Reducing 14-3-3ζ expression influences adipocyte maturity and impairs function

Abstract

One of the primary metabolic functions of a mature adipocyte is to supply energy via lipolysis, or the catabolism of stored lipids. Adipose triacylglycerol lipase (ATGL) and hormone-sensitive lipase (HSL) are critical lipolytic enzymes, and their phosphorylation generates phospho-binding sites for 14-3-3 proteins, a ubiquitously expressed family of molecular scaffolds. Although we previously identified essential roles of the 14-3-3ζ isoform in murine adipogenesis, the presence of 14-3-3 protein binding sites on ATGL and HSL suggests that 14-3-3ζ could also influence mature adipocyte processes like lipolysis. Here we demonstrate that 14-3-3ζ is necessary for lipolysis in male mice and fully differentiated 3T3-L1 adipocytes, as depletion of 14-3-3ζ significantly impaired glycerol and free fatty acid (FFA) release. Unexpectedly, reducing 14-3-3ζ expression was found to significantly impact adipocyte maturity, as observed by reduced abundance of peroxisome proliferator-activated receptor (PPAR)γ2 protein and expression of mature adipocyte genes and those associated with de novo triglyceride synthesis and lipolysis. The impact of 14-3-3ζ depletion on adipocyte maturity was further examined with untargeted lipidomics, which revealed that reductions in 14-3-3ζ abundance promoted the acquisition of a lipidomic signature that resembled undifferentiated preadipocytes. Collectively, these findings reveal a novel aspect of 14-3-3ζ in adipocytes, as reducing 14-3-3ζ was found to have a negative effect on adipocyte maturity and adipocyte-specific processes like lipolysis.

Regulation of lipolysis by 14-3-3 proteins on human adipocyte lipid droplets

“…In the context of adipocyte function, 14-3-3 proteins have been reported to interact with ACACA, ACLY, and FASN (32), suggesting a key role in the regulation of fatty acid biosynthesis. YWHAZ has been implicated in adipocyte differentiation, potentially through effects on RNA splicing (33)(34)(35), and consistent with our finding that knockdown of YHWAZ, but not other isoforms, impairs human adipocyte differentiation. Interestingly, YWHAB has been implicated in insulin signaling, through interactions with the insulin receptor, IRS1 and also phosphodiesterase 3B.…”

Plakoglobin regulates adipocyte differentiation independently of the Wnt/β-catenin signalling pathway

“…hADSCs are maintained for an additional week in induction media without rosiglitazone and IBMX. To assess lipid incorporation, differentiated cells were stained with Oil Red-O (ORO) and absorbance of ORO was quantified, as previously described (24,28).…”

“…NIH-3T3 and 3T3-L1 pre-adipocyte cells (ZenBio, Durham, North Carolina) were maintained in 25mM glucose DMEM (Life Technologies Corporation, Grand Island, NY), supplemented with 10% newborn calf serum (NBCS) and 1% penicillin/streptomycin (P/S). Differentiation of confluent 3T3-L1 cells was induced by a cocktail of 500 μM IBMX (Sigma-Aldrich, Oakville, Ontario), 500 nM dexamethasone (Sigma-Aldrich) and 172 nM insulin (Sigma-Aldrich) for 48 hours, as previously described (24,25,28), and cells were maintained with 25mM glucose DMEM with 10% FBS, 1% P/S and 172 nM insulin for up to 7 days. Human adipocyte stem cells (hADSCs) (ZenBio) were maintained in DMEM/F-12 (Wisent, Saint-Bruno, Quebec) supplemented with 10% FBS and 1% P/S.…”

14-3-3ζ regulates adipogenesis by modulating chromatin accessibility during the early stages of adipocyte differentiation

“…Even within a given adipocyte (i.e., Adipoq+) or APC (i.e., Pdgfra+) population, different niches exist with different cellular and physiological fates (23,(79)(80)(81). With our previous discovery of heterogenous expression of 14-3-3ζ among mature gonadal adipocytes (16) and the heterogeneous expression of Ywhaz among APCs (Sup. Fig.…”

“…Through recognition of specific phosphorylated serine or threonine motifs (RSXpS/TXP and RXXXpS/TXP), all seven mammalian 14-3-3 protein isoforms interact with a broad variety of enzymes, transcription factors, and transporters. Thus, 14-3-3 proteins can regulate diverse cellular processes, such as cell cycle progression (8,9), apoptosis (8,10), secretion, and metabolism (7,9,(11)(12)(13)(14)(15)(16)(17)(18). Despite a high degree of sequence-homology, 14-3-3 family members can perform isoformspecific biological functions, and our group has identified critical roles of the 14-3-3ζ isoform in the regulation of whole-body adiposity, adipogenesis, adipocyte function, and glucose (7,9,(12)(13)(14)(15)(16)(17).…”