Functional analysis reveals splicing mutations of the CASQ2 gene in patients with CPVT: implication for genetic counselling and clinical management.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a rare and severe arrhythmogenic disorder. Although usually transmitted in a recessive form, few cases of dominant mutations have been reported. Thirteen mutations in the CASQ2 gene have been reported so far in association with CPVT. We performed molecular analysis of the CASQ2 gene in 43 probands with CPVT and identified eight mutations in five patients. Six mutations were novel: one was a single nucleotide deletion, three affected consensus splice sites, and two had unknown consequences: the c.939 + 5G>C and the synonymous c.381C>T variations. We demonstrated that these two variations affected CASQ2 splicing using a splicing minigene assay. These data increased significantly the number of CASQ2 mutations described in association with CPVT, revealed the high prevalence of splicing and truncating mutations in this gene and brought new insight regarding the dominant inheritance of the disease. Moreover, our report of the first splicing abnormalities in CASQ2 caused by intronic mutation or synonymous change underlines the absolute necessity to perform extensive molecular analysis for genetic diagnosis and counseling of CPVT.

TRAM (Transcriptome Mapper): database-driven creation and analysis of transcriptome maps from multiple sources.

BACKGROUND: Several tools have been developed to perform global gene expression profile data analysis, to search for specific chromosomal regions whose features meet defined criteria as well as to study neighbouring gene expression. However, most of these tools are tailored for a specific use in a particular context (e.g. they are species-specific, or limited to a particular data format) and they typically accept only gene lists as input. RESULTS: TRAM (Transcriptome Mapper) is a new general tool that allows the simple generation and analysis of quantitative transcriptome maps, starting from any source listing gene expression values for a given gene set (e.g. expression microarrays), implemented as a relational database. It includes a parser able to assign univocal and updated gene symbols to gene identifiers from different data sources. Moreover, TRAM is able to perform intra-sample and inter-sample data normalization, including an original variant of quantile normalization (scaled quantile), useful to normalize data from platforms with highly different numbers of investigated genes. When in 'Map' mode, the software generates a quantitative representation of the transcriptome of a sample (or of a pool of samples) and identifies if segments of defined lengths are over/under-expressed compared to the desired threshold. When in 'Cluster' mode, the software searches for a set of over/under-expressed consecutive genes. Statistical significance for all results is calculated with respect to genes localized on the same chromosome or to all genome genes. Transcriptome maps, showing differential expression between two sample groups, relative to two different biological conditions, may be easily generated. We present the results of a biological model test, based on a meta-analysis comparison between a sample pool of human CD34+ hematopoietic progenitor cells and a sample pool of megakaryocytic cells. Biologically relevant chromosomal segments and gene clusters with differential expression during the differentiation toward megakaryocyte were identified. CONCLUSIONS: TRAM is designed to create, and statistically analyze, quantitative transcriptome maps, based on gene expression data from multiple sources. The release includes FileMaker Pro database management runtime application and it is freely available at http://apollo11.isto.unibo.it/software/, along with preconfigured implementations for mapping of human, mouse and zebrafish transcriptomes.

Motif discovery in promoters of genes co-localized and co-expressed during myeloid cells differentiation.

Genes co-expressed may be under similar promoter-based and/or position-based regulation. Although data on expression, position and function of human genes are available, their true integration still represents a challenge for computational biology, hampering the identification of regulatory mechanisms. We carried out an integrative analysis of genomic position, functional annotation and promoters of genes expressed in myeloid cells. Promoter analysis was conducted by a novel multi-step method for discovering putative regulatory elements, i.e. over-represented motifs, in a selected set of promoters, as compared with a background model. The combination of transcriptional, structural and functional data allowed the identification of sets of promoters pertaining to groups of genes co-expressed and co-localized in regions of the human genome. The application of motif discovery to 26 groups of genes co-expressed in myeloid cells differentiation and co-localized in the genome showed that there are more over-represented motifs in promoters of co-expressed and co-localized genes than in promoters of simply co-expressed genes (CEG). Motifs, which are similar to the binding sequences of known transcription factors, non-uniformly distributed along promoter sequences and/or occurring in highly co-expressed subset of genes were identified. Co-expressed and co-localized gene sets were grouped in two co-expressed genomic meta-regions, putatively representing functional domains of a high-level expression regulation.

Desmosomal dysfunction due to mutations in desmoplakin causes arrhythmogenic right ventricular dysplasia/cardiomyopathy.

Arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C) is characterized by progressive degeneration of the right ventricular myocardium, ventricular arrhythmias, fibrous-fatty replacement, and increased risk of sudden death. Mutations in 6 genes, including 4 encoding desmosomal proteins (Junctional plakoglobin (JUP), Desmoplakin (DSP), Plakophilin 2, and Desmoglein 2), have been identified in patients with ARVD/C. Mutation analysis of 66 probands identified 4 variants in DSP; V30M, Q90R, W233X, and R2834H. To establish a cause and effect relationship between those DSP missense mutations and ARVD/C, we performed in vitro and in vivo analyses of the mutated proteins. Unlike wild-type (WT) DSP, the N-terminal mutants (V30M and Q90R) failed to localize to the cell membrane in desomosome-forming cell line and failed to bind to and coimmunoprecipitate JUP. Multiple attempts to generate N-terminal DSP (V30M and Q90R) cardiac-specific transgenes have failed: analysis of embryos revealed evidence of profound ventricular dilation, which likely resulted in embryonic lethality. We were able to develop transgenic (Tg) mice with cardiac-restricted overexpression of the C-terminal mutant (R2834H) or WT DSP. Whereas mice overexpressing WT DSP had no detectable histologic, morphological, or functional cardiac changes, the R2834H-Tg mice had increased cardiomyocyte apoptosis, cardiac fibrosis, and lipid accumulation, along with ventricular enlargement and cardiac dysfunction in both ventricles. These mice also displayed interruption of DSP-desmin interaction at intercalated discs (IDs) and marked ultra-structural changes of IDs. These data suggest DSP expression in cardiomyocytes is crucial for maintaining cardiac tissue integrity, and DSP abnormalities result in ARVD/C by cardiomyocyte death, changes in lipid metabolism, and defects in cardiac development.

Novel definition files for human GeneChips based on GeneAnnot.

BACKGROUND: Improvements in genome sequence annotation revealed discrepancies in the original probeset/gene assignment in Affymetrix microarray and the existence of differences between annotations and effective alignments of probes and transcription products. In the current generation of Affymetrix human GeneChips, most probesets include probes matching transcripts from more than one gene and probes which do not match any transcribed sequence. RESULTS: We developed a novel set of custom Chip Definition Files (CDF) and the corresponding Bioconductor libraries for Affymetrix human GeneChips, based on the information contained in the GeneAnnot database. GeneAnnot-based CDFs are composed of unique custom-probesets, including only probes matching a single gene. CONCLUSION: GeneAnnot-based custom CDFs solve the problem of a reliable reconstruction of expression levels and eliminate the existence of more than one probeset per gene, which often leads to discordant expression signals for the same transcript when gene differential expression is the focus of the analysis. GeneAnnot CDFs are freely distributed and fully compliant with Affymetrix standards and all available software for gene expression analysis. The CDF libraries are available from http://www.xlab.unimo.it/GA_CDF, along with supplementary information (CDF libraries, installation guidelines and R code, CDF statistics, and analysis results).

Mutations in desmoglein-2 gene are associated with arrhythmogenic right ventricular cardiomyopathy.

BACKGROUND: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiomyopathy characterized by progressive myocardial atrophy with fibrofatty replacement. The recent identification of causative mutations in plakoglobin, desmoplakin (DSP), and plakophilin-2 (PKP2) genes led to the hypothesis that ARVC is due to desmosomal defects. Therefore, desmoglein-2 (DSG2), the only desmoglein isoform expressed in cardiac myocytes, was screened in subjects with ARVC. METHODS AND RESULTS: In a series of 80 unrelated ARVC probands, 26 carried a mutation in DSP (16%), PKP2 (14%), and transforming growth factor-beta3 (2.5%) genes; the remaining 54 were screened for DSG2 mutations by denaturing high-performance liquid chromatography and direct sequencing. Nine heterozygous DSG2 mutations (5 missense, 2 insertion-deletions, 1 nonsense, and 1 splice site mutation) were detected in 8 probands (10%). All probands fulfilled task force criteria for ARVC. An endomyocardial biopsy was obtained in 5, showing extensive loss of myocytes with fibrofatty tissue replacement. In 3 patients, electron microscopy investigation was performed, showing intercalated disc paleness, decreased desmosome number, and intercellular gap widening. CONCLUSIONS: This is the first investigation demonstrating DSG2 gene mutations in a significant number of ARVC-unrelated probands. Cardiac phenotype is characterized clinically by typical ARVC features with frequent left ventricular involvement and morphologically by fibrofatty myocardial replacement and desmosomal remodeling. The presence of mutations in desmosomal encoding genes in 40% of cases confirms that many forms of ARVC are due to alterations in the desmosome complex.

Genomic expression during human myelopoiesis.

BACKGROUND: Human myelopoiesis is an exciting biological model for cellular differentiation since it represents a plastic process where multipotent stem cells gradually limit their differentiation potential, generating different precursor cells which finally evolve into distinct terminally differentiated cells. This study aimed at investigating the genomic expression during myeloid differentiation through a computational approach that integrates gene expression profiles with functional information and genome organization. RESULTS: Gene expression data from 24 experiments for 8 different cell types of the human myelopoietic lineage were used to generate an integrated myelopoiesis dataset of 9,425 genes, each reliably associated to a unique genomic position and chromosomal coordinate. Lists of genes constitutively expressed or silent during myelopoiesis and of genes differentially expressed in commitment phase of myelopoiesis were first identified using a classical data analysis procedure. Then, the genomic distribution of myelopoiesis genes was investigated integrating transcriptional and functional characteristics of genes. This approach allowed identifying specific chromosomal regions significantly highly or weakly expressed, and clusters of differentially expressed genes and of transcripts related to specific functional modules. CONCLUSION: The analysis of genomic expression during human myelopoiesis using an integrative computational approach allowed discovering important relationships between genomic position, biological function and expression patterns and highlighting chromatin domains, including genes with coordinated expression and lineage-specific functions.

Missense mutations in desmocollin-2 N-terminus, associated with arrhythmogenic right ventricular cardiomyopathy, affect intracellular localization of desmocollin-2 in vitro.

BACKGROUND: Mutations in genes encoding desmosomal proteins have been reported to cause arrhythmogenic right ventricular cardiomyopathy (ARVC), an autosomal dominant disease characterised by progressive myocardial atrophy with fibro-fatty replacement. We screened 54 ARVC probands for mutations in desmocollin-2 (DSC2), the only desmocollin isoform expressed in cardiac tissue. METHODS: Mutation screening was performed by denaturing high-performance liquid chromatography and direct sequencing. To evaluate the pathogenic potentials of the DSC2 mutations detected in patients affected with ARVC, full-length wild-type and mutated cDNAs were cloned in eukaryotic expression vectors to obtain a fusion protein with green fluorescence protein (GFP); constructs were transfected in neonatal rat cardiomyocytes and in HL-1 cells. RESULTS: We identified two heterozygous mutations (c.304G>A (p.E102K) and c.1034T>C (p.I345T)) in two probands and in four family members. The two mutations p.E102K and p.I345T map to the N-terminal region, relevant to adhesive interactions. In vitro functional studies demonstrated that, unlike wild-type DSC2, the two N-terminal mutants are predominantly localised in the cytoplasm. CONCLUSION: The two missense mutations in the N-terminal domain affect the normal localisation of DSC2, thus suggesting the potential pathogenic effect of the reported mutations. Identification of additional DSC2 mutations associated with ARVC may result in increased diagnostic accuracy with implications for genetic counseling.

Detecting seeded motifs in DNA sequences.

The problem of detecting DNA motifs with functional relevance in real biological sequences is difficult due to a number of biological, statistical and computational issues and also because of the lack of knowledge about the structure of searched patterns. Many algorithms are implemented in fully automated processes, which are often based upon a guess of input parameters from the user at the very first step. In this paper, we present a novel method for the detection of seeded DNA motifs, composed by regions with a different extent of variability. The method is based on a multi-step approach, which was implemented in a motif searching web tool (MOST). Overrepresented exact patterns are extracted from input sequences and clustered to produce motifs core regions, which are then extended and scored to generate seeded motifs. The combination of automated pattern discovery algorithms and different display tools for the evaluation and selection of results at several analysis steps can potentially lead to much more meaningful results than complete automation can produce. Experimental results on different yeast and human real datasets proved the methodology to be a promising solution for finding seeded motifs. MOST web tool is freely available at http://telethon.bio.unipd.it/bioinfo/MOST.

REEF: searching REgionally Enriched Features in genomes.

BACKGROUND: In Eukaryotic genomes, different features including genes are not uniformly distributed. The integration of annotation information and genomic position of functional DNA elements in the Eukaryotic genomes opened the way to test novel hypotheses of higher order genome organization and regulation of expression. RESULTS: REEF is a new tool, aimed at identifying genomic regions enriched in specific features, such as a class or group of genes homogeneous for expression and/or functional characteristics. The method for the calculation of local feature enrichment uses test statistic based on the Hypergeometric Distribution applied genome-wide by using a sliding window approach and adopting the False Discovery Rate for controlling multiplicity. REEF software, source code and documentation are freely available at http://telethon.bio.unipd.it/bioinfo/reef/. CONCLUSION: REEF can aid to shed light on the role of organization of specific genomic regions in the determination of their functional role.

Computational reconstruction of the human skeletal muscle secretome.

In multicellular organisms, secreted proteins play pivotal regulatory roles in intercellular communication. Proteins secreted by skeletal muscle can act locally on muscle cells through autocrine/paracrine loops and on surrounding tissues such as muscle blood vessels, or they can be released into the blood stream, thus producing systemic effects. By a computational approach, we have screened 6255 products of genes expressed in normal human skeletal muscle. Putatively secreted proteins were identified by sequential steps of sieving, through prediction of signal peptide, recognition of transmembrane regions, and analysis of protein annotation. The resulting putative skeletal muscle secretome consists of 319 proteins, including 78 still uncharacterized proteins. This is the first human skeletal muscle secretome produced by computational analysis. Knowledge of proteins secreted by skeletal muscle could stimulate development of novel treatments for different diseases, including muscle atrophy and dystrophy. In addition, better knowledge of the secretion process in skeletal muscle can be useful for future gene therapy approaches.

Regulatory mutations in transforming growth factor-beta3 gene cause arrhythmogenic right ventricular cardiomyopathy type 1.

OBJECTIVE: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a genetically heterogeneous disorder characterized by fibro-fatty replacement of the right ventricular myocardium, associated with high risk of sudden death. The objective of this study is to identify the gene involved in ARVD1, which has been elusive ever since its locus was mapped to chromosome 14q24.3. METHODS AND RESULTS: Mutation screening of the promoter and untranslated regions (UTRs) of the transforming growth factor-beta3 (TGFbeta3) gene was performed by direct sequencing of genomic DNA of one index case belonging to an ARVD1 family including 38 members in four generations. We detected a nucleotide substitution (c.-36G>A) in 5' UTR of TGFbeta3 gene, invariably associated with the typical ARVC clinical phenotype in the affected family members, according to the established diagnostic criteria. Investigation extended to 30 unrelated ARVC patients, performed by denaturing high-performance liquid chromatography (DHPLC), led to the identification of an additional mutation (c.1723C>T) in the 3' UTR of one proband. Neither nucleotide change was found in 300 control subjects. In vitro expression assays with constructs containing the mutations showed that mutated UTRs were twofold more active than wild-types. CONCLUSION: We identified TGFbeta3 as the disease gene involved in ARVD1. The identification of a novel ARVC gene will increase the power of the genetic screening for early diagnosis of asymptomatic carriers among relatives of ARVC patients.

A multistep bioinformatic approach detects putative regulatory elements in gene promoters.

BACKGROUND: Searching for approximate patterns in large promoter sequences frequently produces an exceedingly high numbers of results. Our aim was to exploit biological knowledge for definition of a sheltered search space and of appropriate search parameters, in order to develop a method for identification of a tractable number of sequence motifs. RESULTS: Novel software (COOP) was developed for extraction of sequence motifs, based on clustering of exact or approximate patterns according to the frequency of their overlapping occurrences. Genomic sequences of 1 Kb upstream of 91 genes differentially expressed and/or encoding proteins with relevant function in adult human retina were analyzed. Methodology and results were tested by analysing 1,000 groups of putatively unrelated sequences, randomly selected among 17,156 human gene promoters. When applied to a sample of human promoters, the method identified 279 putative motifs frequently occurring in retina promoters sequences. Most of them are localized in the proximal portion of promoters, less variable in central region than in lateral regions and similar to known regulatory sequences. COOP software and reference manual are freely available upon request to the Authors. CONCLUSION: The approach described in this paper seems effective for identifying a tractable number of sequence motifs with putative regulatory role.

Novel genes, possibly relevant for molecular diagnosis or therapy of human rhabdomyosarcoma, detected by genomic expression profiling.

Transcriptional profiles of an alveolar rhabdomyosarcoma (RMS) and of a RMS cell line were reconstructed by a computational and statistical approach. Expression data of 29,963 genes in 11 adult human healthy tissues and in 37 tumour tissues were analysed for comparison. We identified 202 genes differentially expressed in at least one RMS sample, as compared with normal skeletal muscle. Among them, 107 resulted specifically overexpressed in RMS, but in no tumour affecting other tissues. Cluster analysis applied to expression data detected a series of genes presumably co-expressed with genes encoding known tumour markers and/or reportedly involved in genesis or development of rhabdomyosarcoma. This study succeeded in identifying a number of genes, which become candidates for in vitro study, thus facilitating discovery of novel tumour markers or targets for drug therapy.

Screening for ryanodine receptor type 2 mutations in families with effort-induced polymorphic ventricular arrhythmias and sudden death: early diagnosis of asymptomatic carriers.

OBJECTIVES: We sought to establish the role of genetic screening for ryanodine receptor type 2 (RyR2) gene mutations in families with effort-induced polymorphic ventricular arrhythmia (PVA), syncope and juvenile sudden death. BACKGROUND: The RyR2 mutations have been associated with PVA, syncope and sudden death in response to physical or emotional stress. METHODS: We studied 81 subjects (39 males and 42 females; mean age 31 +/- 20 years) belonging to eight families with pathogenic RyR2 mutations. All subjects underwent screening for RyR2 mutations, electrocardiography (ECG), 24-h Holter monitoring, signal-averaged electrocardiography (SAECG), two-dimensional echocardiography and exercise stress testing. Electrophysiologic (EP) study was performed in nine patients. RESULTS: Six different RyR2 mutations were found in eight families. Forty-three family members carried the gene mutation. Of these, 28 (65%) showed effort-induced arrhythmic symptoms or signs and one died suddenly during follow-up. Family history revealed 19 juvenile cases of sudden death during effort or emotion. In two families sharing the same mutation, no subject presented with PVA during the stress test; thus, sudden death and syncope were the only clinical manifestations. The 12-lead ECG was normal in all but two subjects, whereas five patients showed positive late potentials on the SAECG. In 17 (39.5%) of 43 subjects, the two-dimensional echocardiogram revealed localized kinetic abnormalities and mild structural alterations of the right ventricle. The EP study was not able to induce PVA. CONCLUSIONS: The absence of symptoms and PVA on the stress test in more than one-third of carriers of RyR2 mutations, as well as the lack of PVA inducibility by the EP study, underlies the importance of genetic screening for the early diagnosis of asymptomatic carriers and prevention of sudden death.

Juvenile sudden death in a family with polymorphic ventricular arrhythmias caused by a novel RyR2 gene mutation: evidence of specific morphological substrates.

We report on a family with a history of sudden death and effort-induced polymorphic ventricular arrhythmias. The index case was a 17-year-old boy who died suddenly and at postmortem had evidence of fibrofatty replacement in the right ventricular free wall, consistent with arrhythmogenic right ventricular cardiomyopathy, as well as calcium phosphate deposits within the myocytes. A molecular genetics investigation carried out in the paraffin-embedded myocardium of the subject and in blood samples of family members disclosed a missense mutation in exon 3 (230C-->T; A77V) of the cardiac ryanodine receptor type 2 gene. The carriers showed effort-induced polymorphic ventricular tachycardia in the setting of normal resting electrocardiogram and trivial echocardiographic abnormalities, consistent with catecholaminergic polymorphic ventricular tachycardia. The observation of both arrhythmogenic right ventricular cardiomyopathy type 2 and catecholaminergic polymorphic ventricular tachycardia in the same family suggests that the two entities might correspond to different degrees of phenotypic expression of the same disease. This experience underscores the importance of a precise autopsy diagnosis in the case of sudden cardiac death, including molecular genetics, and the mission of pathologists to guide further clinical investigation of family members.

Detection of chromosomal regions showing differential gene expression in human skeletal muscle and in alveolar rhabdomyosarcoma.

BACKGROUND: Rhabdomyosarcoma is a relatively common tumour of the soft tissue, probably due to regulatory disruption of growth and differentiation of skeletal muscle stem cells. Identification of genes differentially expressed in normal skeletal muscle and in rhabdomyosarcoma may help in understanding mechanisms of tumour development, in discovering diagnostic and prognostic markers and in identifying novel targets for drug therapy. RESULTS: A Perl-code web client was developed to automatically obtain genome map positions of large sets of genes. The software, based on automatic search on Human Genome Browser by sequence alignment, only requires availability of a single transcribed sequence for each gene. In this way, we obtained tissue-specific chromosomal maps of genes expressed in rhabdomyosarcoma or skeletal muscle. Subsequently, Perl software was developed to calculate gene density along chromosomes, by using a sliding window. Thirty-three chromosomal regions harbouring genes mostly expressed in rhabdomyosarcoma were identified. Similarly, 48 chromosomal regions were detected including genes possibly related to function of differentiated skeletal muscle, but silenced in rhabdomyosarcoma. CONCLUSION: In this study we developed a method and the associated software for the comparative analysis of genomic expression in tissues and we identified chromosomal segments showing differential gene expression in human skeletal muscle and in alveolar rhabdomyosarcoma, appearing as candidate regions for harbouring genes involved in origin of alveolar rhabdomyosarcoma representing possible targets for drug treatment and/or development of tumor markers.

IDEG6: a web tool for detection of differentially expressed genes in multiple tag sampling experiments.

Here we present a novel web tool for the statistical analysis of gene expression data in multiple tag sampling experiments. Differentially expressed genes are detected by using six different test statistics. Result tables, linked to the GenBank, UniGene, or LocusLink database, can be browsed or searched in different ways. Software is freely available at the site: http://telethon.bio.unipd.it/bioinfo/IDEG6_form/, together with additional information on statistical methodologies.

Arrhythmogenic right ventricular cardiomyopathy type 1 (ARVD1): confirmation of locus assignment and mutation screening of four candidate genes.

Arrhythmogenic right ventricular cardiomyopathy type 1 (ARVD1) is an autosomal dominant disorder characterised by progressive degeneration of right ventricular myocardium, arrhythmias and risk of sudden death. By linkage analysis, we previously mapped the involved gene to chromosome 14q24.3. In the present study we report on linkage analysis of one additional and unrelated family, which enabled to confirm previous locus assignment. Another family is reported, in which genetic and clinical data suggest linkage to the same locus. Direct sequencing of DNA from individuals belonging to established ARVD1 families failed to detect causative mutations in exonic sequences of four genes (POMT2, TGFbeta3, KIAAA1036 and KIAA0759) expressed in the heart and which defects could possibly induce plasma membrane instability or apoptosis, key features of ARVD pathogenesis.

TAIL1: an isthmin-like gene, containing type 1 thrombospondin-repeat and AMOP domain, mapped to ARVD1 critical region.

Isthmins represent a novel family of vertebrate secreted proteins containing one copy of the thrombospondin type 1 repeat (TSR), which in mammals is shared by several proteins with diverse biological functions, including cell adhesion, angiogenesis, and patterning of developing nervous system. We have determined the genomic organization of human TAIL1 (thrombospondin and AMOP containing isthmin-like 1), a novel isthmin-like gene encoding a protein that contains a TSR and a C-terminal AMOP domain (adhesion-associated domain in MUC4 and other proteins), characteristic of extracellular proteins involved in adhesion processes. TAIL1 gene encompasses more than 24.4 kb. Analysis of the DNA sequence surrounding the putative transcriptional start region revealed a TATA-less promoter located in a CpG island. Several consensus binding sites for the transcription factors Sp1 and MZF-1 were identified in this promoter region. In humans, TAIL1 gene is located on chromosome 14q24.3 within ARVD1 (arrhythmogenic right ventricular dysplasia/cardiomyopathy, type 1) critical region; preliminary evidence suggests that it is expressed in several tissues, showing multiple alternative splicing.