The U-box-type ubiquitin ligase PRP19β regulates astrocyte differentiation via ubiquitination of PTP1B

Brain Res. 2013 Aug 2;1524:12-25. doi: 10.1016/j.brainres.2013.06.007. Epub 2013 Jun 14.

Abstract

U-box protein PRP19β, a splicing variant of PRP19α, suppresses neuronal differentiation and conversely promotes astrocyte differentiation as a neuron/glia switch molecule. However, the mechanistic basis of PRP19β in astrocyte differentiation is not well understood. Here, we demonstrated that PRP19β regulates the stability of protein tyrosine phosphatase 1B (PTP1B) via ubiquitination during N(6),2'-O-dibutyryl cyclic AMP (cAMP)-induced astrocyte differentiation of C6 cells. Only overexpression of PRP19β conferred astrocyte properties at a certain level, and induced more astrocyte markers, glial fibrillary acidic protein (GFAP) and S100β, in the presence of cAMP, whereas its down-regulation by antisense RNA showed a suppressive effect. In addition, ectopic expression of PRP19β led to robust phosphorylation of signal transducer and activator of transcription 3 (STAT3) accompanying the reduction in PTP1B stability during astrocyte differentiation. Immunological analysis revealed that PRP19β interacted with PTP1B and ubiquitinated PTP1B via its U-box region. Forced expression of the U-box deletion mutant of PRP19β resulted in inhibition of astrocyte differentiation. Moreover, down-regulation of PTP1B by short hairpin (sh)RNA enhanced astrocyte differentiation, while forced expression of PTP1B showed an inhibitory effect. Thus, these results indicate that PRP19β activates the gp130/Janus kinase (JAK)/STAT signaling pathway during astrocyte differentiation of C6 cells via PTP1B ubiquitination.

Keywords: Astrocyte differentiation; CHX; GAPDH; GFAP; IL-6; JAK; Janus kinase; N(6),2-O-dibutyryl cyclic AMP; Neuron/glia switch; PRP19β; PTP1B; RA; SHP2; STAT3; Src homology domain 2 (SH2)-containing protein phosphatase; Ub; Ubiquitination; WB; Western blotting; cAMP; cycloheximide; glial fibrillary acidic protein; glyceraldehyde 3′-phosphate dehydrogenase; interleukin-6; pSTAT3; phosphorylated STAT3; protein tyrosine phosphatase 1B; retinoic acid; signal transducer and activator of transcription 3; ubiquitin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Astrocytes / cytology*
  • Astrocytes / metabolism*
  • Blotting, Northern
  • Blotting, Western
  • Cell Differentiation / physiology*
  • Cell Line
  • Immunohistochemistry
  • Immunoprecipitation
  • Protein Tyrosine Phosphatase, Non-Receptor Type 1 / metabolism*
  • Rats
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction / physiology*
  • Ubiquitin-Protein Ligases / metabolism*
  • Ubiquitination*

Substances

  • Prpf19 protein, rat
  • Ubiquitin-Protein Ligases
  • Protein Tyrosine Phosphatase, Non-Receptor Type 1
  • Ptpn1 protein, rat