Dataset Information

Monoallelic Mutations in CC2D1A Suggest a Novel Role in Human Heterotaxy and Ciliary Dysfunction.

ABSTRACT:

Background

Human heterotaxy is a group of congenital disorders characterized by misplacement of one or more organs according to the left-right axis. The genetic causes of human heterotaxy are highly heterogeneous.

Methods

We performed exome sequencing in a cohort of 26 probands with heterotaxy followed by gene burden analysis for the enrichment of novel rare damaging mutations. Transcription activator-like effector nuclease was used to generate somatic loss-of-function mutants in a zebrafish model. Ciliary defects were examined by whole-mount immunostaining of acetylated α-tubulin.

Results

We identified a significant enrichment of novel rare damaging mutations in the CC2D1A gene. Seven occurrences of CC2D1A mutations were found to affect 4 highly conserved amino acid residues of the protein. Functional analyses in the transcription activator-like effector nuclease-mediated zebrafish knockout models were performed, and heterotaxy phenotypes of the cardiovascular and gastrointestinal systems in both somatic and germline mutants were observed. Defective cilia were demonstrated by whole-mount immunostaining of acetylated α-tubulin. These abnormalities were rescued by wild-type cc2d1a mRNA but not cc2d1a mutant mRNA, strongly suggesting a loss-of-function mechanism. On the other hand, overexpression of cc2d1a orthologous mutations cc2d1a P559L and cc2d1a G808V (orthologous to human CC2D1A P532L and CC2D1A G781V) did not affect embryonic development.

Conclusions

Using a zebrafish model, we were able to establish a novel association of CC2D1A with heterotaxy and ciliary dysfunction in the F2 generation via a loss-of-function mechanism. Future mechanistic studies are needed for a better understanding of the role of CC2D1A in left-right patterning and ciliary dysfunction.

SUBMITTER: Ma ACH

PROVIDER: S-EPMC7748040 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Publications

Monoallelic Mutations in <i>CC2D1A</i> Suggest a Novel Role in Human Heterotaxy and Ciliary Dysfunction.

Ma Alvin Chun Hang ACH Mak Christopher Chun Yu CCY Yeung Kit San KS Pei Steven Lim Cho SLC Ying Dingge D Yu Mullin Ho Chung MHC Hasan Kazi Md Mahmudul KMM Chen Xiangke X Chow Pak Cheong PC Cheung Yiu Fai YF Chung Brian Hon Yin BHY

Circulation. Genomic and precision medicine 20201116 6

<h4>Background</h4>Human heterotaxy is a group of congenital disorders characterized by misplacement of one or more organs according to the left-right axis. The genetic causes of human heterotaxy are highly heterogeneous.<h4>Methods</h4>We performed exome sequencing in a cohort of 26 probands with heterotaxy followed by gene burden analysis for the enrichment of novel rare damaging mutations. Transcription activator-like effector nuclease was used to generate somatic loss-of-function mutants in ...[more]

PMID: 33196317

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Project description:Heterotaxy, a birth defect involving left-right patterning defects, and primary ciliary dyskinesia (PCD), a sinopulmonary disease with dyskinetic/immotile cilia in the airway are seemingly disparate diseases. However, they have an overlapping genetic etiology involving mutations in cilia genes, a reflection of the common requirement for motile cilia in left-right patterning and airway clearance. While PCD is a monogenic recessive disorder, heterotaxy has a more complex, largely non-monogenic etiology. In this study, we show mutations in the novel dynein gene DNAH6 can cause heterotaxy and ciliary dysfunction similar to PCD. We provide the first evidence that trans-heterozygous interactions between DNAH6 and other PCD genes potentially can cause heterotaxy. DNAH6 was initially identified as a candidate heterotaxy/PCD gene by filtering exome-sequencing data from 25 heterotaxy patients stratified by whether they have airway motile cilia defects. dnah6 morpholino knockdown in zebrafish disrupted motile cilia in Kupffer's vesicle required for left-right patterning and caused heterotaxy with abnormal cardiac/gut looping. Similarly DNAH6 shRNA knockdown disrupted motile cilia in human and mouse respiratory epithelia. Notably a heterotaxy patient harboring heterozygous DNAH6 mutation was identified to also carry a rare heterozygous PCD-causing DNAI1 mutation, suggesting a DNAH6/DNAI1 trans-heterozygous interaction. Furthermore, sequencing of 149 additional heterotaxy patients showed 5 of 6 patients with heterozygous DNAH6 mutations also had heterozygous mutations in DNAH5 or other PCD genes. We functionally assayed for DNAH6/DNAH5 and DNAH6/DNAI1 trans-heterozygous interactions using subthreshold double-morpholino knockdown in zebrafish and showed this caused heterotaxy. Similarly, subthreshold siRNA knockdown of Dnah6 in heterozygous Dnah5 or Dnai1 mutant mouse respiratory epithelia disrupted motile cilia function. Together, these findings support an oligogenic disease model with broad relevance for further interrogating the genetic etiology of human ciliopathies.