Mutations in TMEM260 Cause a Pediatric Neurodevelopmental, Cardiac, and Renal Syndrome

Abstract

Despite the accelerated discovery of genes associated with syndromic traits, the majority of families affected by such conditions remain undiagnosed. Here, we employed whole-exome sequencing in two unrelated consanguineous kindreds with central nervous system (CNS), cardiac, renal, and digit abnormalities. We identified homozygous truncating mutations in TMEM260, a locus predicted to encode numerous splice isoforms. Systematic expression analyses across tissues and developmental stages validated two such isoforms, which differ in the utilization of an internal exon. The mutations in both families map uniquely to the long isoform, raising the possibility of an isoform-specific disorder. Consistent with this notion, RT-PCR of lymphocyte cell lines from one of the kindreds showed reduced levels of only the long isoform, which could be ameliorated by emetine, suggesting that the mutation induces nonsense-mediated decay. Subsequent in vivo testing supported this hypothesis. First, either transient suppression or CRISPR/Cas9 genome editing of zebrafish tmem260 recapitulated key neurological phenotypes. Second, co-injection of morphants with the long human TMEM260 mRNA rescued CNS pathology, whereas the short isoform was significantly less efficient. Finally, immunocytochemical and biochemical studies showed preferential enrichment of the long TMEM260 isoform to the plasma membrane. Together, our data suggest that there is overall reduced, but not ablated, functionality of TMEM260 and that attenuation of the membrane-associated functions of this protein is a principal driver of pathology. These observations contribute to an appreciation of the roles of splice isoforms in genetic disorders and suggest that dissection of the functions of these transcripts will most likely inform pathomechanism.Whole-exome and whole-genome sequencing (WES and WGS, respectively) have accelerated the expansion of the repertoire of human loci underscoring Mendelian phenotypes to account for~3,000 human genes (15%).1 Nonetheless, the diagnostic yield remains <50%, 2-4 suggesting that the majority of disease-causing lesions are yet to be identified and that a significant fraction of this genetic burden is likely to be contributed by ultra-rare or private mutations. To contribute to the saturation of the morbid human genome as a means of improving both diagnostic value and mechanistic insight, we undertook the systematic sequencing of families affected by suspected genetic disorders, with an emphasis on congenital syndromic traits. Here, we report the genetic and functional dissection of affected individuals with a genetically undiagnosed syndrome characterized by cardiac, CNS, renal, and digit abnormalities in two unrelated consanguineous pedigrees. We show that the likely molecular cause, homozygous truncating mutations in TMEM260, affect the functions of one of the two established isoforms transcribed from this locus, thereby demonstrating the necessity of the long isoform for the development of multiple tissues. We consulted family 1, a con...

Haploinsufficiency of the Chromatin Remodeler BPTF Causes Syndromic Developmental and Speech Delay, Postnatal Microcephaly, and Dysmorphic Features

“…We and others have shown previously that neuroanatomical defects can be modeled in zebrafish as a direct readout for analogous defects observed in humans. 31,[44][45][46][47] We designed a CRISPR single-guide RNA (sgRNA) targeting exon 9 of bptf, injected 75 pg sgRNA and 150 pg of CAS9 into the cell of 1-cellstage zebrafish embryos, and detected efficient disruption of the locus through the introduction of small insertions or deletions (90% mosaicism; Figure S1, Table S3). First, we asked whether we could detect any differences in the head size of 3 dpf F0 mutant larvae.…”

Bi-allelic Variants in DYNC1I2 Cause Syndromic Microcephaly with Intellectual Disability, Cerebral Malformations, and Dysmorphic Facial Features

“…Individuals in each pedigree (1-VI:1, 2-II-1, and 3-II-1) showed hypogenesis or agenesis of the corpus callosum ( Figure 1E and Table 1), so we first measured the number of commissural axon tracts connecting the zebrafish optic tecta in controls, dync1i2a MO-injected animals, and animals in-jected with MO plus WT DYNC1I2 mRNA, as described. 46 We observed a modest but significant reduction in the number of axon tracts crossing the dorsal midline in MO-injected batches (p < 0.0001 versus controls), and this defect could be rescued significantly by the presence of WT mRNA (p < 0.0001 versus MO alone; replicate batches; Figures S7A and S7B). Additionally, we measured the size of the optic tecta, neuron-rich sensory processing centers in the zebrafish brain whose size correlates with overall head size.…”

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