In silico and functional characterisation of an ultra-rare CFTR mutation identifies novel lasso motif interactions regulating channel gating

Characterisation of I37R – a novel mutation in the lasso motif of ABC-transporter CFTR, a chloride channel – was conducted by theratyping using CFTR potentiators which increase channel gating activity and correctors which repair protein trafficking defects. I37R-CFTR function was characterised using intestinal current measurements (ICM) in rectal biopsies, forskolin-induced swelling (FIS) in intestinal organoids and short circuit current measurements (Isc) in organoid-derived monolayers from an individual with I37R/F508del CFTR genotype. We demonstrated that the I37R-CFTR mutation results in a residual function defect amenable to treatment with potentiators and type III, but not to type I, correctors. Molecular dynamics of I37R-CFTR using an extended model of the phosphorylated, ATP-bound human CFTR identified an altered lasso motif conformation which results in an unfavourable strengthening of the interactions between the lasso motif, the regulatory (R) domain and the transmembrane domain two (TMD2). In conclusion, structural and functional characterisation of the I37R- CFTR mutation increases understanding of CFTR channel regulation and provides a potential pathway to access CFTR modulator treatments for individuals with CF caused by ultra-rare CFTR mutations. ### Competing Interest Statement SAW is the recipient of a Vertex Innovation Grant (2018) and a TSANZ/Vertex Research Award (2020). Both are unrelated and outside of the submitted manuscript. AJ has received consulting fees from Vertex on projects unrelated to this study. CYO has acted as consultant and is on advisory boards for Vertex pharmaceuticals. These works are unrelated to this project and manuscript. All other authors declare no conflict of interest.