Advances in understanding the pathophysiology of facioscapulohumeral dystrophy (FSHD) have led to several therapeutic approaches entering clinical trials and an increased need to develop biomarkers of disease activity and progression. Multiple prior studies have shown early elevation of RNAs encoding components of the complement pathways and relatively widespread activated complement complexes by immunodetection in FSHD muscle. The current study tested plasma from two independent cohorts of FSHD and control subjects and found elevated complement components in both FSHD cohorts. Combining subjects from both cohorts identified complement factors that best distinguished FSHD and controls. Within the FSHD group, a subset of subjects showed elevation in multiple complement components. Together these findings suggest the need for future studies to determine whether measurements of complement activation can be used as a non-invasive measurement of FSHD disease activity, progression and/or response to therapies. In addition, with the ongoing expansion of complement therapeutic approaches, consideration for precision-based targeting of this pathway is appropriate.

Increasing evidences suggest that mitochondrial dysfunction is implicated in diseases and aging, and whole-genome sequencing (WGS) is the most unbiased method in analyzing the mitochondrial genome (mtDNA). However, the genetic landscape of mtDNA in the Chinese population has not been fully examined. Here, we described the genetic landscape of mtDNA using WGS data from Chinese individuals (n = 3241). We identified 3892 mtDNA variants, of which 3349 (86%) were rare variants. Interestingly, we observed a trend toward extreme heterogeneity of mtDNA variants. Our study observed a distinct purifying selection on mtDNA, which inhibits the accumulation of harmful heteroplasmies at the individual level: (1) mitochondrial dN/dS ratios were much <1; (2) the dN/dS ratio of heteroplasmies was higher than homoplasmies; (3) heteroplasmies had more indels and predicted deleterious variants than homoplasmies. Furthermore, we found that haplogroup M (20.27%) and D (20.15%) had the highest frequencies in the Chinese population, followed by B (18.51%) and F (16.45%). The number of variants per individual differed across haplogroup groups, with a higher number of homoplasmies for the M lineage. Meanwhile, mtDNA copy number was negatively correlated with age but positively correlated with the female sex. Finally, we developed an mtDNA variation database of Chinese populations called MTCards (http://genemed.tech/mtcards/) to facilitate the query of mtDNA variants in this study. In summary, these findings contribute to different aspects of understanding mtDNA, providing a better understanding of the genetic basis of mitochondrial-related diseases.

Thyroid dysfunction is a common endocrine disease measured by thyroid-stimulating hormone (TSH) level. Although >70 genetic loci associated with TSH have been reported through genome-wide association studies (GWASs), the variants can only explain a small fraction of the thyroid function heritability. To identify novel candidate genes for thyroid function, we conducted the first large-scale transcriptome-wide association study (TWAS) for thyroid function using GWAS-summary data for TSH levels in up to 119 715 individuals combined with precomputed gene expression weights of six panels from four tissue types. The candidate genes identified by TWAS were further validated by TWAS replication and gene expression profiles. We identified 74 conditionally independent genes significantly associated with thyroid function, such as PDE8B (P = 1.67 × 10-282), PDE10A (P = 7.61 × 10-119), NR3C2 (P = 1.50 × 10-92) and CAPZB (P = 3.13 × 10-79). After TWAS replication using UKBB datasets, 26 genes were replicated for significant associations with thyroid-relevant diseases/traits. Among them, 16 genes were causal for their associations to thyroid-relevant diseases/traits and further validated in differential expression analyses, including two novel genes (MFSD6 and RBM47) that did not implicate in previous GWASs. Enrichment analyses detected several pathways associated with thyroid function, such as the cAMP signaling pathway (P = 7.27 × 10-4), hemostasis (P = 3.74 × 10-4), and platelet activation, signaling and aggregation (P = 9.98 × 10-4). Our study identified multiple candidate genes and pathways associated with thyroid function, providing novel clues for revealing the genetic mechanisms of thyroid function and disease.

Hereditary spastic paraplegia (HSP) is a disease in which dieback degeneration of corticospinal tracts, accompanied by axonal swellings, leads to gait deficiencies. SPG4-HSP, the most common form of the disease, results from mutations of human spastin gene (SPAST), which is the gene that encodes spastin, a microtubule-severing protein. The lack of a vertebrate model that recapitulates both the etiology and symptoms of SPG4-HSP has stymied the development of effective therapies for the disease. hSPAST-C448Y mice, which express human mutant spastin at the ROSA26 locus, display corticospinal dieback and gait deficiencies but not axonal swellings. On the other hand, mouse spastin gene (Spast)-knockout (KO) mice display axonal swellings but not corticospinal dieback or gait deficiencies. One possibility is that reduced spastin function, resulting in axonal swellings, is not the cause of the disease but exacerbates the toxic effects of the mutant protein. To explore this idea, Spast-KO and hSPAST-C448Y mice were crossbred, and the offspring were compared with the parental lines via histological and behavioral analyses. The crossbred animals displayed axonal swellings as well as earlier onset, worsened gait deficiencies and corticospinal dieback compared with the hSPAST-C448Y mouse. These results, together with observations on changes in histone deacetylases 6 and tubulin modifications in the axon, indicate that each of these three transgenic mouse lines is valuable for investigating a different component of the disease pathology. Moreover, the crossbred mice are the best vertebrate model to date for testing potential therapies for SPG4-HSP.

BACKGROUND: Higher birthweight is associated with higher adult body mass index (BMI). Alleles that predispose to greater adult adiposity might act in fetal life to increase fetal growth and birthweight. Whether there are fetal effects of recently identified adult metabolically favorable adiposity alleles on birthweight is unknown. AIM: We aimed to test the effect on birthweight of fetal genetic predisposition to higher metabolically favorable adult adiposity and compare that with the effect of fetal genetic predisposition to higher adult BMI. METHODS: We used published genome wide association study data (n = upto 406 063) to estimate fetal effects on birthweight (adjusting for maternal genotype) of alleles known to raise metabolically favorable adult adiposity or BMI. We combined summary data across single nucleotide polymorphisms (SNPs) with random effects meta-analyses. We performed weighted linear regression of SNP-birthweight effects against SNP-adult adiposity effects to test for a dose-dependent association. RESULTS: Fetal genetic predisposition to higher metabolically favorable adult adiposity and higher adult BMI were both associated with higher birthweight (3 g per effect allele (95% CI: 1-5) averaged over 14 SNPs; P = 0.002; 0.5 g per effect allele (95% CI: 0-1) averaged over 76 SNPs; P = 0.042, respectively). SNPs with greater effects on metabolically favorable adiposity tended to have greater effects on birthweight (R2 = 0.2912, P = 0.027). There was no dose-dependent association for BMI (R2 = -0.0019, P = 0.602). CONCLUSIONS: Fetal genetic predisposition to both higher adult metabolically favorable adiposity and BMI is associated with birthweight. Fetal effects of metabolically favorable adiposity-raising alleles on birthweight are modestly proportional to their effects on future adiposity, but those of BMI-raising alleles are not.

Understanding the genetic effects on non-coding RNA (ncRNA) expression facilitates functional characterization of disease-associated genetic loci. Among several classes of ncRNAs, microRNAs (miRNAs) are key post-transcriptional gene regulators. Despite its biological importance, previous studies on the genetic architecture of miRNA expression focused mostly on the European individuals, underrepresented in other populations. Here, we mapped miRNA expression quantitative trait loci (miRNA-eQTL) for 343 miRNAs in 141 Japanese using small RNA sequencing and whole-genome sequencing, identifying 1275 cis-miRNA-eQTL variants for 40 miRNAs (false discovery rate < 0.2). Of these, 25 miRNAs having eQTL were unreported in the European studies, including 5 miRNAs with their lead variant monomorphic in the European populations, which demonstrates the value of miRNA-eQTL analysis in diverse ancestral populations. MiRNAs with eQTL effect showed allele-specific expression (ASE; e.g. miR-146a-3p), and ASE analysis further detected cis-regulatory variants not captured by the conventional miRNA-eQTL mapping (e.g. miR-933). We identified a copy number variation associated with miRNA expression (e.g. miR-570-3p, P = 7.2 × 10-6), which contributes to a more comprehensive landscape of miRNA-eQTLs. To elucidate a post-transcriptional modification in miRNAs, we created a catalog of miRNA-editing sites, including 10 canonical and 6 non-canonical sites. Finally, by integrating the miRNA-eQTLs and Japanese genome-wide association studies of 25 complex traits (mean n = 192 833), we conducted a transcriptome-wide association study, identifying miR-1908-5p as a potential mediator for adult height, colorectal cancer and type 2 diabetes (P < 9.1 × 10-5). Our study broadens the population diversity in ncRNA-eQTL studies and contributes to functional annotation of disease-associated loci found in non-European populations.

Familial dysautonomia (FD) is an autosomal recessive neurodegenerative disease caused by a splicing mutation in the gene encoding Elongator complex protein 1 (ELP1, also known as IKBKAP). This mutation results in tissue-specific skipping of exon 20 with a corresponding reduction of ELP1 protein, predominantly in the central and peripheral nervous system. Although FD patients have a complex neurological phenotype caused by continuous depletion of sensory and autonomic neurons, progressive visual decline leading to blindness is one of the most problematic aspects of the disease, as it severely affects their quality of life. To better understand the disease mechanism as well as to test the in vivo efficacy of targeted therapies for FD, we have recently generated a novel phenotypic mouse model, TgFD9; IkbkapΔ20/flox. This mouse exhibits most of the clinical features of the disease and accurately recapitulates the tissue-specific splicing defect observed in FD patients. Driven by the dire need to develop therapies targeting retinal degeneration in FD, herein, we comprehensively characterized the progression of the retinal phenotype in this mouse, and we demonstrated that it is possible to correct ELP1 splicing defect in the retina using the splicing modulator compound (SMC) BPN-15477.

A highly evolutionarily conserved myeloid ecotropic viral integration site 1 (MEIS1) intronic region is strongly associated with restless legs syndrome (RLS) and insomnia. To understand its regulatory function, we dissected the region by analyzing chromatin accessibility, enhancer-promoter contacts, DNA methylation and expression quantitative trait locus (eQTLs) in different human neural cell types and tissues. We observed specific activity with respect to cell type and developmental maturation, indicating a prominent role for distinct highly conserved intronic elements in forebrain inhibitory neuron differentiation. Two elements were hypomethylated in neural cells with higher MEIS1 expression, suggesting a role of enhancer demethylation in gene regulation. MEIS1 eQTLs showed a striking modular chromosomal distribution, with forebrain eQTLs clustering in intron 8/9. Clustered regularly interspersed short palindromic repeats interference targeting of individual elements in this region attenuated MEIS1 expression, revealing a complex regulatory interplay of distinct elements. In summary, we found that MEIS1 regulation is organized in a modular pattern. Disease-associated intronic regulatory elements control MEIS1 expression with cell type and maturation stage specificity, particularly in the inhibitory neuron lineage. The precise spatiotemporal activity of these elements likely contributes to the pathogenesis of insomnia and RLS.

Cerebral spinal fluid (CSF) is a promising biospecimen for the detection of central nervous system biomarkers to monitor therapeutic efficacy at the cellular level in neurological diseases. Spinal muscular atrophy (SMA) patients receiving intrathecal antisense oligonucleotide (nusinersen) therapy tend to show improved motor function, but the treatment effect on cellular health remains unknown. The objective of this study was to assess the potential of extracellular RNAs and microRNAs in SMA patient CSF as indicators of neuron and glial health following nusinersen treatment. Extracellular RNA analysis of CSF samples revealed ongoing cellular stress related to inflammation and glial differentiation, even after treatment administration. Downregulated microRNA expression associated with SMA-specific or general motor neuron dysfunction in animal and cellular models, tended to increase in nusinersen-treated patient CSF samples and correlated with SMA Type 1 and 2 motor functioning improvements. However, miR-146a, known to be upregulated in SMA-induced pluripotent stem cell (iPSC)-derived astrocytes, showed increased expression in nusinersen-treated CSF samples. We then used mRNA sequencing and multi-electrode arrays to assess the transcriptional and functional effects of miR-146a on healthy and SMA iPSC-derived motor neurons. miR-146a treatment on iPSC-derived motor neurons led to a downregulation of extracellular matrix genes associated with synaptic perineuronal net and alterations in spontaneous electrophysiological activity. Altogether, this study suggests that extracellular RNAs and microRNAs may serve as useful biomarkers to monitor cellular health during nusinersen treatment. Moreover, these data highlight the importance of addressing astrocyte health and response to nusinersen in SMA pathogenesis and treatment strategies.

X-linked lissencephaly with abnormal genitalia (XLAG) and developmental epileptic encephalopathy-1 (DEE1) are caused by mutations in the Aristaless-related homeobox (ARX) gene, which encodes a transcription factor responsible for brain development. It has been unknown whether the phenotypically diverse XLAG and DEE1 phenotypes may converge on shared pathways. To address this question, a label-free quantitative proteomic approach was applied to the neonatal brain of Arx knockout (ArxKO/Y) and knock-in polyalanine (Arx(GCG)7/Y) mice that are respectively models for XLAG and DEE1. Gene ontology and protein-protein interaction analysis revealed that cytoskeleton, protein synthesis and splicing control are deregulated in an allelic-dependent manner. Decreased α-tubulin content was observed both in Arx mice and Arx/alr-1(KO) Caenorhabditis elegans ,and a disorganized neurite network in murine primary neurons was consistent with an allelic-dependent secondary tubulinopathy. As distinct features of Arx(GCG)7/Y mice, we detected eIF4A2 overexpression and translational suppression in cortex and primary neurons. Allelic-dependent differences were also established in alternative splicing (AS) regulated by PUF60 and SAM68. Abnormal AS repertoires in Neurexin-1, a gene encoding multiple pre-synaptic organizers implicated in synaptic remodelling, were detected in Arx/alr-1(KO) animals and in Arx(GCG)7/Y epileptogenic brain areas and depolarized cortical neurons. Consistent with a conserved role of ARX in modulating AS, we propose that the allelic-dependent secondary synaptopathy results from an aberrant Neurexin-1 repertoire. Overall, our data reveal alterations mirroring the overlapping and variant effects caused by null and polyalanine expanded mutations in ARX. The identification of these effects can aid in the design of pathway-guided therapy for ARX endophenotypes and NDDs with overlapping comorbidities.

Refractive errors are associated with a range of pathological conditions, such as myopic maculopathy and glaucoma, and are highly heritable. Studies of missense and putative loss of function (pLOF) variants identified via whole exome sequencing (WES) offer the prospect of directly implicating potentially causative disease genes. We performed a genome-wide association study for refractive error in 51 624 unrelated adults, of European ancestry, aged 40-69 years from the UK and genotyped using WES. After testing 29 179 pLOF and 495 263 missense variants, 1 pLOF and 18 missense variants in 14 distinct genomic regions were taken forward for fine-mapping analysis. This yielded 19 putative causal variants of which 18 had a posterior inclusion probability >0.5. Of the 19 putative causal variants, 12 were novel discoveries. Specific variants were associated with a more myopic refractive error, while others were associated with a more hyperopic refractive error. Association with age of onset of spectacle wear (AOSW) was examined in an independent validation sample (38 100 early AOSW cases and 74 243 controls). Of 11 novel variants that could be tested, 8 (73%) showed evidence of association with AOSW status. This work identified COL4A4 and ATM as novel candidate genes associated with refractive error. In addition, novel putative causal variants were identified in the genes RASGEF1, ARMS2, BMP4, SIX6, GSDMA, GNGT2, ZNF652 and CRX. Despite these successes, the study also highlighted the limitations of community-based WES studies compared with high myopia case-control WES studies.

To highlight the genetic architecture for epigenetic aging, McCartney et al. recently identified 137 significant single-nucleotide polymorphisms based on genome-wide association study (GWAS) meta-analyses of four epigenetic clocks and two epigenetic surrogate markers. However, none Asian ancestry studies have been included in this or previous meta-analyses. I performed a GWAS on blood DNA methylation (DNAm) levels of 2309 Taiwan Biobank (TWB) participants. Owing to the fact that the sample size of an individual GWAS of DNAm data is still not large, I adopted the 'prioritized subset analysis' (PSA) method to boost the power of a GWAS. The four epigenetic clocks and the two epigenetic surrogate markers were investigated, respectively. I replicated 21 out of the 137 aging-associated genetic loci by applying the PSA method to the TWB DNAm data. Moreover, I identified five novel loci, including rs117530284 that was associated with the 'epigenetic age acceleration' (EAA) according to Lu et al.'s GrimAge (called 'GrimEAA'). Considering 16 covariates (sex, BMI, smoking status, drinking status, regular exercise, educational attainment and the first 10 ancestry principal components), each 'A' allele of rs117530284 in the IBA57 gene was found to be associated with a 1.5943-year GrimEAA (95% confidence interval = [1.0748, 2.1138]). IBA57 is a protein coding gene and is associated with multiple mitochondrial dysfunctions syndromes. A decline in mitochondrial activity and quality is associated with aging and many age-related diseases. This is one of the first DNAm GWAS for individuals of Asian ancestry.

The purpose of our study is to understand the impact of a partial dynamin-related protein 1 (Drp1) on cognitive behavior, mitophagy, autophagy and mitochondrial and synaptic activities in transgenic Tau mice in Alzheimer's disease (AD). Our laboratory reported increased levels of amyloid-beta (Aβ) and phosphorylated Tau (P-Tau) and reported that abnormal interactions between Aβ and Drp1, P-Tau and Drp1 induced increased mitochondrial fragmentation and reduced fusion and synaptic activities in AD. These abnormal interactions result in the proliferation of dysfunctional mitochondria in AD neurons. Recent research on mitochondria revealed that fission protein Drp1 is largely implicated in mitochondrial dynamics in AD. To determine the impact of reduced Drp1 in AD, we recently crossed transgenic Tau mice with Drp1 heterozygote knockout (Drp1+/-) mice and generated double mutant (P301LDrp1+/-) mice. In the current study, we assessed the cognitive behavior, mRNA and protein levels of mitophagy, autophagy, mitochondrial biogenesis, dynamics and synaptic genes, mitochondrial morphology and mitochondrial function and dendritic spines in Tau mice relative to double mutant mice. When compared with Tau mice, double mutant mice did better on the Morris Maze (reduced latency to find hidden platform, increased swimming speed and time spent on quadrant) and rotarod (stayed a longer period of time) tests. Both mRNA- and protein-level autophagy, mitophagy, mitochondrial biogenesis and synaptic proteins were increased in double mutant mice compared with Tau (P301L) mice. Dendritic spines were significantly increased; mitochondrial number was reduced and length was increased in double mutant mice. Based on these observations, we conclude that reduced Drp1 is beneficial in a symptomatic-transgenic Tau (P301L) mice.

The SLC25A26 gene encodes a mitochondrial inner membrane carrier that transports S-adenosylmethionine (SAM) into the mitochondrial matrix in exchange for S-adenosylhomocysteine (SAH). SAM is the predominant methyl-group donor for most cellular methylation processes, of which SAH is produced as a by-product. Pathogenic, biallelic SLC25A26 variants are a recognized cause of mitochondrial disease in children, with a severe neonatal onset caused by decreased SAM transport activity. Here, we describe two, unrelated adult cases, one of whom presented with recurrent episodes of severe abdominal pain and metabolic decompensation with lactic acidosis. Both patients had exercise intolerance and mitochondrial myopathy associated with biallelic variants in SLC25A26, which led to marked respiratory chain deficiencies and mitochondrial histopathological abnormalities in skeletal muscle that are comparable to those previously described in early-onset cases. We demonstrate using both mouse and fruit fly models that impairment of SAH, rather than SAM, transport across the mitochondrial membrane is likely the cause of this milder, late-onset phenotype. Our findings associate a novel pathomechanism with a known disease-causing protein and highlight the quests of precision medicine in optimizing diagnosis, therapeutic intervention and prognosis.

Renal tract defects and autism spectrum disorder (ASD) deficits represent the phenotypic core of the 19q12 deletion syndrome caused by the loss of one copy of the TSHZ3 gene. Although a proportion of Tshz3 heterozygous (Tshz3+/lacZ) mice display ureteral defects, no kidney defects have been reported in these mice. The purpose of this study was to characterize the expression of Tshz3 in adult kidney as well as the renal consequences of embryonic haploinsufficiency of Tshz3 by analyzing the morphology and function of Tshz3 heterozygous adult kidney. Here, we described Tshz3 expression in the smooth muscle and stromal cells lining the renal pelvis, the papilla and glomerular endothelial cells (GEnCs) of the adult kidney as well as in the proximal nephron tubules in neonatal mice. Histological analysis showed that Tshz3+/lacZ adult kidney had an average of 29% fewer glomeruli than wild-type kidney. Transmission electron microscopy of Tshz3+/lacZ glomeruli revealed a reduced thickness of the glomerular basement membrane and a larger foot process width. Compared to wild type, Tshz3+/lacZ mice showed lower blood urea, phosphates, magnesium and potassium at 2 months of age. At the molecular level, transcriptome analysis identified differentially expressed genes related to inflammatory processes in Tshz3+/lacZ compare to wild-type (control) adult kidneys. Lastly, analysis of the urinary peptidome revealed 33 peptides associated with Tshz3+/lacZ adult mice. These results provide the first evidence that in the mouse Tshz3 haploinsufficiency leads to cellular, molecular and functional abnormalities in the adult mouse kidney.

Acute pancreatitis (AP) is widely recognized to be an inflammation-related disease, in which HDAC was upregulated. The anti-inflammatory role of suberoylanilide hydroxamic acid (SAHA), a HDAC inhibitor, has been documented. In this context, this research was implemented to figure out whether SAHA manipulated inflammation in AP. Subsequent to induction of AP mouse model, HDAC5 expression was detected. The binding of HDAC5 and SLIT2 was detected by Co-Immunoprecipitation and Chromatin immunoprecipitation assays. SAHA treatment and gain- and loss-of-function approaches were used in AP mice and lipopolysaccharide (LPS)-induced pancreatic acinar cells. In mice, biochemical methods were implemented to measure activities of pancreatic lipase, trypsin, myeloperoxidase (MPO) and pancreatic edema, TUNEL staining to determine pancreatic cell apoptosis, and flow cytometry to assess the total number of leukocytes and neutrophils in pancreas. In pancreatic acinar cells, CCK-8 was performed to evaluate cell viability. HDAC5 exhibited overexpression in AP mice. Mechanical analysis showed that HDAC5 facilitated SLIT2 deacetylation to downregulate SLIT2, thus activating Akt/β-catenin pathway in pancreatic acinar cells. SAHA treatment, HDAC5 silencing or SLIT2 overexpression diminished inflammation in AP in vivo and in vitro. SAHA treatment, HDAC5 silencing or SLIT2 overexpression reduced activities of pancreatic lipase, trypsin, MPO, pancreatic edema and cell apoptosis in AP mice as well as elevated viability of LPS-induced pancreatic acinar cells. SAHA might exert anti-inflammatory effects in AP mice via HDAC5/SLIT2/Akt/β-catenin axis.

The multi-systemic genetic disorder tuberous sclerosis complex (TSC) impacts multiple neurodevelopmental processes including neuronal morphogenesis, neuronal migration, myelination and gliogenesis. These alterations contribute to the development of cerebral cortex abnormalities and malformations. Although TSC is caused by mTORC1 hyperactivation, cognitive and behavioral impairments are not improved through mTORC1 targeting, making the study of the downstream effectors of this complex important for understanding the mechanisms underlying TSC. As mTORC1 has been shown to promote the activity of the transcriptional co-activator Yap, we hypothesized that altered Yap/Taz signaling contributes to the pathogenesis of TSC. We first observed that the levels of Yap/Taz are increased in human cortical tuber samples and in embryonic cortices of Tsc2 conditional knockout (cKO) mice. Next, to determine how abnormal upregulation of Yap/Taz impacts the neuropathology of TSC, we deleted Yap/Taz in Tsc2 cKO mice. Importantly, Yap/Taz/Tsc2 triple conditional knockout (tcKO) animals show reduced cortical thickness and cortical neuron cell size, despite the persistence of high mTORC1 activity, suggesting that Yap/Taz play a downstream role in cytomegaly. Furthermore, Yap/Taz/Tsc2 tcKO significantly restored cortical and hippocampal lamination defects and reduced hippocampal heterotopia formation. Finally, the loss of Yap/Taz increased the distribution of myelin basic protein in Tsc2 cKO animals, consistent with an improvement in myelination. Overall, our results indicate that targeting Yap/Taz lessens the severity of neuropathology in a TSC animal model. This study is the first to implicate Yap/Taz as contributors to cortical pathogenesis in TSC and therefore as potential novel targets in the treatment of this disorder.

K63-linked polyubiquitination of the neurodegenerative disease-associated misfolded protein copper-zinc superoxide dismutase 1 (SOD1) is associated with the formation of inclusion bodies. Highly expressed E3 ligase Smad ubiquitylation regulatory factor 1 (Smurf1) promotes cellular homeostasis through the enhanced capability of aggregate degradation. However, it is not well explored the role of Smurf1 in the dynamics of SOD1 aggresomes. In this study, we report that Smurf1 promotes the recruitment of SOD1 to form aggresomes. Mechanistically, Smurf1 interacts with mutant SOD1 to promote aggresome formation by modification of its K63-linked polyubiquitination. Moreover, overexpressed Smurf1 enhances mutant SOD1 aggresome formation and autophagic degradation to prevent cell death. Thus, our data suggest that Smurf1 plays an important role in attenuating protein misfolding-induced cell toxicity by both driving the sequestration of misfolded SOD1 into aggresomes and autophagic degradation.

Osteoarthritis is a prevalent joint disease and a major cause of disability worldwide with no curative therapy. Development of disease-modifying therapies requires a better understanding of the molecular mechanisms underpinning disease. A hallmark of osteoarthritis is cartilage degradation. To define molecular events characterizing osteoarthritis at the whole transcriptome level, we performed deep RNA sequencing in paired samples of low- and high-osteoarthritis grade knee cartilage derived from 124 patients undergoing total joint replacement. We detected differential expression between low- and high-osteoarthritis grade articular cartilage for 365 genes and identified a 38-gene signature in osteoarthritis cartilage by replicating our findings in an independent dataset. We also found differential expression for 25 novel long non-coding RNA genes (lncRNAs) and identified potential lncRNA interactions with RNA-binding proteins in osteoarthritis. We assessed alterations in the relative usage of individual gene transcripts and identified differential transcript usage for 82 genes, including ABI3BP, coding for an extracellular matrix protein, AKT1S1, a negative regulator of the mTOR pathway and TPRM4, coding for a transient receptor potential channel. We further assessed genome-wide differential splicing, for the first time in osteoarthritis, and detected differential splicing for 209 genes, which were enriched for extracellular matrix, proteoglycans and integrin surface interactions terms. In the largest study of its kind in osteoarthritis, we find that isoform and splicing changes, in addition to extensive differences in both coding and non-coding sequence expression, are associated with disease and demonstrate a novel layer of genomic complexity to osteoarthritis pathogenesis.

Prostate cancer is among the most common cancers in men, with a large fraction of the individual risk attributable to heritable factors. A majority of the diagnosed cases does not lead to a lethal disease, and hence biological markers that can distinguish between indolent and fatal forms of the disease are of great importance for guiding treatment decisions. Although over 300 genetic variants are known to be associated with prostate cancer risk, few have been associated with the risk of an aggressive disease. One such variant is rs77559646 located in ANO7. This variant has a dual function. It constitutes a missense mutation in the short isoform of ANO7 and a splice region mutation in full-length ANO7. In this study, we have analyzed the impact of the variant allele of rs77559646 on ANO7 mRNA splicing using a minigene splicing assay and by performing splicing analysis with the tools IRFinder (intron retention finder), rMATS (replicate multivariate analysis of transcript splicing) and LeafCutter on RNA sequencing data from prostate tissue of six rs77559646 variant allele carriers and 43 non-carriers. The results revealed a severe disruption of ANO7 mRNA splicing in rs77559646 variant allele carriers. Immunohistochemical analysis of prostate samples from patients homozygous for the rs77559646 variant allele demonstrated a loss of apically localized ANO7 protein. Our study is the first to provide a mechanistic explanation for the impact of a prostate cancer risk SNP on ANO7 protein production. Furthermore, the rs77559646 variant is the first known germline loss-of-function mutation described for ANO7. We suggest that loss of ANO7 contributes to prostate cancer progression.

BACKGROUND: FCGR2A binds antibody-antigen complexes to regulate the abundance of circulating and deposited complexes along with downstream immune and autoimmune responses. Although the abundance of FCRG2A may be critical in immune-mediated diseases, little is known about whether its surface expression is regulated through cis genomic elements and non-coding variants. In the current study, we aimed to characterize the regulation of FCGR2A expression, the impact of genetic variation and its association with autoimmune disease. METHODS: We applied CRISPR-based interference and editing to scrutinize 1.7 Mb of open chromatin surrounding the FCGR2A gene to identify regulatory elements. Relevant transcription factors (TFs) binding to these regions were defined through public databases. Genetic variants affecting regulation were identified using luciferase reporter assays and were verified in a cohort of 1996 genotyped healthy individuals using flow cytometry. RESULTS: We identified a complex proximal region and five distal enhancers regulating FCGR2A. The proximal region split into subregions upstream and downstream of the transcription start site, was enriched in binding of inflammation-regulated TFs, and harbored a variant associated with FCGR2A expression in primary myeloid cells. One distal enhancer region was occupied by CCCTC-binding factor (CTCF) whose binding site was disrupted by a rare genetic variant, altering gene expression. CONCLUSIONS: The FCGR2A gene is regulated by multiple proximal and distal genomic regions, with links to autoimmune disease. These findings may open up novel therapeutic avenues where fine-tuning of FCGR2A levels may constitute a part of treatment strategies for immune-mediated diseases.

Recent studies have demonstrated a relevant role of the host genetics in the coronavirus disease 2019 (COVID-19) prognosis. Most of the 7000 rare diseases described to date have a genetic component, typically highly penetrant. However, this vast spectrum of genetic variability remains yet unexplored with respect to possible interactions with COVID-19. Here, a mathematical mechanistic model of the COVID-19 molecular disease mechanism has been used to detect potential interactions between rare disease genes and the COVID-19 infection process and downstream consequences. Out of the 2518 disease genes analyzed, causative of 3854 rare diseases, a total of 254 genes have a direct effect on the COVID-19 molecular disease mechanism and 207 have an indirect effect revealed by a significant strong correlation. This remarkable potential of interaction occurs for >300 rare diseases. Mechanistic modeling of COVID-19 disease map has allowed a holistic systematic analysis of the potential interactions between the loss of function in known rare disease genes and the pathological consequences of COVID-19 infection. The results identify links between disease genes and COVID-19 hallmarks and demonstrate the usefulness of the proposed approach for future preventive measures in some rare diseases.

In the framework of the UK 100 000 Genomes Project, we investigated the genetic origin of a previously undescribed recessive dermatological condition, which we named LIPHAK (LTV1-associated Inflammatory Poikiloderma with Hair abnormalities and Acral Keratoses), in four affected individuals from two UK families of Pakistani and Indian origins, respectively. Our analysis showed that only one gene, LTV1, carried rare biallelic variants that were shared in all affected individuals, and specifically they bore the NM_032860.5:c.503A > G, p.(Asn168Ser) change, found homozygously in all of them. In addition, high-resolution homozygosity mapping revealed the presence of a small 652-kb stretch on chromosome 6, encompassing LTV1, that was haploidentical and common to all affected individuals. The c.503A > G variant was predicted by in silico tools to affect the correct splicing of LTV1's exon 5. Minigene-driven splicing assays in HEK293T cells and in a skin sample from one of the patients confirmed that this variant was indeed responsible for the creation of a new donor splice site, resulting in aberrant splicing and in a premature termination codon in exon 6 of this gene. LTV1 encodes one of the ribosome biogenesis factors that promote the assembly of the small (40S) ribosomal subunit. In yeast, defects in LTV1 alter the export of nascent ribosomal subunits to the cytoplasm; however, the role of this gene in human pathology is unknown to date. Our data suggest that LIPHAK could be a previously unrecognized ribosomopathy.

Frataxin (FXN) deficiency is responsible for Friedreich's ataxia (FRDA) in which, besides the characteristic features of spinocerebellar ataxia, two thirds of patients develop hypertrophic cardiomyopathy that often progresses to heart failure and premature death. Different mechanisms might underlie FRDA pathogenesis. Among them, the role of miRNAs deserves investigations. We carried out an miRNA PCR-array analysis of plasma samples of early-, intermediate- and late-onset FRDA groups, defining a set of 30 differentially expressed miRNAs. Hsa-miR223-3p is the only miRNA shared between the three patient groups and appears upregulated in all of them. The up-regulation of hsa-miR223-3p was further validated in all enrolled patients (n = 37, Fc = +2.3; P < 0.0001). Using a receiver operating characteristic curve analysis, we quantified the predictive value of circulating hsa-miR223-3p for FRDA, obtaining an area under the ROC curve value of 0.835 (P < 0.0001) for all patients. Interestingly, we found a significant positive correlation between hsa-miR223-3p expression and cardiac parameters in typical FRDA patients (onset < 25 years). Moreover, a significant negative correlation between hsa-miR223-3p expression and HAX-1 (HCLS1-associated protein X-1) at mRNA and protein level was observed in all FRDA patients. In silico analyses suggested HAX-1 as a target gene of hsa-miR223-3p. Accordingly, we report that HAX-1 is negatively regulated by hsa-miR223-3p in cardiomyocytes (AC16) and neurons (SH-SY5Y), which are critically affected cell types in FRDA. This study describes for the first time the association between hsa-miR223-3p and HAX-1 expression in FRDA, thus supporting a potential role of this microRNA as non-invasive epigenetic biomarker for FRDA.

Tubulin-associated unit (Tau) is a microtubule-associated protein, whose abnormal phosphorylation and deposition in the brain characterizes a range of neurodegenerative diseases called tauopathies. Recent clinical (post-mortem) and pre-clinical evidence suggests that Huntington's disease (HD), an autosomal dominant neurodegenerative disorder, could be considered as a tauopathy. Studies have found the presence of hyperphosphorylated tau, altered tau isoform ratio and aggregated tau in HD brains. However, little is known about the implication of tau in the development of HD pathophysiology, which includes motor, cognitive and affective symptoms. To shine a light on the involvement of tau in HD, our present study aimed at (i) knocking out tau expression and (ii) expressing a transgene encoding mutant human tau in the R6/1 mouse model of HD. We hypothesized that expression of the mutant human tau transgene in HD mice would worsen the HD phenotype, while knocking out endogenous mouse tau in HD mice would improve some behavioral deficits displayed by HD mice. Our data suggest that neither the expression of a tau transgene nor the ablation of tau expression impacted the progression of the HD motor, cognitive and affective phenotypes. Supporting these behavioral findings, we also found that modulating tau expression had no effect on brain weights in HD mice. We also report that expression of the tau transgene increased the weight of WT and HD male mice, whereas tau ablation increased the weight of HD females only. Together, our results indicate that tau might not be as important in regulating the onset and progression of HD symptomatology as previously proposed.

Identifying causative genes via genetic testing is useful for screening, preventing and treating cancer. Several hereditary syndromes occur in patients with renal cell carcinoma (RCC). However, the evidence is from the European population; it remains unclear how the RCC-related genes and other cancer-predisposing genes contribute to RCC development in the Japanese population. A case-control study of 14 RCC-related genes and 26 cancer-predisposing genes was performed in 1563 Japanese patients with RCC and 6016 controls. The patients were stratified into clear cell RCC (ccRCC) or non-ccRCC (nccRCC). Gene-based analysis of germline pathogenic variants in patients with each subtype and cancer-free subjects was performed. Following quality control, 1532 patients with RCC and 5996 controls were analyzed. For ccRCC, 52 of 1283 (4.05%) patients carried pathogenic variants mainly in the cancer-predisposing genes such as TP53 (P = 1.73 × 10-4; OR, 5.8; 95% CI, 2.2-15.7). Approximately 80% of patients with pathogenic variants in TP53 had p.Ala189Val that was specific in East Asian population. For nccRCC, 14 of 249 (5.62%) patients carried pathogenic variants mainly in the RCC-related genes such as BAP1 and FH (P = 6.27 × 10-5; OR, Inf; 95% CI, 10.0-Inf). The patients with the pathogenic variants in the associated genes were diagnosed 15.8 years earlier and had a higher proportion of patients with a family history of RCC (OR, 20.0; 95% CI, 1.3-237.4) than the non-carriers. We showed different and population-specific contributions of risk genes between ccRCC and nccRCC in Japanese for improved personalized medicine.