RT Journal Article SR Electronic T1 Interpreting the pathogenicity of Joubert Syndrome missense variants in Caenorhabditis elegans JF bioRxiv FD Cold Spring Harbor Laboratory SP 2020.05.22.110668 DO 10.1101/2020.05.22.110668 A1 Karen I. Lange A1 Sofia Tsiropoulou A1 Katarzyna Kucharska A1 Oliver E. Blacque YR 2020 UL http://biorxiv.org/content/early/2020/05/22/2020.05.22.110668.abstract AB Ciliopathies are a large class of inherited disorders caused by defects in cilia. Variants in ciliopathy genes are highly pleiotropic and represent excellent case studies for interrogating genotype-phenotype correlation. However, in most cell and animal studies, ciliopathy mechanisms are not investigated using alleles as they occur in patients. We have employed Caenorhabditis elegans to model and characterise two pathogenic biallelic missense variants in B9D2/mksr-2 associated with Joubert Syndrome (JBTS). B9D2 functions within the MKS module at the transition zone (TZ) ciliary subcompartment, and regulates the cilium’s molecular composition and signaling function. Using CRISPR-Cas9 genome editing with a novel mutation detection strategy, P74S and G155S mutations were engineered in mksr-2. Quantitative assays of cilium/TZ structure and function, as well as knock-in reporters expressed at endogenous levels, revealed that the mutations act recessively and differentially. Whilst both variants disrupt the cilium/TZ and MKS module assembly, G155S causes a more severe overall phenotype, approaching that of a null allele. In addition, G155S, but not P74S, severely disrupts endogenous MKSR-2 organisation at the TZ. Recapitulation of the JBTS patient biallelic genotype showed that compound heterozygous worms (P74S/G155S) phenocopy worms recessive for P74S. Finally, this study provides new insight into the MKS module assembly mechanism and organisation, revealing a close functional association between the B9 complex and TMEM216/MKS-2. Together, these data provide a thorough quantitative assessment of JBTS alleles, establish C. elegans as a paradigm for interpreting TZ ciliopathy mutations, and provide insight into MKS module organisation at the TZ.Competing Interest StatementThe authors have declared no competing interest.