Van der Woude syndrome (VWS) is an autosomal dominant disorder characterized by lower lip pits and orofacial clefts (OFCs). With a prevalence of approximately 1 in 35,000 live births, it is the most common form of syndromic clefting and may account for ∼2% of all OFCs. The majority of VWS is attributed to genetic variants in IRF6 (∼70%) or GRHL3 (∼5%), leaving up to 25% of individuals with VWS without a molecular diagnosis. Both IRF6 and GRHL3 function in a transcriptional regulatory network governing differentiation of periderm, a single layer of epithelial cells that prevents pathological adhesions during palatogenesis. Disruption of this layer results in a spectrum of phenotypes ranging from lip pits and OFCs to severe pterygia and other congenital anomalies that can be incompatible with life. Understanding the mechanisms of peridermopathies is vital in improving health outcomes for affected individuals. We reasoned that genes encoding additional members of the periderm gene regulatory network, including kinases acting upstream of IRF6 (i.e., atypical protein kinase C family members, RIPK4, and CHUK), are candidates to harbor variants resulting in VWS. Consistent with this prediction, we identified 6 de novo variants (DNs) and 11 rare variants in PRKCI, an atypical protein kinase C, in 17 individuals with clinical features consistent with syndromic OFCs and peridermopathies. Of the identified DNs, 4 were identical p.(Asn383Ser) variants in unrelated individuals with syndromic OFCs, indicating a likely hotspot mutation. We also performed functional validation of 12 variants using the enveloping layer in zebrafish embryos, a structure analogous to the periderm. Three patient-specific alleles (p.Arg130His, p.(Asn383Ser), and p.Leu385Phe) were confirmed to be loss-of-function variants. In summary, we identified PRKCI as a novel causal gene for VWS and syndromic OFC with other features of peridermopathies.

The authors have declared no competing interest.

Sequencing services for CPSeq were provided by the Center for Inherited Disease Research (CIDR). CIDR is fully funded through a federal contract from the National Institutes of Health to The Johns Hopkins University, contract number HHSN268201700006I. Additional whole genome sequencing was funded by National Institutes of Health (NIH) grants: X01-HL136465 (MLM: US, European trios), X01-HL132363 (MLM: Colombian trios), X01-DE0300701 (MLM: Latin American trios), and X01-HG010835 (EJL, MLM: CPseq trios). Patient recruitment, assembly of phenotypic information, and data analysis were supported by National Institutes of Health (NIH) grants: F31-DE032588 (KR), X01-HG010835 (EJL), R01 DE027983 (EJL), R01 DE028342 (EJL), R01 DE030342 (EJL), R01-DE028300 (AB), R01DE032710 (RJL), DE008559 (JCM), R01-DE016148 (MLM, SMW), R01-DE008559 (JCM, MLM), R01-DE032122 (MLM), R01-DE0332319 (MLM).

I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.

Yes

The details of the IRB/oversight body that provided approval or exemption for the research described are given below:

Ethics committee/IRB of Emory University gave ethical approval for this work.

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Sequence and phenotype data is available from the Database of Genotypes and Phenotypes (dbGaP, ncbi.nlm.nih.gov) under study accession # phs002220.v1.p1 (CPSeq, VWS probands), accession # phs001420.v1.p1 (Latin American), accession # phs001168.v2.p2 (US, European), accession # phs001997,v1.p1 (African, Asian). Additional information for probands in the GMKF cohort can be viewed at https://kidsfirstdrc.org/studies/.