Abstract
Centrioles are the major constituents of the animal centrosome, in which Plk4 kinase serves as a master regulator of the duplication cycle. Many eukaryotes also contain numerous peripheral particles known as centriolar satellites. While centriolar satellites aid centriole assembly and primary cilium formation, it is unknown whether Plk4 plays any regulatory roles in centriolar satellite integrity. Here we show that Plk4 is a critical determinant of centriolar satellite organisation. Plk4 depletion leads to the dispersion of centriolar satellites and perturbed ciliogenesis. Plk4 interacts with the satellite component PCM1, and its kinase activity is required for phosphorylation of the conserved S372. The nonphosphorylatable PCM1 mutant recapitulates phenotypes of Plk4 depletion, while the phosphomimetic mutant partially rescues the dispersed centriolar satellite patterns and ciliogenesis in cells depleted of PCM1. We show that S372 phosphorylation occurs during the G1 phase of the cell cycle and is important for PCM1 dimerisation and interaction with other satellite components. Our findings reveal that Plk4 is required for centriolar satellite function, which may underlie the ciliogenesis defects caused by Plk4 dysfunction.
Highlights
The centrosome consists of two orthogonally arranged centrioles that are surrounded by the pericentriolar material, and plays a multifaceted role in a wide range of biological processes as a major microtubule-organising centre (MTOC) [1]
While Plk4-WT* effectively ameliorated the pericentrosomal localisation of PCM1, the introduction of Plk4-KD* was not able to rescue the dispersion of PCM1 (Figs 1H and I, and EV1J and K)
In order to address whether the centrosome/centriole has any impact on centriolar satellite integrity, we first examined the emergence of centriolar satellites under conditions where Plk4 was depleted
Summary
The centrosome consists of two orthogonally arranged centrioles that are surrounded by the pericentriolar material, and plays a multifaceted role in a wide range of biological processes as a major microtubule-organising centre (MTOC) [1]. We show that Plk is required for the spatial distribution and organisation of centriolar satellites separable from its role in centriole duplication This role is executed by Plk4-dependent phosphorylation of PCM1. We found that Plk knockdown in U2OS cells led to the dispersal of centriolar satellite foci away from the vicinity of the centrosome (detected by an anti-PCM1 antibody) (Fig 1A–C). In order to address whether satellite dispersion is induced independently of Plk4’s role in centriole duplication, U2OS cells were first arrested in G1 phase, followed by Plk siRNA treatment (Fig EV1G–I). U2OS cells were treated with control, Plk or hSAS-6 siRNA, and immunoblotting was performed with the indicated antibodies.
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