Meta-Analysis of Cell Therapy Studies in Heart Failure and Acute Myocardial Infarction.

Heart failure (HF) is one of the leading causes of death worldwide and has reached epidemic proportions in most industrialized nations. Despite major improvements in the treatment and management of the disease, the prognosis for patients with HF remains poor with approximately only half of patients surviving for 5 years or longer after diagnosis. The poor prognosis of HF patients is in part because of irreparable damage to cardiac tissue and concomitant maladaptive changes associated with the disease. Cell-based therapies may have the potential to transform the treatment and prognosis of HF through regeneration or repair of damaged cardiac tissue. Accordingly, numerous phase I and II randomized clinical trials have tested the clinical benefits of cell transplant, mostly autologous bone marrow-derived mononuclear cells, in patients with HF, ischemic heart disease, and acute myocardial infarction. Although many of these trials were relatively small, meta-analyses of cell-based therapies have attempted to apply rigorous statistical methodology to assess the potential clinical benefits of the intervention. As a prelude to larger phase III trials, meta-analyses, therefore, remain the obvious means of evaluating the available clinical evidence. Here, we review the different meta-analyses of randomized clinical trials that evaluate the safety and potential beneficial effect of cell therapies in HF and acute myocardial infarction spanning nearly 2 decades since the first pioneering trials were conducted.

Role of major and brain-specific Sgce isoforms in the pathogenesis of myoclonus-dystonia syndrome.

Loss-of-function mutations in SGCE, which encodes ε-sarcoglycan (ε-SG), cause myoclonus-dystonia syndrome (OMIM159900, DYT11). A "major" ε-SG protein derived from CCDS5637.1 (NM_003919.2) and a "brain-specific" protein, that includes sequence derived from alternative exon 11b (CCDS47642.1, NM_001099400.1), are reportedly localized in post- and pre-synaptic membrane fractions, respectively. Moreover, deficiency of the "brain-specific" isoform and other isoforms derived from exon 11b may be central to the pathogenesis of DYT11. However, no animal model supports this hypothesis. Gene-trapped ES cells (CMHD-GT_148G1-3, intron 9 of NM_011360) were used to generate a novel Sgce mouse model (C57BL/6J background) with markedly reduced expression of isoforms derived from exons 3' to exon 9 of NM_011360. Among those brain regions analyzed in adult (2month-old) wild-type (WT) mice, cerebellum showed the highest relative expression of isoforms incorporating exon 11b. Homozygotes (SgceGt(148G1)Cmhd/Gt(148G1)Cmhd or SgceGt/Gt) and paternal heterozygotes (Sgcem+/pGt, m-maternal, p-paternal) showed 60 to 70% reductions in expression of total Sgce. Although expression of the major (NM_011360) and brain-specific (NM_001130189) isoforms was markedly reduced, expression of short isoforms was preserved and relatively small amounts of chimeric ε-SG/ß-galactosidase fusion protein was produced by the Sgce gene-trap locus. Immunoaffinity purification followed by mass spectrometry assessments of Sgcem+/pGt mouse brain using pan- or brain-specific ε-SG antibodies revealed significant reductions of ε-SG and other interacting sarcoglycans. Genome-wide gene-expression data using RNA derived from adult Sgcem+/pGt mouse cerebellum showed that the top up-regulated genes were involved in cell cycle, cellular development, cell death and survival, while the top down-regulated genes were associated with protein synthesis, cellular development, and cell death and survival. In comparison to WT littermates, Sgcem+/pGt mice exhibited "tiptoe" gait and stimulus-induced appendicular posturing between Postnatal Days 14 to 16. Abnormalities noted in older Sgcem+/pGt mice included reduced body weight, altered gait dynamics, and reduced open-field activity. Overt spontaneous or stimulus-sensitive myoclonus was not apparent on the C57BL/6J background or mixed C57BL/6J-BALB/c and C57BL/6J-129S2 backgrounds. Our data confirm that mouse Sgce is a maternally imprinted gene and suggests that short Sgce isoforms may compensate, in part, for deficiency of major and brain-specific Sgce isoforms.

C9orf72 ablation in mice does not cause motor neuron degeneration or motor deficits.

OBJECTIVE: How hexanucleotide (GGGGCC) repeat expansions in C9ORF72 cause amyotrophic lateral sclerosis (ALS) remains poorly understood. Both gain- and loss-of-function mechanisms have been proposed. Evidence supporting these mechanisms in vivo is, however, incomplete. Here we determined the effect of C9orf72 loss-of-function in mice. METHODS: We generated and analyzed a conditional C9orf72 knockout mouse model. C9orf72(fl/fl) mice were crossed with Nestin-Cre mice to selectively remove C9orf72 from neurons and glial cells. Immunohistochemistry was performed to study motor neurons and neuromuscular integrity, as well as several pathological hallmarks of ALS, such as gliosis and TDP-43 mislocalization. In addition, motor function and survival were assessed. RESULTS: Neural-specific ablation of C9orf72 in conditional C9orf72 knockout mice resulted in significantly reduced body weight but did not induce motor neuron degeneration, defects in motor function, or altered survival. INTERPRETATION: Our data suggest that C9orf72 loss-of-function, by itself, is insufficient to cause motor neuron disease. These results may have important implications for the development of therapeutic strategies for C9orf72-associated ALS.

Myoclonus dystonia and muscular dystrophy: ɛ-sarcoglycan is part of the dystrophin-associated protein complex in brain.

BACKGROUND: Myoclonus-dystonia is a neurogenic movement disorder caused by mutations in the gene encoding ɛ-sarcoglycan. By contrast, mutations in the α-, ß-, γ-, and δ-sarcoglycan genes cause limb girdle muscular dystrophies. The sarcoglycans are part of the dystrophin-associated protein complex in muscle that is disrupted in several types of muscular dystrophy. Intriguingly, patients with myoclonus-dystonia have no muscle pathology; conversely, limb-girdle muscular dystrophy patients have not been reported to have dystonia-associated features. To gain further insight into the molecular mechanisms underlying these differences, we searched for evidence of a sarcoglycan complex in the brain. METHODS: Immunoaffinity chromatography and mass spectrometry were used to purify ubiquitous and brain-specific ɛ-sarcoglycan directly from tissue. Cell models were used to determine the effect of mutations on the trafficking and assembly of the brain sarcoglycan complex. RESULTS: Ubiquitous and brain-specific ɛ-sarcoglycan isoforms copurify with ß-, δ-, and ζ-sarcoglycan, ß-dystroglycan, and dystrophin Dp71 from brain. Incorporation of a muscular dystrophy-associated ß-sarcoglycan mutant into the brain sarcoglycan complex impairs the formation of the ßδ-sarcoglycan core but fails to abrogate the association and membrane trafficking of ɛ- and ζ-sarcoglycan. CONCLUSIONS: ɛ-Sarcoglycan is part of the dystrophin-associated protein complex in brain. Partial preservation of ɛ- and ζ-sarcoglycan in brain may explain the absence of myoclonus dystonia-like features in muscular dystrophy patients. © 2016 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.

SGCE mutations cause psychiatric disorders: clinical and genetic characterization.

Myoclonus dystonia syndrome is a childhood onset hyperkinetic movement disorder characterized by predominant alcohol responsive upper body myoclonus and dystonia. A proportion of cases are due to mutations in the maternally imprinted SGCE gene. Previous studies have suggested that patients with SGCE mutations may have an increased rate of psychiatric disorders. We established a cohort of patients with myoclonus dystonia syndrome and SGCE mutations to determine the extent to which psychiatric disorders form part of the disease phenotype. In all, 89 patients with clinically suspected myoclonus dystonia syndrome were recruited from the UK and Ireland. SGCE was analysed using direct sequencing and for copy number variants. In those patients where no mutation was found TOR1A (GAG deletion), GCH1, THAP1 and NKX2-1 were also sequenced. SGCE mutation positive cases were systematically assessed using standardized psychiatric interviews and questionnaires and compared with a disability-matched control group of patients with alcohol responsive tremor. Nineteen (21%) probands had a SGCE mutation, five of which were novel. Recruitment of family members increased the affected SGCE mutation positive group to 27 of whom 21 (77%) had psychiatric symptoms. Obsessive-compulsive disorder was eight times more likely (P < 0.001) in mutation positive cases, compulsivity being the predominant feature (P < 0.001). Generalized anxiety disorder (P = 0.003) and alcohol dependence (P = 0.02) were five times more likely in mutation positive cases than tremor controls. SGCE mutations are associated with a specific psychiatric phenotype consisting of compulsivity, anxiety and alcoholism in addition to the characteristic motor phenotype. SGCE mutations are likely to have a pleiotropic effect in causing both motor and specific psychiatric symptoms.

Adeno-associated virus 9 mediated FKRP gene therapy restores functional glycosylation of α-dystroglycan and improves muscle functions.

Mutations in the FKRP gene are associated with a wide range of muscular dystrophies from mild limb-girdle muscular dystrophy (LGMD) 2I to severe Walker-Warburg syndrome and muscle-eye-brain disease. The characteristic biochemical feature of these diseases is the hypoglycosylation of α-dystroglycan (α-DG). Currently there is no effective treatment available. In this study, we examined the adeno-associated virus serotype 9 vector (AAV9)-mediated gene therapy in the FKRP mutant mouse model with a proline to leucine missense mutation (P448L). Our results showed that intraperitoneal administration of AAV9-FKRP resulted in systemic FKRP expression in all striated muscles examined with the highest levels in cardiac muscle. Consistent with our previous observations, FKRP protein is localized in the Golgi apparatus in myofibers. Expression of FKRP consequently restored functional glycosylation of α-DG in the skeletal and cardiac muscles. Significant improvement in dystrophic pathology, serum creatine kinase levels and muscle function was observed. Only limited FKRP transgene expression was detected in kidney and liver with no detectable toxicity. Our results provided evidence for the utility of AAV-mediated gene replacement therapy for FKRP-related muscular dystrophies.

Hermansky-Pudlak syndrome type 7 (HPS-7) results from mutant dysbindin, a member of the biogenesis of lysosome-related organelles complex 1 (BLOC-1).

Hermansky-Pudlak syndrome (HPS; MIM 203300) is a genetically heterogeneous disorder characterized by oculocutaneous albinism, prolonged bleeding and pulmonary fibrosis due to abnormal vesicle trafficking to lysosomes and related organelles, such as melanosomes and platelet dense granules. In mice, at least 16 loci are associated with HPS, including sandy (sdy; ref. 7). Here we show that the sdy mutant mouse expresses no dysbindin protein owing to a deletion in the gene Dtnbp1 (encoding dysbindin) and that mutation of the human ortholog DTNBP1 causes a novel form of HPS called HPS-7. Dysbindin is a ubiquitously expressed protein that binds to alpha- and beta-dystrobrevins, components of the dystrophin-associated protein complex (DPC) in both muscle and nonmuscle cells. We also show that dysbindin is a component of the biogenesis of lysosome-related organelles complex 1 (BLOC-1; refs. 9-11), which regulates trafficking to lysosome-related organelles and includes the proteins pallidin, muted and cappuccino, which are associated with HPS in mice. These findings show that BLOC-1 is important in producing the HPS phenotype in humans, indicate that dysbindin has a role in the biogenesis of lysosome-related organelles and identify unexpected interactions between components of DPC and BLOC-1.

Misfolding of fukutin-related protein (FKRP) variants in congenital and limb girdle muscular dystrophies.

Fukutin-related protein (FKRP, MIM ID 606596) variants cause a range of muscular dystrophies associated with hypo-glycosylation of the matrix receptor, α-dystroglycan. These disorders are almost exclusively caused by homozygous or compound heterozygous missense variants in the FKRP gene that encodes a ribitol phosphotransferase. To understand how seemingly diverse FKRP missense mutations may contribute to disease, we examined the synthesis, intracellular dynamics, and structural consequences of a panel of missense mutations that encompass the disease spectrum. Under non-reducing electrophoresis conditions, wild type FKRP appears to be monomeric whereas disease-causing FKRP mutants migrate as high molecular weight, disulfide-bonded aggregates. These results were recapitulated using cysteine-scanning mutagenesis suggesting that abnormal disulfide bonding may perturb FKRP folding. Using fluorescence recovery after photobleaching, we found that the intracellular mobility of most FKRP mutants in ATP-depleted cells is dramatically reduced but can, in most cases, be rescued with reducing agents. Mass spectrometry showed that wild type and mutant FKRP differentially associate with several endoplasmic reticulum (ER)-resident chaperones. Finally, structural modelling revealed that disease-associated FKRP missense variants affected the local environment of the protein in small but significant ways. These data demonstrate that protein misfolding contributes to the molecular pathophysiology of FKRP-deficient muscular dystrophies and suggest that molecules that rescue this folding defect could be used to treat these disorders.

Functional analysis of TCF4 missense mutations that cause Pitt-Hopkins syndrome.

Pitt-Hopkins syndrome (PTHS) is a rare developmental disorder associated with severe mental retardation, facial abnormalities, and intermittent hyperventilation. Autosomal dominant PTHS is caused by mutations in the transcription factor 4 (TCF4) gene, whereas NRXN1 and CNTNAP2 mutations are associated with autosomal recessive PTHS. To determine the impact of missense mutations on TCF4 function, we tested a panel of PTHS-associated mutations using a range of quantitative techniques. Mutations in the basic helix-loop-helix (bHLH) domain of TCF4 alter the subnuclear localization of the mutant protein and can attenuate homo- and heterodimer formation in homogenous time-resolved fluorescence (HTRF) assays. By contrast, mutations proximal to the bHLH domain do not alter the location of TCF4 or impair heterodimer formation. In addition, we show that TCF4 can transactivate the NRXN1ß and CNTNAP2 promoters in luciferase assays. Here we find variable, context-specific deficits in the ability of the different PTHS-associated TCF4 mutants to transactivate these promoters when coexpressed with different bHLH transcription factors. These data demonstrate that PTHS-associated missense mutations can have multiple effects on the function of the protein, and suggest that TCF4 may modulate the expression of NRXN1 and CNTNAP2 thereby defining a regulatory network in PTHS.

Fukutin-related protein is essential for mouse muscle, brain and eye development and mutation recapitulates the wide clinical spectrums of dystroglycanopathies.

Mutations in fukutin-related protein (FKRP) cause a common subset of muscular dystrophies characterized by aberrant glycosylation of alpha-dystroglycan (α-DG), collectively known as dystroglycanopathies. The clinical variations associated with FKRP mutations range from mild limb-girdle muscular dystrophy type 2I with predominantly muscle phenotypes to severe Walker-Warburg syndrome and muscle-eye-brain disease with striking structural brain and eye defects. In the present study, we have generated animal models and demonstrated that ablation of FKRP functions is embryonic lethal and that the homozygous-null embryos die before reaching E12.5. The homozygous knock-in mouse carrying the missense P448L mutation almost completely lacks functional glycosylation of α-DG in muscles and brain, validating the essential role of FKRP in the functional glycosylation of α-DG. However, the knock-in mouse survives and develops a wide range of structural abnormalities in the central nervous system, characteristics of neuronal migration defects. The brain and eye defects are highly reminiscent of the phenotypes seen in severe dystroglycanopathy patients. In addition, skeletal muscles develop progressive muscular dystrophy. Our results confirm that post-translational modifications of α-DG are essential for normal development of the brain and eyes. In addition, both the mutation itself and the levels of FKRP expression are equally critical for the survival of the animals. The exceptionally wide clinical spectrums recapitulated in the P448L mice also suggest the involvement of other factors in the disease progression. The mutant mouse represents a valuable model to further elucidate the functions of FKRP and develop therapies for FKRP-related muscular dystrophies.

Recent advances in the genetics of the ALS-FTLD complex.

There is a clinical and pathological overlap between amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). A number of autosomal-dominant genes have been described that primarily cause ALS or FTLD such as progranulin (GRN), valosin-containing protein (VCP), and TAR DNA-Binding Protein (TARDBP), and for each of these conditions there are a small number of cases with both ALS and FTLD. Two major genes were described in 2011, which cause FTLD and/or ALS within extended kindreds. Ubiquilin2 (UBQLN2) is responsible for X-linked FTLD/ALS. A hexanucleotide repeat expansion in C9ORF72 causes chromosome 9p linked FTLD/ALS and is the most common cause of familial ALS accounting for about 40 % of familial cases. Both UBQLN2 and C9ORF72 mutations lead to TDP-43 positive neuropathology, and C9ORF72-positive cases have p62/ubiquitin-positive pathology, which is not stained by TDP-43 antibodies. Ubiquilin2 is one of a family of proteins thought to be important in targeting abnormal proteins for degradation via lysosomal and proteasomal routes. The pathogenic mechanism of the C9ORF72 expansion is unknown but may involve partial haploinsufficiency of C9ORF72 and/or the formations of toxic RNA inclusions. The identification of mutations in these genes represents an important step forward in our understanding of the clinical, pathological, and genetic spectrum of ALS/FTLD diseases.

CMYA5 is a novel interaction partner of FHL2 in cardiac myocytes.

Four-and-a-half LIM domains protein 2 (FHL2) is an anti-hypertrophic adaptor protein that regulates cardiac myocyte signalling and function. Herein, we identified cardiomyopathy-associated 5 (CMYA5) as a novel FHL2 interaction partner in cardiac myocytes. In vitro pull-down assays demonstrated interaction between FHL2 and the N- and C-terminal regions of CMYA5. The interaction was verified in adult cardiac myocytes by proximity ligation assays. Immunofluorescence and confocal microscopy demonstrated co-localisation in the same subcellular compartment. The binding interface between FHL2 and CMYA5 was mapped by peptide arrays. Exposure of neonatal rat ventricular myocytes to a CMYA5 peptide covering one of the FHL2 interaction sites led to an increase in cell area at baseline, but a blunted response to chronic phenylephrine treatment. In contrast to wild-type hearts, loss or reduced FHL2 expression in Fhl2-targeted knockout mouse hearts or in a humanised mouse model of hypertrophic cardiomyopathy led to redistribution of CMYA5 into the perinuclear and intercalated disc region. Taken together, our results indicate a direct interaction of the two adaptor proteins FHL2 and CMYA5 in cardiac myocytes, which might impact subcellular compartmentation of CMYA5.

Transcriptional programs regulating neuronal differentiation are disrupted in DLG2 knockout human embryonic stem cells and enriched for schizophrenia and related disorders risk variants.

Coordinated programs of gene expression drive brain development. It is unclear which transcriptional programs, in which cell-types, are affected in neuropsychiatric disorders such as schizophrenia. Here we integrate human genetics with transcriptomic data from differentiation of human embryonic stem cells into cortical excitatory neurons. We identify transcriptional programs expressed during early neurogenesis in vitro and in human foetal cortex that are down-regulated in DLG2-/- lines. Down-regulation impacted neuronal differentiation and maturation, impairing migration, morphology and action potential generation. Genetic variation in these programs is associated with neuropsychiatric disorders and cognitive function, with associated variants predominantly concentrated in loss-of-function intolerant genes. Neurogenic programs also overlap schizophrenia GWAS enrichment previously identified in mature excitatory neurons, suggesting that pathways active during prenatal cortical development may also be associated with mature neuronal dysfunction. Our data from human embryonic stem cells, when combined with analysis of available foetal cortical gene expression data, de novo rare variants and GWAS statistics for neuropsychiatric disorders and cognition, reveal a convergence on transcriptional programs regulating excitatory cortical neurogenesis.

A gain-of-glycosylation mutation associated with myoclonus-dystonia syndrome affects trafficking and processing of mouse ε-sarcoglycan in the late secretory pathway.

Missense mutations in the SGCE gene encoding ε-sarcoglycan account for approximately 15% of SGCE-positive cases of myoclonus-dystonia syndrome (MDS) in humans. In this study, we show that while the majority of MDS-associated missense mutants modeled with a murine ε-sarcoglycan cDNA are substrates for endoplasmic reticulum-associated degradation, one mutant, M68T (analogous to human c.275T>C, p.M92T), located in the Ig-like domain of ε-sarcoglycan, results in a gain-of-glycosylation mutation producing a protein that is targeted to the plasma membrane, albeit at reduced levels compared to wild-type ε-sarcoglycan. Removal of the ectopic N-linked glycan failed to restore efficient plasma membrane targeting of M68T demonstrating that the substitution rather than the glycan was responsible for the trafficking defect of this mutant. M68T also colocalized with CD63-positive vesicles in the endosomal-lysosomal system and was found to be more susceptible to lysosomal proteolysis than wild-type ε-sarcoglycan. Finally, we demonstrate impaired ectodomain shedding of M68T, a process that occurs physiologically for ε-sarcoglycan resulting in the lysosomal trafficking of the intracellular C-terminal domain of the protein. Our findings show that functional analysis of rare missense mutations can provide a mechanistic insight into the pathogenesis of MDS and the physiological role of ε-sarcoglycan.

TRIM32 is an E3 ubiquitin ligase for dysbindin.

Mutations in the gene encoding tripartite motif protein 32 (TRIM32) cause two seemingly diverse diseases: limb-girdle muscular dystrophy type 2H (LGMD2H) or sarcotubular myopathy (STM) and Bardet-Biedl syndrome type 11(BBS11). Although TRIM32 is involved in protein ubiquitination, its substrates and the molecular consequences of disease-causing mutations are poorly understood. In this paper, we show that TRIM32 is a widely expressed ubiquitin ligase that is localized to the Z-line in skeletal muscle. Using the yeast two-hybrid system, we found that TRIM32 binds and ubiquitinates dysbindin, a protein implicated in the genetic aetiology of schizophrenia, augmenting its degradation. Small-interfering RNA-mediated knock-down of TRIM32 in myoblasts resulted in elevated levels of dysbindin. Importantly, the LGMD2H/STM-associated TRIM32 mutations, D487N and R394H impair ubiquitin ligase activity towards dysbindin and were mislocalized in heterologous cells. These mutants were able to self-associate and also co-immunoprecipitated with wild-type TRIM32 in transfected cells. Furthermore, the D487N mutant could bind to both dysbindin and its E2 enzyme but was defective in monoubiquitination. In contrast, the BBS11 mutant P130S did not show any biochemical differences compared with the wild-type protein. Our data identify TRIM32 as a regulator of dysbindin and demonstrate that the LGMD2H/STM mutations may impair substrate ubiquitination.

Dysbindin-1 in dorsolateral prefrontal cortex of schizophrenia cases is reduced in an isoform-specific manner unrelated to dysbindin-1 mRNA expression.

DTNBP1 (dystrobrevin binding protein 1) remains a top candidate gene in schizophrenia. Reduced expression of this gene and of its encoded protein, dysbindin-1, have been reported in the brains of schizophrenia cases. It has not been established, however, if the protein reductions encompass all dysbindin-1 isoforms or if they are associated with decreased DTNBP1 gene expression. Using a matched pairs design in which each of 28 Caucasian schizophrenia cases was matched in age and sex to a normal Caucasian control, Western blotting of whole-tissue lysates of dorsolateral prefrontal cortex (DLPFC) revealed significant reductions in dysbindin-1C (but not in dysbindin-1A or -1B) in schizophrenia (P = 0.022). These reductions occurred without any significant change in levels of the encoding transcript in the same tissue samples and in the absence of the only DTNBP1 risk haplotype for schizophrenia reported in the USA. Indeed, no significant correlations were found between case-control differences in any dysbindin-1 isoform and the case-control differences in its encoding mRNA. Consequently, the mean 60% decrease in dysbindin-1C observed in 71% of our case-control pairs appears to reflect abnormalities in mRNA translation and/or processes promoting dysbindin-1C degradation (e.g. oxidative stress, phosphorylation and/or ubiquitination). Given the predominantly post-synaptic localization of dysbindin-1C and known post-synaptic effects of dysbindin-1 reductions in the rodent equivalent of the DLPFC, the present findings suggest that decreased dysbindin-1C in the DLPFC may contribute to the cognitive deficits of schizophrenia by promoting NMDA receptor hypofunction in fast-spiking interneurons.

Psychiatric disorders, myoclonus dystonia, and the epsilon-sarcoglycan gene: a systematic review.

BACKGROUND: Mutations in the maternally imprinted epsilon-sarcoglycan gene occur in 30%-50% of myoclonus-dystonia cases. Psychiatric symptoms, particularly obsessive-compulsive disorder, have been described in some patients. METHODS: We systematically reviewed 22 reports of psychiatric symptoms in myoclonus-dystonia, dividing individuals according to clinical and mutation status. RESULTS: Clinically manifesting mutation carriers demonstrated an excess of psychiatric disorders compared with nonmutation carriers (P < .001). No differences were seen between non-motor-manifesting carriers and nonmutation carriers with the exception of alcohol excess/dependence, higher in non-motor-manifesting carriers. CONCLUSIONS: The results confirm the association of epsilon-sarcoglycan gene mutations with psychiatric disease and suggest a possible separation of the motor and psychiatric effects.

Isoform-Specific Reduction of the Basic Helix-Loop-Helix Transcription Factor TCF4 Levels in Huntington's Disease.

Huntington's disease (HD) is an inherited neurodegenerative disorder with onset of characteristic motor symptoms at midlife, preceded by subtle cognitive and behavioral disturbances. Transcriptional dysregulation emerges early in the disease course and is considered central to HD pathogenesis. Using wild-type (wt) and HD knock-in mouse striatal cell lines we observed a HD genotype-dependent reduction in the protein levels of transcription factor 4 (TCF4), a member of the basic helix-loop-helix (bHLH) family with critical roles in brain development and function. We characterized mouse Tcf4 gene structure and expression of alternative mRNAs and protein isoforms in cell-based models of HD, and in four different brain regions of male transgenic HD mice (R6/1) from young to mature adulthood. The largest decrease in the levels of TCF4 at mRNA and specific protein isoforms were detected in the R6/1 mouse hippocampus. Translating this finding to human disease, we found reduced expression of long TCF4 isoforms in the postmortem hippocampal CA1 area and in the cerebral cortex of HD patients. Additionally, TCF4 protein isoforms showed differential synergism with the proneural transcription factor ASCL1 in activating reporter gene transcription in hippocampal and cortical cultured neurons. Induction of neuronal activity increased these synergistic effects in hippocampal but not in cortical neurons, suggesting brain region-dependent differences in TCF4 functions. Collectively, this study demonstrates isoform-specific changes in TCF4 expression in HD that could contribute to the progressive impairment of transcriptional regulation and neuronal function in this disease.

Intragenic deletion of TRIM32 in compound heterozygotes with sarcotubular myopathy/LGMD2H.

In 2005 the commonality of sarcotubular myopathy (STM) and limb girdle muscular dystrophy type 2H (LGMD2H) was demonstrated, as both are caused by the p D487N missense mutation in TRIM32 originally found in the Manitoba Hutterite population. Recently, three novel homozygous TRIM32 mutations have been described in LGMD patients. Here we describe a three generation Swedish family clinically presenting with limb girdle muscular weakness and histological features of a microvacuolar myopathy. The two index patients were compound heterozygotes for a frameshift mutation in TRIM32 (c.1560delC ) and a 30 kb intragenic deletion, encompassing parts of intron 1 and the entire exon 2 of TRIM32. In these patients, no full-length or truncated TRIM32 could be detected. Interestingly, heterozygous family members carrying only one mutation showed mild clinical symptoms and vacuolar changes in muscle. In our family, the phenotype encompasses additionally a mild demyelinating polyneuropathic syndrome. Thus STM and LGMD2H are the result of loss of function mutations that can be either deletions or missense mutations.

Transcriptional Changes following Cellular Knockdown of the Schizophrenia Risk Gene SETD1A Are Enriched for Common Variant Association with the Disorder.

Loss of function mutations in SETD1A are the first experiment-wide significant findings to emerge from exome sequencing studies of schizophrenia. Although SETD1A is known to encode a histone methyltransferase, the consequences of reduced S ETD1A activity on gene expression in neural cells have, to date, been unknown. To explore transcriptional changes through which genetic perturbation of SETD1A could confer risk for schizophrenia, we have performed genome-wide gene expression profiling of a commonly used human neuroblastoma cell line in which SETD1A expression has been experimentally reduced using RNA interference (RNAi). We identified 1,031 gene expression changes that were significant in two separate RNAi conditions compared with control, including effects on genes of known neurodevelopmental importance such as DCX and DLX5. Genes that were differentially expressed following SETD1A knockdown were enriched for annotation to metabolic pathways, peptidase regulator activity and integrin-mediated regulation of cell adhesion. Moreover, differentially expressed genes were enriched for common variant association with schizophrenia, suggesting a degree of molecular convergence between this rare schizophrenia risk factor and susceptibility variants for the disorder operating more generally.