Human disease variants of KATNIP fail to support CILK1 activation and control of primary cilia.

Pathogenic variants in KATNIP (encoding katanin-interacting protein) are linked to Joubert syndrome, a prototypical ciliopathy. KATNIP is a scaffold protein that binds and potentiates ciliogenesis-associated kinase 1 (CILK1) activation and function to control cilia length and frequency. We previously showed that of the three predicted 'domains of unknown functions' (DUFs) in KATNIP, the DUF2 domain alone supports binding to CILK1 without activating CILK1. Here, we report three human disease variants of KATNIP with different lengths that exhibit loss of function. The longest variant of KATNIP M1474C, which is truncated near the C-terminus, binds to CILK1 but does not support the activating TDY phosphorylation in CILK1, the phosphorylation of CILK1 substrates, or the restriction of cilia length and ciliation rate. Deletion analysis of KATNIP further revealed that residues 1524-1573 encompassing predicted β-sheets and an α-helix are essential for CILK1 activation and function. The results support a model where KATNIP uses separate domains to bind and to enhance activation of CILK1, enabling CILK1 function in control of cilia formation and elongation.