Integrating genomic and deep clinical data to facilitate discovery of novel genes and disease mechanisms in rare diseases 

De novo indels are a recurrent cause of Robinow Syndrome (RS), a skeletal dysplasia accompanied by facial cleft, macrocephaly, and cardiac malformations. The molecular cause of the dominant form remained mostly unknown for 50 years. The Carvalho Lab teamed up with the Centers for Mendelian Genomics (CMG) to perform high-coverage, exome sequencing in probands with RS and could finally resolve the mystery ¹. 

About 30% of the dominant form of the disease is caused by indels at specific exons affecting one of the three disheveled paralogous, DVL1, DVL2 and DVL3, none of them ever reported in disease before. DVL is a conserved key mediator of the Wnt pathway acting in downstream developmental signaling branches, including the planar cell polarity pathway (PCP) which have key roles in cell migration and bone formation. 

Intriguingly, ALL RS individuals harbor distinct genomic variants resulting in -1 frameshifting of the reading frame of the penultimate or last exons in DVL1 or DVL3. Those variants produce long, truncated C-termini with at least 85 amino acids replaced with a novel, highly basic tail, for which contribution to PCP perturbation remains unknown. The discovery of a group of genes that cause genetic diseases only when carrying pathogenic alleles created by a specific frameshift change was an important scientific discovery with implications to how the genetic field interprets indels. For instance, those genes have characteristically low pLI scores (pLI scores represent the tolerance of a gene or transcript to DNA variants leading protein truncation), contrary to expected for heterozygous pathogenic alleles which leads them to be overlooked as candidates for dominant diseases. Other genes discovered to cause RS in the same cohort include FZD2 and NXN.

The Carvalho Lab’s goal for this project is to evaluate the functional effects of pathogenic DNA variants in human development and cell maintenance. This PNRI lab is investigating variants affecting DVL1, DVL2 and DVL3 leading to Robinow Syndrome given their intriguing disease mechanism. The goal is to identify cellular readouts and disease biomarkers that can help informing pathogenicity of variants of uncertain significance (VUS) in patient cohorts with overlapping clinical phenotypes. 

c.

Figure –

The Carvalho Lab is investigating the function of indels as an underlying cause of genetic disease and developmental malformations.

a, b. DVL1 and DVL3 are now classic examples of transcripts that can cause diseases exclusively due to the presence of indels alleles that escape nonsense mediated decay (NMD)^2-7;

c. HEK293T cells transfected with mutant and wild-type DVL1 (left plot) and DVL3 (right plot). Wild-type and -1 frameshifted proteins stimulated with WNT ligands accumulate (blue bars) in WT cells but stay as punctate in the presence of truncated DVL proteins. NMD^– escaping DVLs does not seem to respond to WNT signaling as the wild-type counterpart according to this experiment, which may help explain the developmental phenotypes observed in patients with Robinow Syndrome. Unpublished preliminary data.