Cystathionine γ lyase–hydrogen sulfide increases peroxisome proliferator-activated receptor γ activity by sulfhydration at C139 site thereby promoting glucose uptake and lipid storage in adipocytes

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Highlights

  • CSE–H2S inhibits PDE activity in adipocyte.

  • H2S sulfhydrates PPARγ at C139 site.

  • PPARγ sulfhydration increases its activity thereby promotes glucose uptake and lipid storage.

  • H2S attenuated insulin resistance but did not accelerate obesity in HFD obese mice.

Abstract

Adipocytes express the cystathionine γ lyase (CSE)–hydrogen sulfide (H2S) system. CSE–H2S promotes adipogenesis but ameliorates adipocyte insulin resistance. We investigated the mechanism of how CSE–H2S induces these paradoxical effects. First, we confirmed that an H2S donor or CSE overexpression promoted adipocyte differentiation. Second, we found that H2S donor inhibited but CSE inhibition increased phosphodiesterase (PDE) activity. H2S replacing isobutylmethylxanthine in the differentiation program induced adipocyte differentiation in part. Inhibiting PDE activity by H2S induced peroxisome proliferator activated receptor γ (PPARγ) protein and mRNA expression. Of note, H2S directly sulfhydrated PPARγ protein. Sulfhydrated PPARγ increased its nuclear accumulation, DNA binding activity and adipogenesis gene expression, thereby increasing glucose uptake and lipid storage, which were blocked by the desulfhydration reagent DTT. H2S induced PPARγ sulfhydration, which was blocked by mutation of the C139 site of PPARγ. In mice fed a high-fat diet (HFD) for 4 weeks, the CSE inhibitor decreased but H2S donor increased adipocyte numbers. In obese mice fed an HFD for 13 weeks, H2S treatment increased PPARγ sulfhydration in adipose tissues and attenuated insulin resistance but did not increase obesity. In conclusion, CSE–H2S increased PPARγ activity by direct sulfhydration at the C139 site, thereby changing glucose into triglyceride storage in adipocytes. CSE–H2S-mediated PPARγ activation might be a new therapeutic target for diabetes associated with obesity.

Introduction

Obesity is a prevalent health hazard and is characterized by adipose tissue expansion due to increasing number of cells via adipocyte differentiation (hyperplasia) and large adipocyte size (hypertrophy) [1]. Obesity is associated with an increase in circulating levels of several amino acids such as homocysteine, cysteine, alanine, phenylalanine, and tyrosine [2], [3], [4], [5]. Cysteine is a metabolic product of the essential amino acid methionine and is synthesized by transsulfuration from homocysteine dependent on cystathionine β synthase (CBS) and cystathionine γ lyase (CSE) [6]. Several large epidemiological studies showed plasma total cysteine (tCys) positively associated with body mass index [2], [7], [8]. However, in animals or humans, intake of cysteine-rich protein did not increase fat mass [9]. Knockout of CBS [10] can reduce body and adipose tissue weight. In humans, homocysteinuria due to CBS deficiency [11] or CSE mutation [12] was associated with lipodystrophy; in contrast, Down's syndrome children with high CBS activity [13] exhibit obesity. Thus, endogenous cysteine biogenesis enzymes but not cysteine per se may be linked with obesity.

CSE is a key enzyme generating cysteine by hydrolyzing cystathionine. CSE and CBS also hydrolyze cysteine to produce hydrogen sulfide (H2S) [6]. Feeding CSE-knockout mice with cysteine-limited food decreased white adipose weight about two thirds as compared with wild-type mice [14], so the CSE–H2S system might be involved in the pathogenesis of obesity. Our previous work found that adipocytes expressed CSE and endogenously generated H2S [15], [16]; the CSE inhibitor dl-propargylglycine (PPG) increased lipolysis in adipocytes and blocked obesity induced by a high-fat diet (HFD) [17]. Thus, the CSE–H2S system affects adipocyte hypertrophy by regulating lipolysis during obesity. Recently, Tsai et al. reported that H2S promotes adipogenesis [18], so H2S might increase adipocyte hyperplasia causing obesity. H2S increased insulin sensitivity in adipocytes with high glucose treatment [19], TNF-α [20] or diabetic mice induced by an HFD [17]. Whether CSE/H2S promotes glucose utility to storage in lipid droplets in adipocytes is unknown as is its possible molecular mechanism.

In the present study, we investigated the CSE–H2S effects on exchange of glucose to triglycerides by sulfhydating peroxisome proliferator-activated receptor γ (PPARγ) in adipocytes.

Section snippets

Animals and materials

All animal procedures complied with the Animal Management Rule of the Ministry of Health, People's Republic of China (document no. 55, 2001) and the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH publication no. 85-23, updated 2011). The care and use of laboratory animals were approved by the Laboratory Animal Ethics Committee of Peking University. A total of 75 C57BL/6J mice (18–20 g, 10 weeks old) were supplied by the Animal Center, Peking University

CSE–H2S system promoted triglyceride accumulation in adipocyte differentiation

3T3L1 preadipocytes expressed CSE mRNA (Fig. S1A) and protein (Fig. S1B), which was upregulated during differentiation into mature adipocytes (Fig. S1A and C, all P < 0.01). Consistent with changes in CSE expression, H2S production was significantly increased during adipocyte differentiation (Fig. S1D). Thus, the CSE–H2S system was upregulated in adipocyte differentiation.

Treating 3T3L1 preadipocytes in the differentiation cocktail with dl-propargylglycine (PPG, 100 μM; the CSE inhibitor)

Discussion

CSE is a key enzyme to catalyze cystathionine generation l-cysteine, concomitantly releasing H2S [6]. CSE-knockout mice fed an l-cysteine-deficient diet showed white fat loss [14]. In contrast, HFD feeding lowered CSE protein expression and H2S production [17]. Therefore, adipocyte CSE–H2S system dysfunction may contribute to obesity. Obese adipocytes are always associated with impaired insulin sensitivity. Interestingly, high glucose downregulated the CSE–H2S system [27], whereas H2S increased

Conflicts of interest

The authors declare no conflicts of interest.

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Acknowledgments

This work was supported by the Major State Basic Research Development Program of the People's Republic of China (no. 2012CB517806) and the National Natural Science Foundation of China (nos. 81170235, 81470552, 91339106 and 91439119).

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    These authors contributed equally to this work.

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