. We obtained evidence that brain endothelial cells are infected and that the main protease of SARS-CoV-2 (M pro ) cleaves NEMO, the essential modulator of nuclear factor-κB. By ablating NEMO, M pro induces the death of human brain endothelial cells and the occurrence of string vessels in mice. Deletion of receptor-interacting protein kinase (RIPK) 3, a mediator of regulated cell death, blocks the vessel rarefaction and disruption of the blood–brain barrier due to NEMO ablation. Importantly, a pharmacological inhibitor of RIPK signaling prevented the M pro -induced microvascular pathology. Our data suggest RIPK as a potential therapeutic target to treat the neuropathology of COVID-19.