Functional characterization of CNOT3 variants identified in familial adenomatous polyposis adenomas.

Germline mutations in the tumor suppressor () define Familial Adenomatous Polyposis (FAP), the genetic predisposition to developing adenomatous polyps. Recent sequencing of FAP adenomas have challenged established dogma that mutations alone represent the adenoma mutational landscape because recurrent somatic mutations in non-WNT pathway genes were also discovered. In particular, one of these novel genes, , presented E20K and E70K mutations that are predicted to be deleterious . We utilized zebrafish embryos to determine if these mutations affect function and perform novel biology in an APC-dependent pathway . Human () and mRNA injection rescued zebrafish knockdown lordosis phenotype while did not. In the FAP zebrafish model, we show that , but not retinoic acid, regulates expression. Injection of and , but not , to homozygous zebrafish initiated gut differentiation while knockdown in wildtype embryos led to decreased intestinal development and differentiation. Finally, targeted sequencing of 37 additional FAP adenomas revealed mutations in 20% of these samples. Overall, our work supports a mechanism where regulates and that overall perturbation could work in concert with germline mutations in advancing adenomas to a more transformed state prior to progression to adenocarcinoma.