RRC ID 61143
Author Kinoshita-Kikuta E, Utsumi T, Miyazaki A, Tokumoto C, Doi K, Harada H, Kinoshita E, Koike T.
Title Protein-N-myristoylation-dependent phosphorylation of serine 13 of tyrosine kinase Lyn by casein kinase 1γ at the Golgi during intracellular protein traffic.
Journal Sci Rep
Abstract Protein N-myristoylation of Src-family kinases (SFKs) is a critical co-translational modification to anchor the enzymes in the plasma membrane. Phosphorylation of SFKs is also an essential modification for regulating their enzymatic activities. In this study, we used Phos-tag SDS-PAGE to investigate N-myristoylation-dependent phosphorylation of SFKs and their non-N-myristoylated G2A mutants. The serine-13 residue of Lyn (Lyn-S13) was shown to be N-myristoylation-dependently phosphorylated. Although there have been more than 40 reports of mass spectrometric studies on phosphorylation at Lyn-S13, the kinase responsible remained unclear. We succeeded in identifying casein kinase 1γ (CK1γ) as the kinase responsible for phosphorylation of Lyn-S13. In HEK293 cells co-expressing Lyn and CK1γ, the phosphorylation level of Lyn-S13 increased significantly. CK1γ is unique among the CK1 family (α, γ, δ, and ε) in carrying an S-palmitoylation site for membrane binding. Co-expression with the non-S-palmitoylated CK1γ mutant, which localized in the cytosol, gave no increase in the phosphorylation level at Lyn-S13. In HEK293 cells expressing the non-S-palmitoylated Lyn-C3A mutant, on the other hand, the Lyn-C3A mutant was phosphorylated at Lyn-S13, and the mutant remained at the Golgi. These results showed that S-palmitoylated CK1γ can phosphorylate S13 of N-myristoylated Lyn at the Golgi during intracellular protein traffic.
Volume 10(1)
Pages 16273
Published 2020-10-1
DOI 10.1038/s41598-020-73248-0
PII 10.1038/s41598-020-73248-0
PMID 33004926
PMC PMC7531007
MeSH Casein Kinase I / metabolism* Cell-Free System Electrophoresis, Polyacrylamide Gel Golgi Apparatus / metabolism* HEK293 Cells / metabolism Humans Mass Spectrometry Microscopy, Fluorescence Phosphorylation Protein Transport* Protein-Tyrosine Kinases / metabolism* Serine
IF 3.998
Resource
DNA material Lyn/pLY30 (RDB01306) Genome Network Project Human cDNA clone IRAL021O14 (HGY088750) IRAK064L13 (HGX025877) IRAL034D12 (HGY093684) IRAL047C19 (HGY098867) W01A107I15 (HGE043007) W01A056G21 (HGE022565) W01A026C12 (HGE010460) pCDNA3.1_SRC_FLAG (RDB19292) pCDNA3.1_SRC_G2A_FLAG (RDB19293) pCDNA3.1_LCK_FLAG (RDB19294) pCDNA3.1_LCK_G2A_FLAG (RDB19295) pCDNA3.1_HCK_FLAG (RDB19296) pCDNA3.1_HCK_G2A_FLAG (RDB19297) pCDNA3.1_BLK_FLAG (RDB19298) pCDNA3.1_BLK_G2A_FLAG (RDB19299) pF25AICET7_LYN (RDB19300) pF25AICET7_LYN_G2A (RDB19301) pF25AICET7_LYN_S13A (RDB19302) pFN21A_Halo_CK1alpha (RDB19303) pFN21A_Halo_CK1alpha_K46R (RDB19304) pFN21A_Halo_CK1gamma1 (RDB19305) pFN21A_Halo_CK1gamma1_K74R (RDB19306) pFN21A_Halo_CK1gamma1_deltaC (RDB19307) pFN21A_Halo_CK1gamma2 (RDB19308) pFN21A_Halo_CK1gamma2_K75R (RDB19309) pFN21A_Halo_CK1gamma2_deltaC (RDB19310) pFN21A_Halo_CK1gamma3 (RDB19311) pFN21A_Halo_CK1gamma3_K72R (RDB19312) pFN21A_Halo_CK1gamma3_deltaC (RDB19313) pFN21A_Halo_CK1delta (RDB19314) pFN21A_Halo_CK1delta_K38R (RDB19315) pFN21A_Halo_CK1epsilon (RDB19316) pFN21A_Halo_CK1epsilon_K38R (RDB19317)