Nonsense mutations, single nucleotide base changes that result in an in frame premature termination codon (PTC), make up ~20% of the ~43,000 disease associated mutations residing in gene coding regions and are responsible for ~11% of all inherited human diseases. Restoring functional protein by inducing readthrough of PTCs has been accomplished using aminoglycosides that have been considered as potential therapies. The overall efficacy of readthrough achieved with and the toxicity caused by currently available compounds has not been desirable. The goal of our project is identify genes that promote readthrough and we have employed cystic fibrosis, which is an autosomal recessive disease caused by loss of function of the cystic fibrosis transmembrane conductance regulator (CFTR), as model disease. Previously, we showed that Yor1 (Yeast Oligomycin Resistance-1, a member of the ABC gene family) could serve as a robust model for genome-wide phenomic analysis of CFTR. The approach demonstrated evolutionary conservation of gene-gene interactions between yeast and human for proteins that regulate CFTR-ΔF508 biogenesis; and resulted in discovery of novel targets for correction of CFTR misfolding. The CFTR-G542X mutation is the most common premature termination codon in CFTR, and the second most common CF-causing allele. We have introduced a homologous mutation of CFTR-G542X, G704X, in YOR1, and using growth on oligomycin as readout show that yor1-G704X behaves in a similar fashion to the yor1-null allele. This data indicates very little or no full-length protein is expressed, analogous to CFTR-G542X. In the presence of G418, an aminoglycoside shown to restore partial CFTR function in mammalian cell lines, we observed oligomycin resistance specifically mediated by the Yor1-G704X mutant, establishing that yor1- G704X is suitable as a reporter for genome-wide yeast phenomic screening and identification of molecular targets that allow readthrough of nonsense variants. Among the hits produced by our unbiased genome-wide phenotypic screen with Yor1-G704X are the members of nonsense mediated decay (NMD) pathway, Nam7 & Upf3. Suppression of NMD pathway was previously shown to lead to recovery of CFTR protein function. We identified some members of the 20S proteasome as modifiers of Yor1-G704X function. These candidates will be further tested in cell lines for CFTR protein function with siRNA knockdown. Resulting genes and pathways will serve as a basis for new therapeutic strategies to achieve functional CFTR expression in CF patients carrying premature stop codons.