PROJECT SUMMARY How much does function vary in individuals bearing WT genes and what is the level of function when only one WT allele is present? Addressing these questions has taken on new relevance in the field of cystic fibrosis (CF), one of the most common lethal autosomal recessive single gene disorders, due to recent large-scale epidemiologic studies reporting significant excess of CF-like respiratory symptoms in CF carriers. The findings are consistent with prior reports indicating that CF carriers have increased rates of sinus and lung disease, even though other features of CF such as sweat chloride concentration are not abnormal. The medical implications are substantial as there are an estimated 10 million CF carriers in the U.S. and an increasing fraction are being identified during prenatal screening or by direct to consumer testing. As testing moves toward sequencing of the CFTR gene, the detection of hypomorphic variants that reduce CFTR function but do not cause CF creates uncertainty for the individual, family and medical team. Treatment of symptomatic CF carriers with effective drugs that target CFTR, the protein defective in CF is conceivable. Although we know a considerable amount about those affected with CF, the range of CFTR function in non-carrier individuals and in CF carriers has yet to be determined. We propose to assess CFTR function in nasal epithelial cell cultures derived from individual subjects. The nasal epithelium (NE) is a native location for CFTR expression and conditionally reprogrammed cells derived from the NE form euploid monolayers that retain respiratory airway cell characteristics. We will apply our experience in culturing nasal epithelial cells and assessing CFTR function, insight into partial function variants and response to CFTR modulators in individuals with CF and CFTR-related disorders to CF carriers and non-carriers. The overall goal of this project is to determine if variation in CFTR function and/or presence of hypomorphic CFTR variants correlates with CF-like symptoms in CF carriers and if CFTR modulators might be effective treatment for symptomatic CF carriers. Four aims are proposed: Aim 1. To determine if CFTR chloride transport function differs between healthy non-CF carriers and asymptomatic CF carriers. Aim 2. To determine if CFTR chloride transport function differs between symptomatic and asymptomatic CF carriers. Aim 3: To assess whether the frequency of hypomorphic CFTR variants differs between asymptomatic and symptomatic CF carriers. Aim 4. To determine CFTR function in CF carriers in response to a CFTR modulator. Pursuit of these aims will address the molecular and medical implications of carrier status for CF that will inform counseling and management of millions of CF carriers worldwide.