A novel regulatory role of RGS4 in STAT5B activation, neurite outgrowth and neuronal differentiation

Neuropharmacology. 2017 May 1:117:408-421. doi: 10.1016/j.neuropharm.2017.02.012. Epub 2017 Feb 17.

Abstract

The Regulator of G protein Signalling 4 (RGS4) is a multitask protein that interacts with and negatively modulates opioid receptor signalling. Previously, we showed that the δ-opioid receptor (δ-OR) forms a multiprotein signalling complex consisting of Gi/Go proteins and the Signal Transducer and Activator of Transcription 5B (STAT5B) that leads to neuronal differentiation and neurite outgrowth upon δ-ΟR activation. Here, we investigated whether RGS4 could participate in signalling pathways to regulate neurotropic events. We demonstrate that RGS4 interacts directly with STAT5B independently of δ-ΟR presence both in vitro and in living cells. This interaction involves the N-terminal portion of RGS4 and the DNA-binding SH3 domain of STAT5B. Expression of RGS4 in HEK293 cells expressing δ-OR and/or erythropoietin receptor results in inhibition of [D-Ser2, Leu5, Thr6]-enkephalin (DSLET)-and erythropoietin-dependent STAT5B phosphorylation and subsequent transcriptional activation. DSLET-dependent neurite outgrowth of neuroblastoma cells is also blocked by RGS4 expression, whereas primary cortical cultures of RGS4 knockout mice (RGS4-/-) exhibit enhanced neuronal sprouting after δ-OR activation. Additional studies in adult brain extracts from RGS4-/- mice revealed increased levels of p-STAT5B. Finally, neuronal progenitor cultures from RGS4-/- mice exhibit enhanced proliferation with concomitant increases in the mRNA levels of the anti-apoptotic STAT5B target genes bcl2 and bcl-xl. These observations suggest that RGS4 is implicated in opioid dependent neuronal differentiation and neurite outgrowth via a "non-canonical" signaling pathway regulating STAT5B-directed responses.

Keywords: Deoxycholate (PubChem CID: 91896239); Differentiation; Erythropoietin; MgCl2(PubChem CID:5360315); Neurite outgrowth; Opioid receptor; PMSF (PubChem CID: 4784); RGS4; Retinoic acid (PubChem CID: 444795); STAT5B; Sodium orthovanadate (PubChem CID: 6167); Triton X-100 (PubChem CID: 5590); [(3)H]-diprenorphine (PubChem CID: 71719127); [D-Ser(2),Leu5, Thr6]-enkephalin (PubChem CID: 107847).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line, Tumor
  • Cell Survival / physiology
  • Cerebral Cortex / metabolism
  • HEK293 Cells
  • Humans
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Neural Stem Cells / metabolism
  • Neurogenesis / physiology*
  • Neuronal Outgrowth / physiology*
  • Neurons / metabolism*
  • Phosphorylation / physiology
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • RGS Proteins / genetics
  • RGS Proteins / metabolism*
  • RNA, Messenger / metabolism
  • Rats
  • Receptors, Erythropoietin / metabolism
  • Receptors, Opioid, delta / metabolism
  • STAT5 Transcription Factor / metabolism*
  • bcl-X Protein / metabolism

Substances

  • Bcl2l1 protein, mouse
  • Proto-Oncogene Proteins c-bcl-2
  • RGS Proteins
  • RNA, Messenger
  • Receptors, Erythropoietin
  • Receptors, Opioid, delta
  • STAT5 Transcription Factor
  • bcl-X Protein
  • Bcl2 protein, mouse
  • RGS4 protein