Ankyrin repeat and SOCS box containing protein 4 (Asb-4) colocalizes with insulin receptor substrate 4 (IRS4) in the hypothalamic neurons and mediates IRS4 degradation

BMC Neurosci. 2011 Sep 28:12:95. doi: 10.1186/1471-2202-12-95.

Abstract

Background: The arcuate nucleus of the hypothalamus regulates food intake. Ankyrin repeat and SOCS box containing protein 4 (Asb-4) is expressed in neuropeptide Y and proopiomelanocortin (POMC) neurons in the arcuate nucleus, target neurons in the regulation of food intake and metabolism by insulin and leptin. However, the target protein(s) of Asb-4 in these neurons remains unknown. Insulin receptor substrate 4 (IRS4) is an adaptor molecule involved in the signal transduction by both insulin and leptin. In the present study we examined the colocalization and interaction of Asb-4 with IRS4 and the involvement of Asb-4 in insulin signaling.

Results: In situ hybridization showed that the expression pattern of Asb-4 was consistent with that of IRS4 in the rat brain. Double in situ hybridization showed that IRS4 colocalized with Asb-4, and both Asb-4 and IRS4 mRNA were expressed in proopiomelanocortin (POMC) and neuropeptide Y (NPY) neurons within the arcuate nucleus of the hypothalamus. In HEK293 cells co-transfected with Myc-tagged Asb-4 and Flag-tagged IRS4, Asb-4 co-immunoprecipitated with IRS4; In these cells endogenous IRS4 also co-immunoprecipitated with transfected Myc-Asb-4; Furthermore, Asb-4 co-immunoprecipitated with IRS4 in rat hypothalamic extracts. In HEK293 cells over expression of Asb-4 decreased IRS4 protein levels and deletion of the SOCS box abolished this effect. Asb-4 increased the ubiquitination of IRS4; Deletion of SOCS box abolished this effect. Expression of Asb-4 decreased both basal and insulin-stimulated phosphorylation of AKT at Thr308.

Conclusions: These data demonstrated that Asb-4 co-localizes and interacts with IRS4 in hypothalamic neurons. The interaction of Asb-4 with IRS4 in cell lines mediates the degradation of IRS4 and decreases insulin signaling.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • CHO Cells
  • Cell Line
  • Cricetinae
  • HEK293 Cells
  • Humans
  • Hypothalamus / cytology*
  • Hypothalamus / metabolism*
  • Insulin / metabolism
  • Insulin / physiology
  • Insulin Receptor Substrate Proteins / antagonists & inhibitors*
  • Insulin Receptor Substrate Proteins / genetics
  • Insulin Receptor Substrate Proteins / metabolism*
  • Male
  • Mice
  • Neurons / cytology
  • Neurons / metabolism*
  • Rats
  • Rats, Sprague-Dawley
  • Signal Transduction / physiology
  • Suppressor of Cytokine Signaling Proteins / metabolism*
  • Suppressor of Cytokine Signaling Proteins / physiology

Substances

  • Asb4 protein, mouse
  • Asb4 protein, rat
  • IRS4 protein, rat
  • Insulin
  • Insulin Receptor Substrate Proteins
  • Suppressor of Cytokine Signaling Proteins