CCRK/CDK20 regulates ciliary retrograde protein trafficking via interacting with BROMI/TBC1D32.

Tatsuro Noguchi,Kentaro Nakamura,Yohei Katoh,Kazuhisa Nakayama,Yuuki Satoda,Oliver E Blacque

doi:10.1371/journal.pone.0258497

Abstract

CCRK/CDK20 was reported to interact with BROMI/TBC1D32 and regulate ciliary Hedgehog signaling. In various organisms, mutations in the orthologs of CCRK and those of the kinase ICK/CILK1, which is phosphorylated by CCRK, are known to result in cilia elongation. Furthermore, we recently showed that ICK regulates retrograde ciliary protein trafficking and/or the turnaround event at the ciliary tips, and that its mutations result in the elimination of intraflagellar transport (IFT) proteins that have overaccumulated at the bulged ciliary tips as extracellular vesicles, in addition to cilia elongation. However, how these proteins cooperate to regulate ciliary protein trafficking has remained unclear. We here show that the phenotypes of CCRK-knockout (KO) cells closely resemble those of ICK-KO cells; namely, the overaccumulation of IFT proteins at the bulged ciliary tips, which appear to be eliminated as extracellular vesicles, and the enrichment of GPR161 and Smoothened on the ciliary membrane. The abnormal phenotypes of CCRK-KO cells were rescued by the exogenous expression of wild-type CCRK but not its kinase-dead mutant or a mutant defective in BROMI binding. These results together indicate that CCRK regulates the turnaround process at the ciliary tips in concert with BROMI and probably via activating ICK.

Highlights

The crucial roles of primary cilia in embryonic development and adult tissue homeostasis are widely acknowledged by the fact that ciliary dysfunctions cause genetically heterogeneous disorders with a wide spectrum of clinical manifestations, collectively referred to as the ciliopathies [1,2]
We recently showed that is the average ciliary length longer and the variation in ciliary length is larger in intestinal cell kinase (ICK)-KO cells than in control cells, and that excessively accumulated proteins at the bulged ciliary tips of ICK-KO cells are eliminated as cell cycle-related kinase (CCRK)–BROMI interplay in ciliary protein trafficking extracellular vesicles (ECVs) [30]
In ICK-KO cells and IFT144-KO cells, as well as in cells with a compromised interaction between the intraflagellar transport (IFT)-A and IFT-B complexes, ciliary proteins that are excessively accumulated at the ciliary tips owing to an impairment in the turnaround process are packaged into ECVs and eliminated to relieve cilia of the stress of protein overaccumulation [30,71]

Summary

Introduction

The crucial roles of primary cilia in embryonic development and adult tissue homeostasis are widely acknowledged by the fact that ciliary dysfunctions cause genetically heterogeneous disorders with a wide spectrum of clinical manifestations, collectively referred to as the ciliopathies [1,2]. Cilia are axonemal microtubule-based protrusions surrounded by the ciliary membrane, which are found in a variety of eukaryotic cells. Via specific receptors and ion channels on the ciliary membrane, cilia function as cellular antennae for extracellular stimuli, such as fluid flow, and for extracellular signaling molecules, such as the Hedgehog (Hh) family of morphogens [3,4].

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