Leptin signaling regulates hypothalamic expression of nescient helix-loop-helix 2 (Nhlh2) through signal transducer and activator 3 (Stat3).
Mice with a deletion of the hypothalamic basic helix-loop-helix transcription factor Nhlh2 display adult onset obesity. We have previously shown that Nhlh2 expression is induced by leptin. In this study, we identify a small proximal leptin-responsive promoter region in the Nhlh2 gene. This 163bp promoter contains five putative binding sites for the leptin-activated Stat3 transcription factor, and two putative binding sites for the NFκB transcription factor. Results of mutagenesis studies reveal that deletion of the NFκB sites have little effect, mutagenesis of the third Stat3 site eliminates both leptin-induced and basal expression of Nhlh2. Mutagenesis of the 4th and 5th sites eliminates leptin-induced expression, and increases basal expression above the WT promoter. Stat3 can be preferentially pulled down from leptin-treated mouse hypothalamic chromatin extracts. This study identifies leptin-induced Stat3 transcription factor as the major transcriptional regulator of Nhlh2. As Nhlh2 transcriptionally regulates genes within the melanocortin pathway, these findings have implications for human body weight control.
Department of Human Nutrition, Foods and Exercise, 1981 Kraft Drive, ILSB Room 1020 (0913), Virginia Tech, Blacksburg, VA 24061, United States.
Numan
N
Al Rayyan
Department of Human Nutrition, Foods and Exercise, 1981 Kraft Drive, ILSB Room 1020 (0913), Virginia Tech, Blacksburg, VA 24061, United States.
Jinhua
J
Zhang
Department of Veterinary and Animal Sciences, Integrated Sciences Building, University of Massachusetts, 661 N. Pleasant Street, Amherst, MA 01003, United States.
Amy S
AS
Burnside
Department of Human Nutrition, Foods and Exercise, 1981 Kraft Drive, ILSB Room 1020 (0913), Virginia Tech, Blacksburg, VA 24061, United States. Electronic address: goodd@vt.edu.
Deborah J
DJ
Good
D000818:Animals; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D002460:Cell Line; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D007031:Hypothalamus / Q000166:cytology / Q000378:metabolism*; D020738:Leptin / Q000235:genetics* / Q000378:metabolism; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D050796:STAT3 Transcription Factor / Q000235:genetics* / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D015398:Signal Transduction; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D051792:Basic Helix-Loop-Helix Proteins; D020738:Leptin; D008156:Luciferases; D016328:NF-kappa B; D050796:STAT3 Transcription Factor; D001616:beta-Galactosidase; C484104:Nhlh2 protein, mouse; C494087:Stat3 protein, mouse
Chromatin immunoprecipitation assay; Electrophoretic mobility shift assay; Luciferase assay; Promoter; Transcription factor; Transfection