Identifying novel regulators of ciliogenesis

Primary cilia are microtubule-based “antennae-like” organelles extending from the apical surface of most mammalian cells. They integrate mechanical or chemical signals essential for cell homeostasis and differentiation through several cooperating compartments. Mutations in genes that encode ciliary proteins or proteins essential for correct ciliary function are the cause of a major group of inherited and variable developmental disorders known as ciliopathies. Functional interactions between ciliary compartments, the molecular basis of variable clinical phenotypes, and the mechanisms of cilia formation are all still poorly understood. The presence or absence of cilia can be easily imaged and quantitated. This lends itself to high-throughput, high-content imaging in reverse genetic screens of cellular phenotypes. siRNA reverse genetic screens were performed to assess increased cilia incidence and identified a key negative regulator of ciliogenesis. Analysis showed this regulator acts through a mechanism involving actin remodelling and acto-myosin contraction. Pharmacological inhibition of this target may therefore comprise a novel therapeutic approach for treatment of a broad group of ciliopathy disease classes. Further screening of a data-set for supernumerary primary cilia, added to the evidence that cytokinesis is not a prerequisite for ciliogenesis. Disruption of the centralspindilin complex caused mitotic failure and maturation of supernumerary centrosomes, leading to the formation of the supernumerary cilia, a known cellular phenotype of severe ciliopathies. Finally, a combinatorial screening approach to generate double genetic perturbations of ciliary genes identified reciprocal synthetic genetic interactions between anterograde intraflagellar transport (the IFT B complex) and the transition zone. Reverse genetics screening techniques have identified novel regulators and pathways of ciliogenesis, and a potential therapeutic target for ciliopathies. Furthermore, combinatorial screening has highlighted a novel and complex interaction in ciliary biology, that may provide potential new insights into ciliary organisation and disease pathomechanisms.