FAM222B is proposed as a novel candidate disease gene for dominant cardiovascular laterality defects, supported by identification of six variants in affected individuals, zebrafish double-knockout showing aberrant cardiac looping, and human mRNA injection experiments demonstrating perturbed cardiogenesis.
Key Findings
Results
Six different variants in FAM222B were identified across individuals with cardiovascular laterality defects through exome sequencing and survey.
Initial discovery used exome sequencing in 16 case-parent trios.
Exome survey was conducted in 2,109 individuals with situs inversus totalis, heterotaxy, or isolated congenital heart defects (CHD).
The variant c.899G > A (p.300Arg > His) was found de novo in two unrelated families.
FAM222B had previously been discussed as a candidate gene for cerebral cavernous malformations, not cardiovascular laterality defects.
Results
Structural modelling suggests that FAM222B Arg300 plays a functional role in Nemo-like kinase (NLK) recognition.
FAM222B has been described as a substrate of Nemo-like kinase (NLK).
NLK is associated with left-right body axis determination.
The specific amino acid change p.300Arg > His is predicted by structural modelling to affect the NLK recognition site.
This mechanistic link provides a potential explanation for how FAM222B variants could disrupt cardiovascular laterality.
Results
Zebrafish double-knockout of fam222ba/bb resulted in aberrant cardiac looping in larvae and enlarged atrium and ventricle in adult zebrafish.
A double-knockout (dd-KO) fam222ba/bb zebrafish line was generated and analyzed.
Aberrant cardiac looping was identified in dd-KO larvae.
Adult dd-KO zebrafish displayed enlarged atrium and ventricle.
Whole-mount in situ hybridization (WISH) was used to characterize the expression pattern and genomic context of the zebrafish FAM222B homologues fam222ba/bb and fam222aa.
Results
Injection of human FAM222B c.899G > A variant mRNA into wildtype zebrafish reporter larvae led to perturbed cardiogenesis.
The c.899G > A variant was tested using human mRNA injections into Tg(kdrl:EGFP) wildtype reporter zebrafish.
This approach allowed visualization of cardiovascular development via the endothelial EGFP reporter.
The resulting perturbed cardiogenesis provides functional evidence that this specific variant is pathogenic rather than benign.
This experiment supports a dominant mechanism of disease, consistent with de novo occurrence in two unrelated families.
Background
Cardiovascular laterality defects occur with an estimated birth prevalence of 1.1/10,000 live births, and known disease genes explain only about 20% of all cases.
The condition is associated with congenital heart defects (CHD) and situs abnormalities.
Known disease genes are often correlated with primary ciliary dyskinesia (PCD).
The study aimed to identify disease genes beyond PCD-related aetiologies.
The low explanatory rate of known genes (approximately 20%) motivated the search for novel candidate genes such as FAM222B.
What This Means
This research suggests that a gene called FAM222B is a new genetic cause of cardiovascular laterality defects — conditions where the heart and other organs are positioned abnormally in the body. These defects affect roughly 1 in 10,000 newborns and are often associated with serious heart malformations. Previously, scientists could only identify the genetic cause in about 20% of cases, mostly linked to a condition called primary ciliary dyskinesia. By analyzing the genomes of affected individuals and their parents, the researchers found six different variants in FAM222B among people with these heart and organ positioning defects, including one specific change (p.Arg300His) that appeared spontaneously — not inherited from parents — in two unrelated families.
To test whether FAM222B is truly responsible for these defects, the researchers used zebrafish as a model organism. When they knocked out the zebrafish versions of this gene, the fish larvae developed abnormal heart looping (the normal twisting of the heart during development), and adult fish had enlarged heart chambers. Additionally, when they injected zebrafish embryos with the human disease-associated version of the FAM222B gene, the heart development was disrupted. The gene appears to work through a protein called Nemo-like kinase (NLK), which is known to help determine left-right body symmetry, and computer modeling suggests the disease-causing variant interferes with this interaction.
This research matters because it identifies a new genetic explanation for cardiovascular laterality defects that operates independently of the previously known causes. This could help improve genetic diagnosis for affected families and potentially inform future understanding of how the heart and organs establish their correct positions during embryonic development. The finding that the same variant arose independently in two unrelated families strengthens the case that FAM222B is a genuine disease gene rather than an incidental finding.