Characterising the loss-of-function impact of 5' untranslated region variants in 15,708 individuals.

Upstream open reading frames (uORFs) are tissue-specific cis-regulators of protein translation. Isolated reports have shown that variants that create or disrupt uORFs can cause disease. Here, in a systematic genome-wide study using 15,708 whole genome sequences, we show that variants that create new upstream start codons, and variants disrupting stop sites of existing uORFs, are under strong negative selection. This selection signal is significantly stronger for variants arising upstream of genes intolerant to loss-of-function variants. Furthermore, variants creating uORFs that overlap the coding sequence show signals of selection equivalent to coding missense variants. Finally, we identify specific genes where modification of uORFs likely represents an important disease mechanism, and report a novel uORF frameshift variant upstream of NF2 in neurofibromatosis. Our results highlight uORF-perturbing variants as an under-recognised functional class that contribute to penetrant human disease, and demonstrate the power of large-scale population sequencing data in studying non-coding variant classes.

ZBTB17 (MIZ1) Is Important for the Cardiac Stress Response and a Novel Candidate Gene for Cardiomyopathy and Heart Failure.

BACKGROUND: Mutations in sarcomeric and cytoskeletal proteins are a major cause of hereditary cardiomyopathies, but our knowledge remains incomplete as to how the genetic defects execute their effects. METHODS AND RESULTS: We used cysteine and glycine-rich protein 3, a known cardiomyopathy gene, in a yeast 2-hybrid screen and identified zinc-finger and BTB domain-containing protein 17 (ZBTB17) as a novel interacting partner. ZBTB17 is a transcription factor that contains the peak association signal (rs10927875) at the replicated 1p36 cardiomyopathy locus. ZBTB17 expression protected cardiac myocytes from apoptosis in vitro and in a mouse model with cardiac myocyte-specific deletion of Zbtb17, which develops cardiomyopathy and fibrosis after biomechanical stress. ZBTB17 also regulated cardiac myocyte hypertrophy in vitro and in vivo in a calcineurin-dependent manner. CONCLUSIONS: We revealed new functions for ZBTB17 in the heart, a transcription factor that may play a role as a novel cardiomyopathy gene.

16p11.2-p12.2 duplication syndrome; a genomic condition differentiated from euchromatic variation of 16p11.2.

Chromosome 16 contains multiple copy number variations (CNVs) that predispose to genomic disorders. Here, we differentiate pathogenic duplications of 16p11.2-p12.2 from microscopically similar euchromatic variants of 16p11.2. Patient 1 was a girl of 18 with autism, moderate intellectual disability, behavioural difficulties, dysmorphic features and a 7.71-Mb (megabase pair) duplication (16:21 521 005-29 233 146). Patient 2 had a 7.81-Mb duplication (16:21 382 561-29 191 527), speech delay and obsessional behaviour as a boy and, as an adult, short stature, macrocephaly and mild dysmorphism. The duplications contain 65 coding genes of which Polo-like kinase 1 (PLK1) has the highest likelihood of being haploinsufficient and, by implication, a triplosensitive gene. An additional 1.11-Mb CNV of 10q11.21 in Patient 1 was a possible modifier containing the G-protein-regulated inducer of neurite growth 2 (GPRIN2) gene. In contrast, the euchromatic variants in Patients 3 and 4 were amplifications from a 945-kb region containing non-functional immunoglobulin heavy chain (IGHV), hect domain pseudogene (HERC2P4) and TP53-inducible target gene 3 (TP53TG3) loci in proximal 16p11.2 (16:31 953 353-32 898 635). Paralogous pyrosequencing gave a total copy number of 3-8 in controls and 8 to >10 in Patients 3 and 4. The 16p11.2-p12.2 duplication syndrome is a recurrent genomic disorder with a variable phenotype including developmental delay, dysmorphic features, mild to severe intellectual disability, autism, obsessive or stereotyped behaviour, short stature and anomalies of the hands and fingers. It is important to differentiate pathogenic 16p11.2-p12.2 duplications from harmless, microscopically similar euchromatic variants of proximal 16p11.2, especially at prenatal diagnosis.

Next generation diagnostics in inherited arrhythmia syndromes : a comparison of two approaches.

Next-generation sequencing (NGS) provides an unprecedented opportunity to assess genetic variation underlying human disease. Here, we compared two NGS approaches for diagnostic sequencing in inherited arrhythmia syndromes. We compared PCR-based target enrichment and long-read sequencing (PCR-LR) with in-solution hybridization-based enrichment and short-read sequencing (Hyb-SR). The PCR-LR assay comprehensively assessed five long-QT genes routinely sequenced in diagnostic laboratories and "hot spots" in RYR2. The Hyb-SR assay targeted 49 genes, including those in the PCR-LR assay. The sensitivity for detection of control variants did not differ between approaches. In both assays, the major limitation was upstream target capture, particular in regions of extreme GC content. These initial experiences with NGS cardiovascular diagnostics achieved up to 89 % sensitivity at a fraction of current costs. In the next iteration of these assays we anticipate sensitivity above 97 % for all LQT genes. NGS assays will soon replace conventional sequencing for LQT diagnostics and molecular pathology.