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Endoplasmic reticulum stress regulates adipocyte resistin expression.

Lefterova MI, Mullican SE, Tomaru T, Qatanani M, Schupp M, Lazar MA - Diabetes (2009)

Bottom Line: The effects of endoplasmic stress inducers on resistin mRNA and secreted protein levels were examined in differentiated 3T3-L1 adipocytes, focusing on the expression and genomic binding of transcriptional regulators of resistin.ER stress reduced resistin mRNA in 3T3-L1 adipocytes in a time- and dose-dependent manner.The effects of ER stress were transcriptional because of downregulation of CAAT/enhancer binding protein-alpha and peroxisome proliferator-activated receptor-gamma transcriptional activators and upregulation of the transcriptional repressor CAAT/enhancer binding protein homologous protein-10 (CHOP10).

View Article: PubMed Central - PubMed

Affiliation: Division of Endocrinology, Diabetes, and Metabolism, Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania.

ABSTRACT

Objective: Resistin is a secreted polypeptide that impairs glucose metabolism and, in rodents, is derived exclusively from adipocytes. In murine obesity, resistin circulates at elevated levels but its gene expression in adipose tissue is paradoxically reduced. The mechanism behind the downregulation of resistin mRNA is poorly understood. We investigated whether endoplasmic reticulum (ER) stress, which is characteristic of obese adipose tissue, regulates resistin expression in cultured mouse adipocytes.

Research design and methods: The effects of endoplasmic stress inducers on resistin mRNA and secreted protein levels were examined in differentiated 3T3-L1 adipocytes, focusing on the expression and genomic binding of transcriptional regulators of resistin. The association between downregulated resistin mRNA and induction of ER stress was also investigated in the adipose tissue of mice fed a high-fat diet.

Results: ER stress reduced resistin mRNA in 3T3-L1 adipocytes in a time- and dose-dependent manner. The effects of ER stress were transcriptional because of downregulation of CAAT/enhancer binding protein-alpha and peroxisome proliferator-activated receptor-gamma transcriptional activators and upregulation of the transcriptional repressor CAAT/enhancer binding protein homologous protein-10 (CHOP10). Resistin protein was also substantially downregulated, showing a close correspondence with mRNA levels in 3T3-L1 adipocytes as well as in the fat pads of obese mice.

Conclusions: ER stress is a potent regulator of resistin, suggesting that ER stress may underlie the local downregulation of resistin mRNA and protein in fat in murine obesity. The paradoxical increase in plasma may be because of various systemic abnormalities associated with obesity and insulin resistance.

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Endoplasmic reticulum stress downregulates resistin in adipocytes by a transcriptional mechanism. A: Changes in resistin mRNA levels over 48 h in response to vehicle or 5 μg/ml tunicamycin in the presence of 5 μg/ml Actinomycin D. Data are presented as percent of the mRNA level at time 0, mean ± SD of a triplicate experiment. B: Normalized luciferase activity of resistin-luc (−13,580 bp to +243 bp) or empty vector (pGL3 basic) in the presence of vehicle or 5 μg/ml tunicamycin for 24 h. Data are presented as mean ± SE, n = 3. **P = 0.003 for pGL3 basic versus resistin-luc activity in vehicle-treated cells. ##P = 0.005 for resistin-luc activity in vehicle- versus tunicamycin-treated cells.
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Figure 3: Endoplasmic reticulum stress downregulates resistin in adipocytes by a transcriptional mechanism. A: Changes in resistin mRNA levels over 48 h in response to vehicle or 5 μg/ml tunicamycin in the presence of 5 μg/ml Actinomycin D. Data are presented as percent of the mRNA level at time 0, mean ± SD of a triplicate experiment. B: Normalized luciferase activity of resistin-luc (−13,580 bp to +243 bp) or empty vector (pGL3 basic) in the presence of vehicle or 5 μg/ml tunicamycin for 24 h. Data are presented as mean ± SE, n = 3. **P = 0.003 for pGL3 basic versus resistin-luc activity in vehicle-treated cells. ##P = 0.005 for resistin-luc activity in vehicle- versus tunicamycin-treated cells.

Mentions: An initial step in dissecting the mechanism by which ER stress regulates resistin expression was to determine whether the downregulation of resistin by tunicamycin is transcriptional. For this purpose, 3T3-L1 adipocytes were treated with 5 μg/ml tunicamycin in the presence of 5 μg/ml of the transcriptional inhibitor Actinomycin D. Tunicamycin treatment did not reduce the half-life of resistin mRNA as would have been expected if ER stress reduced resistin mRNA by a post-transcriptional mechanism (Fig. 3A). The effect of ER stress was further explored using a luciferase reporter vector (resistin-luc) driven by a large fragment of the resistin gene including the promoter and transcriptional start site (−13,580 bp to +243 bp). The resistin-luc reporter was active in mature adipocytes, but most of this activity was lost when the cells were treated with 5 μg/ml tunicamycin for 24 h (Fig. 3B), further demonstrating that ER stress causes reduced resistin gene transcription.


Endoplasmic reticulum stress regulates adipocyte resistin expression.

Lefterova MI, Mullican SE, Tomaru T, Qatanani M, Schupp M, Lazar MA - Diabetes (2009)

Endoplasmic reticulum stress downregulates resistin in adipocytes by a transcriptional mechanism. A: Changes in resistin mRNA levels over 48 h in response to vehicle or 5 μg/ml tunicamycin in the presence of 5 μg/ml Actinomycin D. Data are presented as percent of the mRNA level at time 0, mean ± SD of a triplicate experiment. B: Normalized luciferase activity of resistin-luc (−13,580 bp to +243 bp) or empty vector (pGL3 basic) in the presence of vehicle or 5 μg/ml tunicamycin for 24 h. Data are presented as mean ± SE, n = 3. **P = 0.003 for pGL3 basic versus resistin-luc activity in vehicle-treated cells. ##P = 0.005 for resistin-luc activity in vehicle- versus tunicamycin-treated cells.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2712799&req=5

Figure 3: Endoplasmic reticulum stress downregulates resistin in adipocytes by a transcriptional mechanism. A: Changes in resistin mRNA levels over 48 h in response to vehicle or 5 μg/ml tunicamycin in the presence of 5 μg/ml Actinomycin D. Data are presented as percent of the mRNA level at time 0, mean ± SD of a triplicate experiment. B: Normalized luciferase activity of resistin-luc (−13,580 bp to +243 bp) or empty vector (pGL3 basic) in the presence of vehicle or 5 μg/ml tunicamycin for 24 h. Data are presented as mean ± SE, n = 3. **P = 0.003 for pGL3 basic versus resistin-luc activity in vehicle-treated cells. ##P = 0.005 for resistin-luc activity in vehicle- versus tunicamycin-treated cells.
Mentions: An initial step in dissecting the mechanism by which ER stress regulates resistin expression was to determine whether the downregulation of resistin by tunicamycin is transcriptional. For this purpose, 3T3-L1 adipocytes were treated with 5 μg/ml tunicamycin in the presence of 5 μg/ml of the transcriptional inhibitor Actinomycin D. Tunicamycin treatment did not reduce the half-life of resistin mRNA as would have been expected if ER stress reduced resistin mRNA by a post-transcriptional mechanism (Fig. 3A). The effect of ER stress was further explored using a luciferase reporter vector (resistin-luc) driven by a large fragment of the resistin gene including the promoter and transcriptional start site (−13,580 bp to +243 bp). The resistin-luc reporter was active in mature adipocytes, but most of this activity was lost when the cells were treated with 5 μg/ml tunicamycin for 24 h (Fig. 3B), further demonstrating that ER stress causes reduced resistin gene transcription.

Bottom Line: The effects of endoplasmic stress inducers on resistin mRNA and secreted protein levels were examined in differentiated 3T3-L1 adipocytes, focusing on the expression and genomic binding of transcriptional regulators of resistin.ER stress reduced resistin mRNA in 3T3-L1 adipocytes in a time- and dose-dependent manner.The effects of ER stress were transcriptional because of downregulation of CAAT/enhancer binding protein-alpha and peroxisome proliferator-activated receptor-gamma transcriptional activators and upregulation of the transcriptional repressor CAAT/enhancer binding protein homologous protein-10 (CHOP10).

View Article: PubMed Central - PubMed

Affiliation: Division of Endocrinology, Diabetes, and Metabolism, Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania.

ABSTRACT

Objective: Resistin is a secreted polypeptide that impairs glucose metabolism and, in rodents, is derived exclusively from adipocytes. In murine obesity, resistin circulates at elevated levels but its gene expression in adipose tissue is paradoxically reduced. The mechanism behind the downregulation of resistin mRNA is poorly understood. We investigated whether endoplasmic reticulum (ER) stress, which is characteristic of obese adipose tissue, regulates resistin expression in cultured mouse adipocytes.

Research design and methods: The effects of endoplasmic stress inducers on resistin mRNA and secreted protein levels were examined in differentiated 3T3-L1 adipocytes, focusing on the expression and genomic binding of transcriptional regulators of resistin. The association between downregulated resistin mRNA and induction of ER stress was also investigated in the adipose tissue of mice fed a high-fat diet.

Results: ER stress reduced resistin mRNA in 3T3-L1 adipocytes in a time- and dose-dependent manner. The effects of ER stress were transcriptional because of downregulation of CAAT/enhancer binding protein-alpha and peroxisome proliferator-activated receptor-gamma transcriptional activators and upregulation of the transcriptional repressor CAAT/enhancer binding protein homologous protein-10 (CHOP10). Resistin protein was also substantially downregulated, showing a close correspondence with mRNA levels in 3T3-L1 adipocytes as well as in the fat pads of obese mice.

Conclusions: ER stress is a potent regulator of resistin, suggesting that ER stress may underlie the local downregulation of resistin mRNA and protein in fat in murine obesity. The paradoxical increase in plasma may be because of various systemic abnormalities associated with obesity and insulin resistance.

Show MeSH
Related in: MedlinePlus