Hypothalamic ER–associated degradation regulates POMC maturation, feeding, and age-associated obesity
Geun Hyang Kim, … , Martin G. Myers Jr., Ling Qi
Geun Hyang Kim, … , Martin G. Myers Jr., Ling Qi
Published February 19, 2018
Citation Information: J Clin Invest. 2018;128(3):1125-1140. https://doi.org/10.1172/JCI96420.
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Research Article Cell biology Metabolism

Hypothalamic ER–associated degradation regulates POMC maturation, feeding, and age-associated obesity

Abstract

Pro-opiomelanocortin (POMC) neurons function as key regulators of metabolism and physiology by releasing prohormone-derived neuropeptides with distinct biological activities. However, our understanding of early events in prohormone maturation in the ER remains incomplete. Highlighting the significance of this gap in knowledge, a single POMC cysteine-to-phenylalanine mutation at position 28 (POMC-C28F) is defective for ER processing and causes early onset obesity in a dominant-negative manner in humans through an unclear mechanism. Here, we report a pathologically important role of Sel1L-Hrd1, the protein complex of ER-associated degradation (ERAD), within POMC neurons. Mice with POMC neuron–specific Sel1L deficiency developed age-associated obesity due, at least in part, to the ER retention of POMC that led to hyperphagia. The Sel1L-Hrd1 complex targets a fraction of nascent POMC molecules for ubiquitination and proteasomal degradation, preventing accumulation of misfolded and aggregated POMC, thereby ensuring that another fraction of POMC can undergo normal posttranslational processing and trafficking for secretion. Moreover, we found that the disease-associated POMC-C28F mutant evades ERAD and becomes aggregated due to the presence of a highly reactive unpaired cysteine thiol at position 50. Thus, this study not only identifies ERAD as an important mechanism regulating POMC maturation within the ER, but also provides insights into the pathogenesis of monogenic obesity associated with defective prohormone folding.

Authors

Geun Hyang Kim, Guojun Shi, Diane R.M. Somlo, Leena Haataja, Soobin Song, Qiaoming Long, Eduardo A. Nillni, Malcolm J. Low, Peter Arvan, Martin G. Myers Jr., Ling Qi

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Figure 6

POMC-C28F readily forms disulfide bond–mediated aggregates.

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POMC-C28F readily forms disulfide bond–mediated aggregates. (A) Schemati...
(A) Schematic diagram showing amino acid sequence of POMC 26–50 and the positions of 2 disulfide bonds and a free thiol in POMC-C28F. (B) Western blot analysis of steady-state levels of POMC proteins in WT and Hrd1–/– N2a cells transfected with POMC-WT and -C28F. mRNA levels of each sample are shown below. (C) Western blot analyses of ubiquitination following immunoprecipitation of POMC in HEK293T cells transfected with POMC-WT-Flag or POMC-C28F-Flag construct with or without HA-Ub, Myc-tagged HRD1-WT, or HRD1 E3 ligase-dead C2A mutant. (D) Representative confocal images of POMC in POMC-transfected WT and Sel1L–/– N2a cells. White arrows point to secreted POMC in granules, while yellow arrows point to perinuclear POMC, possibly in the form of aggregates. KDEL marks the ER. (E) Sucrose gradient fractionation (fractions 1–7 from top to bottom of gradient) of HEK293T cells expressing POMC-WT or -C28F under nonreducing (–β-ME) and reducing (+β-ME) SDS-PAGE. (F) Western blot analysis of Myc-tagged POMC in WT and Hrd1–/– N2a cells transfected with POMC-WT or -C28F under reducing and nonreducing SDS-PAGE. Red box marks HMW aggregates, while green box marks monomers and dimers. HMW, high molecular weight. (POMC)2, POMC dimers. Representative data from at least 2 independent experiments are shown.