title: Phosphorylation of ERK5 on Thr732 is associated with ERK5 nuclear localization and ERK5-dependent transcription.
abstract: Extracellular signal-regulated kinases (ERKs) play critical roles in numerous cellular processes, including proliferation and differentiation. ERK5 contains a kinase domain at the N-terminal, and the unique extended C-terminal includes multiple autophosphorylation sites that enhance ERK5-dependent transcription. However, the impact of phosphorylation at the various sites remain unclear. In this study, we examined the role of phosphorylation at the ERK5 C-terminal. We found that a constitutively active MEK5 mutant phosphorylated ERK5 at the TEY motif, resulting in the sequential autophosphorylation of multiple C-terminal residues, including Thr732 and Ser769/773/775. However, when ERK1/2 was selectively activated by an oncogenic RAS mutant, ERK5 phosphorylation at Thr732 was induced without affecting the phosphorylation status at TEY or Ser769/773/775. The Thr732 phosphorylation was U0126-sensitive and was observed in a kinase-dead mutant of ERK5 as well, suggesting that ERK1/2 can phosphorylate ERK5 at Thr732. This phosphorylation was also promoted by epidermal growth factor and nerve growth factor in HEK293 and PC12 cells, respectively. The ERK5-T732A mutant was localized in the cytosol under basal conditions. In contrast, ERK5 phosphorylated at Thr732 via the RAS-ERK1/2 pathway and ERK5-T732E, which mimics the phosphorylated form, were localized in both the nucleus and cytosol. Finally, ER-32A and U0126 blocked ERK5-dependent MEF2C transcriptional activity. Based on these findings, we propose a novel cross-talk mechanism in which ERK1/2, following activation by growth factor stimulation, phosphorylates ERK5 at Thr732. This phosphorylation event is responsible for ERK5 nuclear localization and ERK5-dependent transcription.
authors.affiliation: Department of Cellular Signaling, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba, Aramaki, Aoba-ku, Sendai 980-8578, Japan.
authors.forename: Takuto
authors.initials: T
authors.lastname: Honda
authors.affiliation: Department of Cellular Signaling, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba, Aramaki, Aoba-ku, Sendai 980-8578, Japan; Department of Pharmacology, Yamagata University School of Medicine, 2-2-2 Iida-Nishi, Yamagata, 990-9585, Japan.
authors.forename: Yutaro
authors.initials: Y
authors.lastname: Obara
authors.affiliation: Department of Cellular Signaling, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba, Aramaki, Aoba-ku, Sendai 980-8578, Japan.
authors.forename: Arata
authors.initials: A
authors.lastname: Yamauchi
authors.affiliation: Cell Signaling Technology, 3 Trask Lane, Danvers, MA 01923, United States of America.
authors.forename: Anthony D
authors.initials: AD
authors.lastname: Couvillon
authors.affiliation: Cell Signaling Technology, 3 Trask Lane, Danvers, MA 01923, United States of America.
authors.forename: Justin J
authors.initials: JJ
authors.lastname: Mason
authors.affiliation: Department of Pharmacology, Yamagata University School of Medicine, 2-2-2 Iida-Nishi, Yamagata, 990-9585, Japan.
authors.forename: Kuniaki
authors.initials: K
authors.lastname: Ishii
authors.affiliation: Department of Cellular Signaling, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba, Aramaki, Aoba-ku, Sendai 980-8578, Japan.
authors.forename: Norimichi
authors.initials: N
authors.lastname: Nakahata
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D048054:Mitogen-Activated Protein Kinase 7 / Q000737:chemistry* / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D051381:Rats; D013912:Threonine / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D013912:Threonine; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048054:Mitogen-Activated Protein Kinase 7