Related Experiment Videos
Insulin regulation of malic enzyme gene expression in rat liver: evidence for nuclear proteins that bind to two
C García-Jiménez1, B Benito, T Jolin
1Instituto de Investigaciones Biomédicas Consejo Superior de Invetigaciones Científicas, Madrid, Spain.
Abstract:
Diabetes in rats is characterized by insulin deficiency accompanied by a decrease in lipogenic enzymes. The malic enzyme (ME) gene, which encodes an important lipogenic enzyme, was used to investigate insulin regulation of gene expression. ME mRNA levels were reduced by more than 90% in the liver of diabetic rats. The administration of insulin (3 U/15 days) to either control or diabetic rats increased ME mRNA by 2- to 10-fold, respectively. Since diabetes reduces circulating T3 and the levels of nuclear T3-receptors, the potential role of thyroid hormone on insulin regulation of ME gene expression was also evaluated in thyroidectomized-diabetic rats. In these animals the levels of ME mRNA were undetectable but were increased by insulin even in the absence of thyroid hormones. These in vivo effects of insulin and T3 were not additive. The transcription rate of the gene was also reduced in the diabetic liver and recovered after insulin therapy. By computer analyses we have identified two different putative insulin response elements (IREs) in the ME gene promoter, hereafter referred to as IRE-I (-683 to -692), which is similar to the phosphoenol pyruvate carboxy kinase promoter IRE and IRE-II (-161 to -170), which is similar to the glyceraldehyde phosphate dehydrogenase gene promoter IRE-A. Results from gel retardation assays suggest that a single nuclear protein binds to IRE-I whereas two different nuclear proteins bind to IRE-II. The protein/IRE-I complex increased in liver nuclear extracts from diabetic rats and decreased after insulin administration.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Insulin therapy restores malic enzyme (ME) gene expression in diabetic rats, even without thyroid hormone. This study identifies key DNA regions in the ME gene promoter that regulate insulin
Area of Science:
- Molecular Endocrinology
- Gene Regulation
- Metabolic Diseases
Background:
- Diabetes mellitus is characterized by insulin deficiency and reduced lipogenic enzyme activity.
- The malic enzyme (ME) gene encodes a key lipogenic enzyme crucial for metabolic regulation.
- Insulin's role in regulating ME gene expression is not fully understood, particularly in the context of diabetes and thyroid hormones.
Purpose of the Study:
- To investigate the regulation of malic enzyme (ME) gene expression by insulin in diabetic rats.
- To elucidate the role of thyroid hormone in insulin-mediated regulation of ME gene expression.
- To identify and characterize insulin response elements (IREs) within the ME gene promoter.
Main Methods:
- Quantitative analysis of ME mRNA levels in rat liver using Northern blotting.
- Administration of insulin and evaluation of its effects on ME mRNA and gene transcription.
- Experimental manipulation of thyroid hormone levels in diabetic rats.
- Computer analysis of the ME gene promoter to identify putative IREs.
- Gel retardation assays to study protein binding to identified IREs.
Main Results:
- Diabetes significantly reduced ME mRNA levels (>90%) and gene transcription in rat liver.
- Insulin administration increased ME mRNA levels in both control and diabetic rats.
- Insulin restored ME mRNA levels and transcription even in thyroidectomized-diabetic rats, indicating thyroid hormone independence.
- Two putative IREs (IRE-I and IRE-II) were identified in the ME gene promoter.
- Specific nuclear proteins bind to IRE-I and IRE-II, with altered binding observed in diabetic and insulin-treated rats.
Conclusions:
- Insulin plays a critical role in regulating ME gene expression and transcription, independent of thyroid hormone.
- The ME gene promoter contains functional insulin response elements that mediate insulin's regulatory effects.
- Altered nuclear protein binding to IRE-I in diabetic rats suggests a mechanism for insulin resistance at the gene expression level.