title: 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.
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.
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109, USA.
authors.forename: Ji-Yao
authors.initials: JY
authors.lastname: Li
authors.forename: Biaoxin
authors.initials: B
authors.lastname: Chai
authors.forename: Weizhen
authors.initials: W
authors.lastname: Zhang
authors.forename: Xiaobin
authors.initials: X
authors.lastname: Wu
authors.forename: Chao
authors.initials: C
authors.lastname: Zhang
authors.forename: Danielle
authors.initials: D
authors.lastname: Fritze
authors.forename: Zefeng
authors.initials: Z
authors.lastname: Xia
authors.forename: Cam
authors.initials: C
authors.lastname: Patterson
authors.forename: Michael W
authors.initials: MW
authors.lastname: Mulholland
mesh_terms: D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D057809:HEK293 Cells; D006801:Humans; D007031:Hypothalamus / Q000166:cytology* / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism / Q000502:physiology; D055504:Insulin Receptor Substrate Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D008297:Male; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology; D050826:Suppressor of Cytokine Signaling Proteins / Q000378:metabolism* / Q000502:physiology
chemical_list: C530435:Asb4 protein, mouse; C573786:Asb4 protein, rat; C491532:IRS4 protein, rat; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D050826:Suppressor of Cytokine Signaling Proteins