TET3 epigenetically controls feeding and stress response behaviors via AGRP neurons
Di Xie, … , Tamas L. Horvath, Yingqun Huang
Di Xie, … , Tamas L. Horvath, Yingqun Huang
Published October 3, 2022
Citation Information: J Clin Invest. 2022;132(19):e162365. https://doi.org/10.1172/JCI162365.
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Research Article Metabolism Neuroscience

TET3 epigenetically controls feeding and stress response behaviors via AGRP neurons

Abstract

The TET family of dioxygenases promote DNA demethylation by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Hypothalamic agouti-related peptide–expressing (AGRP-expressing) neurons play an essential role in driving feeding, while also modulating nonfeeding behaviors. Besides AGRP, these neurons produce neuropeptide Y (NPY) and the neurotransmitter GABA, which act in concert to stimulate food intake and decrease energy expenditure. Notably, AGRP, NPY, and GABA can also elicit anxiolytic effects. Here, we report that in adult mouse AGRP neurons, CRISPR-mediated genetic ablation of Tet3, not previously known to be involved in central control of appetite and metabolism, induced hyperphagia, obesity, and diabetes, in addition to a reduction of stress-like behaviors. TET3 deficiency activated AGRP neurons, simultaneously upregulated the expression of Agrp, Npy, and the vesicular GABA transporter Slc32a1, and impeded leptin signaling. In particular, we uncovered a dynamic association of TET3 with the Agrp promoter in response to leptin signaling, which induced 5hmC modification that was associated with a chromatin-modifying complex leading to transcription inhibition, and this regulation occurred in both the mouse models and human cells. Our results unmasked TET3 as a critical central regulator of appetite and energy metabolism and revealed its unexpected dual role in the control of feeding and other complex behaviors through AGRP neurons.

Authors

Di Xie, Bernardo Stutz, Feng Li, Fan Chen, Haining Lv, Matija Sestan-Pesa, Jonatas Catarino, Jianlei Gu, Hongyu Zhao, Christopher E. Stoddard, Gordon G. Carmichael, Marya Shanabrough, Hugh S. Taylor, Zhong-Wu Liu, Xiao-Bing Gao, Tamas L. Horvath, Yingqun Huang

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Figure 2

TET3 knockdown increases AGRP expression and AGRP neuronal activity.

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TET3 knockdown increases AGRP expression and AGRP neuronal activity. (A)...
(A) Schematic diagram of AAV-sgTet3 (top) with the sgRNA design for targeting the mouse Tet3 genomic locus (bottom). (B) Schematic diagram of bilateral virus injection into the ARC of Cas9+ mice. (C) Representative photomicrographs of AGRP neurons (green) expressing injected AAV-sgTet3 (red). (D) Representative photomicrographs and corresponding statistical analysis of TET3+ (green) AGRP neurons (red) showing decreased TET3 expression in AGRP neurons in AAV-sgTet3–injected mice. n = 5 mice per group. ***P < 0.001, by 2-tailed Student’s t test. (E) Representative photomicrographs of AGRP neurons (red) and AGRP (green) showing a marked increase in AGRP in the ARC of ad libitum–fed mice injected with AAV-sgTet3. (F) Increased expression of Agrp mRNA in the ARC of mice injected with AAV-sgTet3. n = 6–7 mice per group. **P < 0.01, by 2-tailed Student’s t test. (G) Quantification of AGRP neurons in the ARCs of Cas9+ mice injected with AAV or AAV-sgTet3 showing no significant difference between the groups. n = 5 mice per group. Significance was determined by 2-tailed Student’s t test. (H) Representative traces of membrane and APs recorded under current clamp in AGRP neurons of Cas9+ mice injected with AAV or AAV-sgTet3. (I) Bar graphs show the frequency of spontaneous APs (AP freq, left) and AP threshold (right) in AGRP cells in control and TET3-knockdown mice. n = 10–11 neurons from 3–4 mice per group. *P < 0.05, by 2-tailed Student’s t test. All data represent the mean ± SEM. Scale bars: 50 μm (C–E).