VRK1 regulates Cajal body dynamics and protects coilin from proteasomal degradation in cell cycle.

Lara Cantarero,Paul Renbaum,Hadar Vinograd-Byk,Marta Sanz-García,Ephrat Levy-Lahad,Pedro A Lazo

doi:10.1038/srep10543

Abstract

Cajal bodies (CBs) are nuclear organelles associated with ribonucleoprotein functions and RNA maturation. CBs are assembled on coilin, its main scaffold protein, in a cell cycle dependent manner. The Ser-Thr VRK1 (vaccinia-related kinase 1) kinase, whose activity is also cell cycle regulated, interacts with and phosphorylates coilin regulating assembly of CBs. Coilin phosphorylation is not necessary for its interaction with VRK1, but it occurs in mitosis and regulates coilin stability. Knockdown of VRK1 or VRK1 inactivation by serum deprivation causes a loss of coilin phosphorylation in Ser184 and of CBs formation, which are rescued with an active VRK1, but not by kinase-dead VRK1. The phosphorylation of coilin in Ser184 occurs during mitosis before assembly of CBs. Loss of coilin phosphorylation results in disintegration of CBs, and of coilin degradation that is prevented by proteasome inhibitors. After depletion of VRK1, coilin is ubiquitinated in nuclei, which is partly mediated by mdm2, but its proteasomal degradation occurs in cytosol and is prevented by blocking its nuclear export. We conclude that VRK1 is a novel regulator of CBs dynamics and stability in cell cycle by protecting coilin from ubiquitination and degradation in the proteasome, and propose a model of CB dynamics.

Highlights

CBs20, but how this aggregation is regulated in proliferating cells is not known
Cajal bodies (CBs) were isolated from nuclei and the presence of coilin and VRK1 was determined by immunofluorescence and immunoblot (Fig. 1A)
Purified CBs were detected by their positivity for coilin (Fig. 1A, detail and immunoblot), but VRK1 was not detected within CBs, if VRK1 is present its level must represent a very minor subpopulation

Summary

Introduction

CBs20, but how this aggregation is regulated in proliferating cells is not known. In addition, CBs contain multiple proteins whose function is not well known[21,22]. CBs dynamic assembly and disassembly requires regulatory mechanisms that are not yet identified, but in which protein phosphorylation is very likely to play an important role. In this work we have studied the role that VRK1 plays in the regulation of CBs. We have identified that VRK1 regulates CBs assembly in cell cycle by a specific phosphorylation and protects coilin from ubiquitin mediated degradation in the proteasome, in which the ubiquitin ligase Mdm[2] is implicated. We have identified that VRK1 regulates CBs assembly in cell cycle by a specific phosphorylation and protects coilin from ubiquitin mediated degradation in the proteasome, in which the ubiquitin ligase Mdm[2] is implicated This effect of VRK1 on coilin stability regulates the assembly and disassembly of Cajal bodies in nuclei as well as its variation in cell cycle progression

Methods

Results

Conclusion