FLS2 is a CDK-like kinase that directly binds IFT70 and is required for proper ciliary disassembly in Chlamydomonas

Abstract

Intraflagellar transport (IFT) is required for ciliary assembly and maintenance. While disruption of IFT may trigger ciliary disassembly, we show here that IFT mediated transport of a CDK-like kinase ensures proper ciliary disassembly. Mutations in flagellar shortening 2 (FLS2), encoding a CDK-like kinase, lead to retardation of cilia resorption and delay of cell cycle progression. Stimulation for ciliary disassembly induces gradual dephosphorylation of FLS2 accompanied with gradual inactivation. Loss of FLS2 or its kinase activity induces early onset of kinesin13 phosphorylation in cilia. FLS2 is predominantly localized in the cell body, however, it is transported to cilia upon induction of ciliary disassembly. FLS2 directly interacts with IFT70 and loss of this interaction inhibits its ciliary transport, leading to dysregulation of kinesin13 phosphorylation and retardation of ciliary disassembly. Thus, this work demonstrates that IFT plays active roles in controlling proper ciliary disassembly by transporting a protein kinase to cilia to regulate a microtubule depolymerizer.

Identification of Regulators for Ciliary Disassembly by a Chemical Screen

“…Ciliary disassembly in Chlamydomonas can be induced experimentally by treatment of the cells with 20 mM NaPPi, a calcium chelator. , Under this condition, cilia are gradually disassembled around 3 h and cells become immotile. Thus, cilia motility can be used as a readout for ciliary disassembly.…”

“…NaPPi induced ciliary disassembly involves two CDK-like kinases, FLS1 and FLS2. , We then tested whether abemaciclib inhibited FLS1, which undergoes phosphorylation with a gel shift during NaPPi induced ciliary disassembly. As shown in Figure e, abemaciclib indeed inhibited FLS1 phosphorylation induced during ciliary disassembly.…”

“…Thus, the effect of abemaciclib on ciliary disassembly is likely mediated at least by inhibiting FLS1; FLS2 and other unknown CDK-related kinases that are involved in ciliary disassembly may also be inhibited. The kinase activities of FLS1 and FLS2 are not involved in ciliary assembly and maintenance. , Thus, they are unlikely the targets of abemaciclib in promoting ciliary length in the steady state cells. It is curious how abemaciclib affects ciliary assembly in steady state cells.…”

“…Chemical screens in this organism have revealed regulators involved in cilia motility, ciliary assembly, and deciliation. , The ciliary disassembly of Chlamydomonas can be induced experimentally by treatment with sodium pyrophosphate (NaPPi) . Previously, we have used genetic screens in this system and uncovered two CDK-like kinases that are involved in ciliary disassembly, , demonstrating that such a system can be used for chemical screen to identify regulators in ciliary disassembly.…”

Phosphorylation and Ubiquitylation Regulate Protein Trafficking, Signaling, and the Biogenesis of Primary Cilia

“…In support of a central role for KIF24 in cilium disassembly, a recent study identified FLS2 as a CDK-like kinase that phosphorylates the KIF24 ortholog CrKIF13 in Chlamydomonas, allowing efficient cilia disassembly (Figure 2D; Zhao et al, 2020). In turn, phosphorylated FLS2 showed lower activity and appears to be dephosphorylated upon cilia disassembly when it enters cilia by binding to the IFT-B component IFT70 (Zhao et al, 2020).…”