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Nuclear clearance and cytoplasmic accumulations of the RNA-binding protein TDP-43 are pathological hallmarks in almost all patients with amyotrophic lateral sclerosis (ALS) and up to 50% of patients with frontotemporal dementia (FTD) and Alzheimers disease. In Alzheimers disease, TDP-43 pathology is predominantly observed in the limbic system and correlates with cognitive decline and reduced hippocampal volume. Disruption of nuclear TDP-43 function leads to abnormal RNA splicing and incorporation of erroneous cryptic exons in numerous transcripts including Stathmin-2 (STMN2, also known as SCG10) and UNC13A, recently reported in tissues from patients with ALS and FTD. Here, we identify both STMN2 and UNC13A cryptic exons in Alzheimers disease patients, that correlate with TDP-43 pathology burden, but not with amyloid-β or tau deposits. We also demonstrate that processing of the STMN2 pre-mRNA is more sensitive to TDP-43 loss of function than UNC13A. In addition, full-length RNAs encoding STMN2 and UNC13A are suppressed in large RNA-seq datasets generated from Alzheimers disease post-mortem brain tissue. Collectively, these results open exciting new avenues to use STMN2 and UNC13A as potential therapeutic targets in a broad range of neurodegenerative conditions with TDP-43 proteinopathy including Alzheimers disease.

Biallelic loss‐of‐function MEGF10 mutations lead to MEGF10 myopathy, also known as early onset myopathy with areflexia, respiratory distress, and dysphagia (EMARDD). MEGF10 is expressed in muscle satellite cells, but the contribution of satellite cell dysfunction to MEGF10 myopathy is unclear. Myofibers and satellite cells were isolated and examined from Megf10−/− and wild‐type mice. A separate set of mice underwent repeated intramuscular barium chloride injections. Megf10−/− muscle satellite cells showed reduced proliferation and migration, while Megf10−/− mouse skeletal muscles showed impaired regeneration. Megf10 deficiency is associated with impaired muscle regeneration, due in part to defects in satellite cell function. Efforts to rescue Megf10 deficiency will have therapeutic implications for MEGF10 myopathy and other inherited muscle diseases involving impaired muscle regeneration. [ABSTRACT FROM AUTHOR]

Gene expression in a tissue-specific context depends on the combined efforts of epigenetic, transcriptional and post-transcriptional processes that lead to the production of specific proteins that are important determinants of cellular identity. Ribosomes are a central component of the protein biosynthesis machinery in cells; however, their regulatory roles in the translational control of gene expression in skeletal muscle remain to be defined. In a genetic screen to identify critical regulators of myogenesis, we identified a DEAD-Box RNA helicase, DDX27, that is required for skeletal muscle growth and regeneration. We demonstrate that DDX27 regulates ribosomal RNA (rRNA) maturation, and thereby the ribosome biogenesis and the translation of specific transcripts during myogenesis. These findings provide insight into the translational regulation of gene expression in myogenesis and suggest novel functions for ribosomes in regulating gene expression in skeletal muscles.

Background: Insect metamorphosis relies on temporal and spatial cues that are precisely controlled. Previous studies in Drosophila have shown that untimely activation of genes that are essential to metamorphosis results in growth defects, developmental delay and death. Multiple factors exist that safeguard these genes against dysregulated expression. The list of identified negative regulators that play such a role in Drosophila development continues to expand. Results: By using RNAi transgene-induced gene silencing coupled to spatio/temporal assessment, we have unraveled an important role for the Drosophila dopamine 1-like receptor, Dop1R2, in development. We show that Dop1R2 knockdown leads to pre-adult lethality. In adults that escape death, abnormal wing expansion and/or melanization defects occur. Furthermore we show that salivary gland expression of this GPCR during the late larval/prepupal stage is essential for the flies to survive through adulthood. In addition to RNAi-induced effects, treatment of larvae with the high affinity D1-like receptor antagonist flupenthixol, also results in developmental arrest, and in morphological defects comparable to those seen in Dop1R2 RNAi flies. To examine the basis for pupal lethality in Dop1R2 RNAi flies, we carried out transcriptome analysis. These studies revealed up-regulation of genes that respond to ecdysone, regulate morphogenesis and/or modulate defense/immunity. Conclusion: Taken together our findings suggest a role for Dop1R2 in the repression of genes that coordinate metamorphosis. Premature release of this inhibition is not tolerated by the developing fly. [ABSTRACT FROM AUTHOR]

Introduction/Aims Methods Results Discussion Heterogeneous nuclear ribonucleoprotein A1 is involved in nucleic acid homeostatic functions. The encoding gene HNRNPA1 has been associated with several neuromuscular disorders including an amyotrophic lateral sclerosis‐like phenotype, distal hereditary motor neuropathy, multisystem proteinopathy, and various myopathies. We report two unrelated individuals with monoallelic stop loss variants affecting the same codon of HNRNPA1.Two individuals with unsolved juvenile‐onset myopathy were enrolled under approved institutional protocols. Phenotype data were collected and genetic analyses were performed, including whole‐exome sequencing (WES).The two probands (MNOT002‐01 and K1440‐01) showed a similar onset of slowly progressive extremity and facial weakness in early adolescence. K1440‐01 presented with facial weakness, winged scapula, elevated serum creatine kinase (CK) levels, and mild neck weakness. MNOT002‐01 also exhibited elevated CK levels along with facial weakness, cardiomyopathy, respiratory dysfunction, pectus excavatum, a mildly rigid spine, and loss of ambulation. On quadriceps muscle biopsy, K1440‐01 displayed rounded myofibers, mild variation in fiber diameter, and type 2 fiber hypertrophy, while MNOT002‐01 displayed rimmed vacuoles. Monoallelic stop‐loss variants in HNRNPA1 were identified for both probands: c.1119A>C p.*373Tyrext*6 (K1440‐01) and c.1118A>C p.*373Serext*6 (MNOT002‐01) affect the same codon and are both predicted to lead to the addition of six amino acids before termination at an alternative stop codon.Both stop‐loss variants in our probands are likely pathogenic. Our findings contribute to the disease characterization of pathogenic variants in HNRNPA1. This gene should be screened in clinical diagnostic testing of unsolved cases of sporadic or dominant juvenile‐onset myopathy. [ABSTRACT FROM AUTHOR]

Background: Pathogenic mutations causing aberrant splicing are often difficult to detect. Standard variant analysis of next‐generation sequence (NGS) data focuses on canonical splice sites. Noncanonical splice sites are more difficult to ascertain. Methods: We developed a bioinformatics pipeline that screens existing NGS data for potentially aberrant novel essential splice sites (PANESS) and performed a pilot study on a family with a myotonic disorder. Further analyses were performed via qRT‐PCR, immunoblotting, and immunohistochemistry. RNAi knockdown studies were performed in Drosophila to model the gene deficiency. Results: The PANESS pipeline identified a homozygous ATP2A1 variant (NC_000016.9:g.28905928G>A; NM_004320.4:c.1287G>A:p.(Glu429=)) that was predicted to cause the omission of exon 11. Aberrant splicing of ATP2A1 was confirmed via qRT‐PCR, and abnormal expression of the protein product sarcoplasmic/endoplasmic reticulum Ca++ ATPase 1 (SERCA1) was demonstrated in quadriceps femoris tissue from the proband. Ubiquitous knockdown of SERCA led to lethality in Drosophila, as did knockdown targeting differentiating or fusing myoblasts. Conclusions: This study confirms the potential of novel in silico algorithms to detect cryptic mutations in existing NGS data; expands the phenotypic spectrum of ATP2A1 mutations beyond classic Brody myopathy; and suggests that genetic testing of ATP2A1 should be considered in patients with clinical myotonia. We developed a novel bioinformatics pipeline to identify potentially aberrant novel essential splice site (PANESS) mutations in next‐generation sequencing data, and used it successfully to identify a pathogenic splice site mutation in ATP2A1 for a family with a myotonic phenotype. The pathogenic nature of the mutation was confirmed via RT‐PCR, western blot, and immunohistochemistry, along with functional studies in Drosophila. [ABSTRACT FROM AUTHOR]

Pathogenic variants in HMGCR were recently linked to a limb-girdle muscular dystrophy (LGMD) phenotype. The protein product HMG CoA reductase (HMGCR) catalyzes a key component of the cholesterol synthesis pathway. The two other muscle diseases associated with HMGCR, statin-associated myopathy (SAM) and autoimmune anti-HMGCR myopathy, are not inherited in a Mendelian pattern. The mechanism linking pathogenic variants in HMGCR with skeletal muscle dysfunction is unclear. We knocked down Hmgcr in mouse skeletal myoblasts, knocked down hmgcr in Drosophila, and expressed three pathogenic HMGCR variants (c.1327C>T, p.Arg443Trp; c.1522_1524delTCT, p.Ser508del; and c.1621G>A, p.Ala541Thr) in Hmgcr knockdown mouse myoblasts. Hmgcr deficiency was associated with decreased proliferation, increased apoptosis, and impaired myotube fusion. Transcriptome sequencing of Hmgcr knockdown versus control myoblasts revealed differential expression involving mitochondrial function, with corresponding differences in cellular oxygen consumption rates. Both ubiquitous and muscle-specific knockdown of hmgcr in Drosophila led to lethality. Overexpression of reference HMGCR cDNA rescued myotube fusion in knockdown cells, whereas overexpression of the pathogenic variants of HMGCR cDNA did not. These results suggest that the three HMGCR-related muscle diseases share disease mechanisms related to skeletal muscle development.