Effects of inhibiting transcription and protein synthesis on basal and insulin-stimulated leptin gene expression and leptin secretion in cultured rat adipocytes

María J. Moreno-Aliaga, Kimber Stanhope, Francine M. Gregoire, Craig H Warden, Peter J Havel

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

We have previously reported that glucose metabolism mediates the effects of insulin to increase leptin gene expression and leptin secretion by isolated adipocytes. The aim of the present study was to investigate the role of transcription and translation in the regulation of basal and insulin-stimulated leptin production. The short-term (4h) and long-term (24-48h) effects of actinomycin D, a transcriptional inhibitor, and cycloheximide, an inhibitor of protein synthesis, on leptin gene expression and leptin secretion by isolated adipocytes were determined. Actinomycin D (5μg/ml) increased both basal and insulin-stimulated (1.6nM) leptin secretion at 4 and 24h (193±14.9% and 153.8±10.4% of respective controls at 24h, both p<0.001). Similar effects of actinomycin D were observed on basal and insulin-stimulated leptin mRNA levels. 5,6-Dichlororibofuranosyl benzimidazole (DRB), another inhibitor of transcription, also increased basal (175.4±18.2% of control; p<0.01) and insulin-stimulated leptin secretion (141.0±11.1% of insulin-treated cells; p<0.05) at 24h. The effect of actinomycin D and DRB to increase basal leptin secretion observed at 4 and 24h was not present at 48h when actinomycin D and DRB both markedly inhibited insulin-stimulated leptin secretion (to 36±16%, p<0.05 and 21.9±5.6% of control, for actinomycin D and DRB, respectively, both p<0.001). Neither actinomycin D nor DRB had any effect on adipocyte glucose utilization between 24 and 48h. The observed effects of inhibitors of transcription on leptin gene expression and leptin secretion are consistent with a long-term transcriptional mechanism for insulin-stimulated glucose metabolism to increase leptin production. Cycloheximide treatment (10μg/ml) abolished the effects of insulin to stimulate leptin secretion (29±11% of control, p<0.01) during the first 4h of treatment and at all later time points, which indicate that de novo protein synthesis is required for insulin-mediated glucose metabolism to increase leptin secretion.

Original languageEnglish (US)
Pages (from-to)907-914
Number of pages8
JournalBiochemical and Biophysical Research Communications
Volume307
Issue number4
DOIs
StatePublished - Aug 8 2003

Fingerprint

Transcription
Leptin
Adipocytes
Gene expression
Rats
Insulin
Gene Expression
Dactinomycin
Proteins
Metabolism
Glucose
Cycloheximide
Protein Synthesis Inhibitors

Keywords

  • Actinomycin
  • Cycloheximide
  • Insulin-mediated glucose metabolism
  • Leptin
  • Transcription

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Effects of inhibiting transcription and protein synthesis on basal and insulin-stimulated leptin gene expression and leptin secretion in cultured rat adipocytes. / Moreno-Aliaga, María J.; Stanhope, Kimber; Gregoire, Francine M.; Warden, Craig H; Havel, Peter J.

In: Biochemical and Biophysical Research Communications, Vol. 307, No. 4, 08.08.2003, p. 907-914.

Research output: Contribution to journalArticle

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abstract = "We have previously reported that glucose metabolism mediates the effects of insulin to increase leptin gene expression and leptin secretion by isolated adipocytes. The aim of the present study was to investigate the role of transcription and translation in the regulation of basal and insulin-stimulated leptin production. The short-term (4h) and long-term (24-48h) effects of actinomycin D, a transcriptional inhibitor, and cycloheximide, an inhibitor of protein synthesis, on leptin gene expression and leptin secretion by isolated adipocytes were determined. Actinomycin D (5μg/ml) increased both basal and insulin-stimulated (1.6nM) leptin secretion at 4 and 24h (193±14.9{\%} and 153.8±10.4{\%} of respective controls at 24h, both p<0.001). Similar effects of actinomycin D were observed on basal and insulin-stimulated leptin mRNA levels. 5,6-Dichlororibofuranosyl benzimidazole (DRB), another inhibitor of transcription, also increased basal (175.4±18.2{\%} of control; p<0.01) and insulin-stimulated leptin secretion (141.0±11.1{\%} of insulin-treated cells; p<0.05) at 24h. The effect of actinomycin D and DRB to increase basal leptin secretion observed at 4 and 24h was not present at 48h when actinomycin D and DRB both markedly inhibited insulin-stimulated leptin secretion (to 36±16{\%}, p<0.05 and 21.9±5.6{\%} of control, for actinomycin D and DRB, respectively, both p<0.001). Neither actinomycin D nor DRB had any effect on adipocyte glucose utilization between 24 and 48h. The observed effects of inhibitors of transcription on leptin gene expression and leptin secretion are consistent with a long-term transcriptional mechanism for insulin-stimulated glucose metabolism to increase leptin production. Cycloheximide treatment (10μg/ml) abolished the effects of insulin to stimulate leptin secretion (29±11{\%} of control, p<0.01) during the first 4h of treatment and at all later time points, which indicate that de novo protein synthesis is required for insulin-mediated glucose metabolism to increase leptin secretion.",
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AU - Havel, Peter J

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AB - We have previously reported that glucose metabolism mediates the effects of insulin to increase leptin gene expression and leptin secretion by isolated adipocytes. The aim of the present study was to investigate the role of transcription and translation in the regulation of basal and insulin-stimulated leptin production. The short-term (4h) and long-term (24-48h) effects of actinomycin D, a transcriptional inhibitor, and cycloheximide, an inhibitor of protein synthesis, on leptin gene expression and leptin secretion by isolated adipocytes were determined. Actinomycin D (5μg/ml) increased both basal and insulin-stimulated (1.6nM) leptin secretion at 4 and 24h (193±14.9% and 153.8±10.4% of respective controls at 24h, both p<0.001). Similar effects of actinomycin D were observed on basal and insulin-stimulated leptin mRNA levels. 5,6-Dichlororibofuranosyl benzimidazole (DRB), another inhibitor of transcription, also increased basal (175.4±18.2% of control; p<0.01) and insulin-stimulated leptin secretion (141.0±11.1% of insulin-treated cells; p<0.05) at 24h. The effect of actinomycin D and DRB to increase basal leptin secretion observed at 4 and 24h was not present at 48h when actinomycin D and DRB both markedly inhibited insulin-stimulated leptin secretion (to 36±16%, p<0.05 and 21.9±5.6% of control, for actinomycin D and DRB, respectively, both p<0.001). Neither actinomycin D nor DRB had any effect on adipocyte glucose utilization between 24 and 48h. The observed effects of inhibitors of transcription on leptin gene expression and leptin secretion are consistent with a long-term transcriptional mechanism for insulin-stimulated glucose metabolism to increase leptin production. Cycloheximide treatment (10μg/ml) abolished the effects of insulin to stimulate leptin secretion (29±11% of control, p<0.01) during the first 4h of treatment and at all later time points, which indicate that de novo protein synthesis is required for insulin-mediated glucose metabolism to increase leptin secretion.

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