Salt-inducible kinase inhibition promotes the adipocyte thermogenic program and adipose tissue browning

Abstract

Norepinephrine stimulates the adipose tissue thermogenic program through a beta-adrenergic receptor (Beta AR) - cyclic adenosine monophosphate (cAMP) - protein kinase A (PKA) signaling cascade. Previous work from our lab showed that noncanonical activation of the mechanistic target of rapamycin complex 1 (mTORC1) by PKA is required for the beta AR-stimulation of adipose tissue browning. However, the downstream events triggered by mTORC1 activation that drive this thermogenic response are not well understood. In this study, we used a proteomic approach of Stable Isotope Labeling by/with Amino acids in Cell culture (SILAC) to characterize the global protein phosphorylation profile in brown adipocytes with beta-agonist isoproterenol stimulation and identified salt-inducible kinase 3 (SIK3) as a candidate mTORC1 substrate. SIK3 interacts with RAPTOR, the defining component of the mTORC1 complex, and is phosphorylated at Ser 884 in a mTORC1-dependent manner. Pharmacological SIK inhibition by a pan-SIK inhibitor (HG-9-91-01) in brown adipocytes increases basal Ucp1 gene expression and restores its expression upon either mTORC1 or PKA blockades. Short-hairpin RNA knockdown of Sik3 augments, while overexpression of SIK3 suppresses, Ucp1 gene expression in brown adipocytes. In addition, CRISPR gene-edited Sik3-/- brown adipocytes show enhanced Ucp1 gene expression at basal conditions and after Iso stimulation. Finally, a SIK inhibitor well-tolerated in vivo (YKL-05-099) can stimulate adipose tissue browning and thermogenic gene expression in mouse subcutaneous inguinal adipose tissue, supporting an essential role of SIK inhibition in regulating the adipose tissue browning program in vivo. Taken together, our data reveal that salt-inducible kinases function as a phosphorylation switch for beta-adrenergic activation to drive the adipose thermogenic program and suggest that maneuvers targeting SIKs could be beneficial for obesity and related metabolic disease.