Identification of amphiphysin 1 as an endogenous substrate for CDKL5, a protein kinase associated with X-linked neurodevelopmental disorder

Mari Sekiguchi, Syouichi Katayama, Naoya Hatano, Yasushi Shigeri, Noriyuki Sueyoshi, Isamu Kameshita

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

Cyclin-dependent kinase-like 5 (CDKL5) is a Ser/Thr protein kinase predominantly expressed in brain and mutations of its gene are known to be associated with neurodevelopmental disorders such as X-linked West syndrome and Rett syndrome. However, the physiological substrates of CDKL5 that are directly linked to these neurodevelopmental disorders are currently unknown. In this study, we explored endogenous substrates for CDKL5 in mouse brain extracts fractionated by a liquid-phase isoelectric focusing. In conjunction with CDKL5 phosphorylation assay, this approach detected a protein band with an apparent molecular mass of 120 kDa that is remarkably phosphorylated by CDKL5. This 120-kDa protein was identified as amphiphysin 1 (Amph1) by LC-MS/MS analysis, and the site of phosphorylation by CDKL5 was determined to be Ser-293. The phosphorylation mimic mutants, Amph1(S293E) and Amph1(S293D), showed significantly reduced affinity for endophilin, a protein involved in synaptic vesicle endocytosis. Introduction of point mutations in the catalytic domain of CDKL5, which are disease-causing missense mutations found in Rett patients, resulted in the impairment of kinase activity toward Amph1. These results suggest that Amph1 is the cytoplasmic substrate for CDKL5 and that its phosphorylation may play crucial roles in the neuronal development.

Original languageEnglish
Pages (from-to)257-267
Number of pages11
JournalArchives of Biochemistry and Biophysics
Volume535
Issue number2
DOIs
Publication statusPublished - Jun 3 2013
Externally publishedYes

Keywords

  • Amphiphysin 1
  • CDKL5
  • Endogenous substrate
  • Endophilin
  • Neurodevelopmental disorder
  • Rett syndrome

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

Fingerprint Dive into the research topics of 'Identification of amphiphysin 1 as an endogenous substrate for CDKL5, a protein kinase associated with X-linked neurodevelopmental disorder'. Together they form a unique fingerprint.

  • Cite this