Abstract
Both the magnitude and duration of insulin signaling are important in executing its cellular functions. Insulin-induced degradation of insulin receptor substrate 1 (IRS1) represents a key negative feedback loop that restricts insulin signaling. Moreover, high concentrations of fatty acids (FAs) and glucose involved in the etiology of obesity-associated insulin resistance also contribute to the regulation of IRS1 degradation. The scavenger receptor CD36 binds many lipid ligands, and its contribution to insulin resistance has been extensively studied, but the exact regulation of insulin sensitivity by CD36 is highly controversial. Herein, we found that CD36 knockdown in C2C12 myotubes accelerated insulin-stimulated Akt activation, but the activated signaling was sustained for a much shorter period of time as compared with WT cells, leading to exacerbated insulin-induced insulin resistance. This was likely due to enhanced insulin-induced IRS1 degradation after CD36 knockdown. Overexpression of WT CD36, but not a ubiquitination-defective CD36 mutant, delayed IRS1 degradation. We also found that CD36 functioned through ubiquitination-dependent binding to IRS1 and inhibiting its interaction with cullin 7, a key component of the multisubunit cullin-RING E3 ubiquitin ligase complex. Moreover, dissociation of the Src family kinase Fyn from CD36 by free FAs or Fyn knockdown/inhibition accelerated insulin-induced IRS1 degradation, likely due to disrupted IRS1 interaction with CD36 and thus enhanced binding to cullin 7. In summary, we identified a CD36-dependent FA-sensing pathway that plays an important role in negative feedback regulation of insulin activation and may open up strategies for preventing or managing type 2 diabetes mellitus.
Highlights
Both the magnitude and duration of insulin signaling are important in executing its cellular functions
We found that Cluster of differentiation 36 (CD36) functioned through ubiquitination-dependent binding to insulin receptor substrate 1 (IRS1) and inhibiting its interaction with cullin 7, a key component of the multisubunit cullin-RING E3 ubiquitin ligase complex
Because IRS1 plays a critical role in the insulin signaling pathway and insulin-induced IRS1 degradation is a key negative feedback mechanism to maintain appropriate signal strength and duration, we further examined whether CD36 mediates dynamics of insulin signaling activation by regulating IRS1 stability
Summary
Previous studies suggested that CD36 regulation of insulin signaling may be highly dependent on nutrient and signaling states of the cells. The insulin-stimulated degradation of IRS1 in CD36 KD myotubes as well as in CHO/hIR cells overexpressing CD36 was completely blocked by MG132 (Fig. S1), suggesting that CD36 regulates proteasomal degradation of IRS1. The phosphorylation level of IRS1 at Ser-307 was increased in CD36 KD myotubes as compared with control myotubes (Fig. 4, C and D), which is consistent with the enhanced insulin-stimulated IRS1 degradation (Fig. 3). We examined the presence of endogenous IRS1-FynCD36 complex in myotubes Both CD36 and Fyn coimmunoprecipitated with IRS1, and insulin treatment, which inhibits CD36 ubiquitination [25], decreased its association with IRS1 (Fig. 5D). Inhibition of Fyn by Src inhibitor-1 (Src-1) decreased CD36 and increased CUL7 coimmunoprecipitated with IRS1 (Fig. 5F), suggesting a key role of Fyn and its kinase activity in CD36 regulation of IRS1 degradation. Fyn activity may be important in CD36 regulation of IRS1 degradation
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