Detection of Leptin Activity in Living Cells Expressing Chicken Leptin Receptor and STAT3

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  • Adachi Hiromi
    College of Agriculture, Ibaraki University, Japan United Graduate School of Agriculture, Tokyo University of Agriculture and Technology, Japan
  • Murase Daisuke
    College of Agriculture, Ibaraki University, Japan United Graduate School of Agriculture, Tokyo University of Agriculture and Technology, Japan
  • Atomura Susumu
    College of Agriculture, Ibaraki University, Japan
  • Ohkubo Takeshi
    College of Agriculture, Ibaraki University, Japan

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Abstract

The chicken leptin receptor (chLEPR) activates Janus kinase (JAK) - signal transducers and activator of transcription (STAT) signaling pathway after leptin stimulation. We have previously developed a bioassay using leptin inducible reporter gene in cultured cells. However, we failed to detect leptin-like activity in chicken blood. In the present study, we expressed green fluorescent protein (GFP) fused chSTAT3 (GFP-chSTAT3) in cells expressing chLEPR and analyzed leptin dependent activation of the chSTAT3. Leptin phosphorylated GFP-chSTAT3 and by which fluorescent signal translocated into nuclei in COS-7 cells transiently expressing GFP-chSTAT3 with chLEPR. Furthermore, we established CHO-K1 cells stably expressing chLEPR and chSTAT3 (CHO-chLEPR/chSTAT3), and in which detected time- and dose-dependent activation of chSTAT3 by leptin. Therefore, the CHO-chLEPR/STAT3 cells would be an excellent tool to detect and monitor leptin-like activity in avian tissues.

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