Identification of the leptin receptor sequences crucial for the STAT3-Independent control of metabolism

Tammy M. Barnes, Kimi Shah, Margaret B. Allison, Gabrielle K. Steinl, Desiree Gordian, Paul V. Sabatini, Abigail J. Tomlinson, Wenwen Cheng, Justin C. Jones, Qing Zhu, Chelsea Faber, Martin G. Myers Jr.

Leptin, a peptide hormone produced by white adipose tissue in proportion to energy stores, plays a central role in the control of feeding and energy balance. Leptin receptor (LepRb) mainly signals trough activation of signal transducer and activator of transcription 3 (STAT3); however, there is also a second, yet unidentified signal. Barnes et al. used CRISPR/Cas9-mediated mutagenesis to generate a panel of mouse lines containing truncations of LepRb. By studying these five novel mouse lines, they identified a region of the intracellular LepRb that is required to mediate the second signal and additionally found a region that mediates a previously undescribed LepRb inhibitory signal.

Objective: Leptin acts via its receptor, LepRb, on specialized neurons in the brain to modulate energy balance and glucose homeostasis. LepRb→STAT3 signaling plays a crucial role in leptin action, but LepRb also mediates an additional as-yet-unidentified signal (Signal 2) that is important for leptin action. Signal 2 requires LepRb regions in addition to those required for JAK2 activation but operates independently of STAT3 and LepRb phosphorylation sites.

Methods: To identify LepRb sequences that mediate Signal 2, we used CRISPR/Cas9 to generate five novel mouse lines containing COOH-terminal truncation mutants of LepRb. We analyzed the metabolic phenotype and measures of hypothalamic function for these mouse lines.

Results: We found that deletion of LepRb sequences between residues 921 and 960 dramatically worsens metabolic control and alters hypothalamic function relative to smaller truncations. We also found that deletion of the regions including residues 1013–1053 and 960–1013 each decreased obesity compared to deletions that included additional COOH-terminal residues.

Conclusions: LepRb sequences between residues 921 and 960 mediate the STAT3 and LepRb phosphorylation-independent second signal that contributes to the control of energy balance and metabolism by leptin/LepRb. In addition to confirming the inhibitory role of the region (residues 961–1013) containing Tyr985, we also identified the region containing residues 1013–1053 (which contains no Tyr residues) as a second potential mediator of LepRb inhibition. Thus, the intracellular domain of LepRb mediates multiple Tyr-independent signals.