Role of Vaccinia-related kinase 2 in the regulation of dysbindin and G3BP1

DoctorVaccinia-related kinase 2 (VRK2) is a serine/threonine kinase highly ex-pressed in proliferating cells. In genome-wide screens, VRK2 has been found to be associated with neurological disorders such as schizophrenia and epilepsy. Especially, In gene expression analysis of patients with schizophrenia, a single-nucleotide polymorphism (SNP) located upstream of VRK2 caused an in-creased risk of schizophrenia. However, few VRK2 substrates are known, and its functionality has yet to be fully explored. It is reported that VRK2 directly interacts with and phosphorylates NFAT1 to induce gene expression of COX-2, which promotes cancer cell invasion. Also, VRK2 phosphorylates and inhibits ubiquitin-specific protease 25 (USP25), a TRiC deubiquitinating enzyme. As a result, it induces recruitment of the E3 ligase COP1 and degrades chaperonin TRiC protein, which also results in increased polyQ aggregation. However, as mentioned above, only a few substrates of VRK2 are known, and there are more functions of VRK2 left to be discovered. In the first chapter, I demonstrated that VRK2 modulates role of dys-bindin by regulating protein stability. Dysbindin is known as one of the strong risk factors for schizophrenia. The expression of dysbindin is indeed signifi-cantly reduced in schizophrenia patients. Moreover, dysbindin is involved in neurite outgrowth and regulation of NMDA receptor signaling. I first identified dysbindin as a novel interacting protein of VRK2 and showed that VRK2 phosphorylates Ser 297 and Ser 299 of dysbindin. In addition, I found that VRK2‐mediated phosphorylation of dysbindin enhanced ubiquitination of dysbindin and consequently resulted in the decrease in its protein stability. Over‐expression of VRK2 reduced neurite outgrowth. Furthermore, a phos-phomimetic mutant of dysbindin alleviated neurite outgrowth and affected sur-face expression of N‐methyl‐d‐aspartate 2A, a subunit of NMDA receptor in mouse hippocampal neurons. Thus, this findings reveals the regulation of dys-bindin by VRK2, providing the association of these two proteins, which are commonly implicated in schizophrenia. In the second chapter, I showed that VRK2 negatively regulates the for-mation of stress granules via phosphorylation of G3BP1. In mammalian cells, global translational repression in response to environmental stress occurs via assembly of cytoplasmic messenger ribonucleoprotein (mRNP) into stress granules (SGs). RasGAP SH3 domain binding protein 1 (G3BP1) is one of the factors involved in stress granule assembly and is known to induce the for-mation of SGs through oligomerization. Previous reports showed that phos-phorylation of G3BP1 at serine 149 (S149) interferes with G3BP1 oligomeriza-tion. However, the identity of all kinases involved in phosphorylation of G3BP1 is still unclear. I identified that G3BP1 as a protein that directly interacts with VRK2. Furthermore, VRK2 phosphorylates G3BP1 at S149, which attenu-ates the aggregation of SGs. Moreover, the phosphorylated forms of G3BP1 and VRK2 levels are consistently decreased under cell stress. These findings suggest a novel function of VRK2 in a cellular defensive response under stress by regulating SG formation as an upstream kinase of G3BP1