Identification and validation of the phosphorylation sites on Aristaless-related homeobox protein

Biosci Rep. 2020 Jul 31;40(7):BSR20194513. doi: 10.1042/BSR20194513.

Abstract

The Aristaless-related homeobox protein (ARX) is a transcription factor expressed in the developing forebrain, skeletal muscle, pancreas, testis, and a variety of other tissues. It is known to have context-dependent transcriptional activator and repressor activity, although how it can achieve these opposing functions remains poorly understood. We hypothesized phosphorylation status might play a role in pivoting ARX between functioning as an activator or repressor. To gain further mechanistic insight as to how ARX functions, we identified multiple phosphorylation sites on ARX. We further established PKA as the kinase that phosphorylates ARX at least at Ser266 in mice. Two other kinases, CK2α and CDK4/cyclin D1, were also identified as kinases that phosphorylate ARX in vitro. Unexpectedly, phosphorylation status did not change either the nuclear localization or transcriptional function of ARX.

Keywords: Aristaless-related homeobox; Cortex development; Mass Spectrometry; phosphorylation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Casein Kinase II / metabolism
  • Cell Nucleus / metabolism
  • Cerebral Cortex / embryology
  • Cerebral Cortex / metabolism
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • Cyclin D1 / metabolism
  • Cyclin-Dependent Kinase 4 / metabolism
  • Embryo, Mammalian
  • Female
  • Gene Expression Regulation, Developmental
  • HEK293 Cells
  • HeLa Cells
  • Homeodomain Proteins / genetics
  • Homeodomain Proteins / metabolism*
  • Humans
  • Mice
  • Mice, Transgenic
  • Phosphorylation / physiology
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Serine / metabolism
  • Sf9 Cells
  • Spodoptera
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*

Substances

  • ARX protein, mouse
  • Homeodomain Proteins
  • Recombinant Proteins
  • Transcription Factors
  • Cyclin D1
  • Serine
  • Casein Kinase II
  • Cyclic AMP-Dependent Protein Kinases
  • Cyclin-Dependent Kinase 4