Branchiootorenal spectrum disorders (BOS) are characterized by craniofacial defects that include malformation of branchial arches (BAs), external ear, middle ear, and inner ear; a subset of patients also has kidney defects. Two causative genes are associated with BOS diagnoses, but these genes account for fewer than half of patient cases: the SIX1 transcription factor and a co-factor protein, EYA1, which binds to SIX1 and modifies its transcriptional activity. Thus, the causative genes for over half of BOS patients are yet to be identified. We hypothesize that there are other key co-factor proteins that bind to SIX1 to regulate its activity, and that mutations in these co-factors contribute to the unknown causes of BOS. The goal of this research program is to identify, in tractable model systems, additional genes whose altered functions contribute to the craniofacial malformations of BOS so that these genes can ultimately be included in human genetic screening. Using the Drosophila interactome data for the fly homologue of Six1, we identified 11 novel putative co-factors in Xenopus and showed that most of these are expressed in the developing BAs, ear and kidney, and therefore are potentially relevant to BOS. These proteins are highly conserved in humans, and our preliminary data show that five of them (Sobp, Zmym2, Zmym4, 2G4, Mcrs1) are required for development of the embryonic precursors of the branchial arches (neural crest [NC]), middle ear (NC) and inner ear (otic placode).
In Aim 1, we will use gain- and loss-of-function approaches to determine whether these candidate cofactors play a role NC formation or migration, branchial arch cartilages or inner ear gene expression and formation.
In Aim 2, we will evaluate the biochemical interactions of these gene products with Six1 and whether they affect Six1 transcriptional function.
In Aim 3, we will determine whether the known BOS mutations in SIX1 affect candidate co-factor binding or function, and map what regions of the protein-protein interaction domains of Six1 and of each co-factor mediate binding and transcriptional activity. Our previous work and established model systems uniquely position us to validate whether these candidates are bone fide Six1 co-factors, and elucidate how they contribute to normal and dysmorphic craniofacial development. These analyses will provide important information that cannot be obtained from the limited patient material available. They also have the future potential to explain the phenotypic variability in BOS patients and provide a rationale for including new causative genes in BOS gene panels.

Public Health Relevance

Craniofacial abnormalities are among the most prevalent birth defect, and often are caused by mutations in genes that regulate the formation and differentiation of cranial sensory placodes and neural crest. Patients diagnosed with one of the Branchiootorenal spectrum of disorders (BOS), whose known causes are mutations in SIX1 or EYA1, suffer from craniofacial deformities and suffer from hearing loss and kidney defects. However, since SIX1 and EYA1 mutations only account for half of the patients, the proposed research will identify additional genes involved in BOS by demonstrating the cellular and molecular functions of novel proteins that cooperate with SIX1 during craniofacial development. !

National Institute of Health (NIH)
National Institute of Dental & Craniofacial Research (NIDCR)
Research Project (R01)
Project #
Application #
Study Section
Skeletal Biology Development and Disease Study Section (SBDD)
Program Officer
Stein, Kathryn K
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
George Washington University
Anatomy/Cell Biology
Schools of Medicine
United States
Zip Code
Sullivan, Charles H; Majumdar, Himani D; Neilson, Karen M et al. (2018) Six1 and Irx1 have reciprocal interactions during cranial placode and otic vesicle formation. Dev Biol :