A Drosophila Su(H) model of Adams-Oliver Syndrome reveals cofactor titration as a mechanism underlying developmental defects.

Notch signaling is a conserved pathway that converts extracellular receptor-ligand interactions into changes in gene expression via a single transcription factor (CBF1/RBPJ in mammals; Su(H) in Drosophila). In humans, RBPJ variants have been linked to Adams-Oliver syndrome (AOS), a rare autosomal dominant disorder characterized by scalp, cranium, and limb defects. Here, we found that a previously described Drosophila Su(H) allele encodes a missense mutation that alters an analogous residue found in an AOS-associated RBPJ variant. Importantly, genetic studies support a model that heterozygous Drosophila with the AOS-like Su(H) allele behave in an opposing manner to heterozygous flies with a Su(H) null allele, due to a dominant activity of sequestering either the Notch co-activator or the antagonistic Hairless co-repressor. Consistent with this model, AOS-like Su(H) and Rbpj variants have decreased DNA binding activity compared to wild type proteins, but these variants do not significantly alter protein binding to the Notch co-activator or the fly and mammalian co-repressors, respectively. Taken together, these data suggest a cofactor sequestration mechanism underlies AOS phenotypes associated with RBPJ variants, whereby the AOS-associated RBPJ allele encodes a protein with compromised DNA binding activity that retains cofactor binding, resulting in Notch target gene dysregulation. Author summary: Adams-Oliver Syndrome (AOS) is a rare disease defined by missing skin/skull tissue, limb malformations, and cardiovascular abnormalities. Human genetic studies have revealed that ~40% of AOS patients inherit dominant mutations within specific genes in the Notch signaling pathway. Notch signaling is a highly conserved cell-to-cell communication pathway found in all metazoans and plays crucial roles during embryogenesis and tissue homeostasis in organisms from Drosophila (fruit-flies) to mammals. The Notch receptor converts cell-to-cell interactions into a Notch signal that enters the nucleus and activates target genes by binding to a highly conserved transcription factor. Here, we took advantage of the unexpected finding that a previously described dominant allele in the Drosophila Notch pathway transcription factor contains a missense variant in an analogous residue found in a family with AOS. Using this novel animal model of AOS along with biochemical DNA binding, protein-protein interaction, and transcriptional reporter assays, we found that this transcription factor variant selectively compromises DNA binding but not binding to the Notch signal nor binding to other proteins in the Notch pathway. Taken together with prior human genetic studies, these data suggest AOS phenotypes associated with variants in the Notch pathway transcription factor are caused by a dominant mechanism that sequesters the Notch signal, leading to Notch target gene dysregulation. [ABSTRACT FROM AUTHOR]

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