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Evolution of pyruvate carboxylase and other biotin containing enzymes in developing rat liver and kidney
R Salto1, M D Girón, M del Mar Sola
1Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Granada, Campus de Cartuja, Spain.
Insights
Pyruvate carboxylase expression in rat liver and kidney develops differently during perinatal development, with liver showing significant activity and kidney remaining low. This study also confirms early biotinylation of carboxylases in both organs.
Area of Science:
- Biochemistry
- Developmental Biology
- Metabolic Regulation
Background:
- Pyruvate carboxylase (PC) is crucial for gluconeogenesis and anaplerosis.
- Its developmental regulation in rat liver and kidney during the perinatal period is not fully understood.
- Biotinylation is essential for carboxylase function, requiring holocarboxylase synthetase (HCS).
Purpose of the Study:
- To investigate the developmental expression of pyruvate carboxylase (PC) in rat liver and kidney during the perinatal period.
- To examine the biotinylation status of PC and other biotin-dependent carboxylases.
- To elucidate the role of renal gluconeogenesis during perinatal development.
Main Methods:
- Assay of PC activity, enzyme amount, and mRNA levels from 2 days before delivery to weaning.
- Western-blot analysis using streptavidin-biotin peroxidase to detect biotinylation of PC and other biotin-enzymes.
- Analysis of mitochondrial samples from liver and kidney.
Main Results:
- Liver PC showed distinct activity and mRNA peaks before and after parturition, while kidney PC remained low.
- All detected PC in both organs was fully biotinylated, indicating early HCS development.
- Other mitochondrial carboxylases, like propionyl-CoA carboxylase, were also differentially expressed, with lower levels in kidney.
Conclusions:
- Rat liver exhibits significant PC induction during perinatal development, supporting hepatic gluconeogenesis.
- Kidney PC expression is minimal, confirming a limited role for renal gluconeogenesis during this period.
- Holocarboxylase synthetase activity develops early, ensuring proper biotinylation of carboxylases in both liver and kidney.
Abstract:
The evolution of pyruvate carboxylase has been studied in rat liver and kidney during perinatal development. The pyruvate carboxylase activity, amount of enzyme and mRNA levels have been assayed from 2 days before delivery to weaning. In liver, there is a peak of activity and amount of enzyme 24 h before delivery and 2 peaks, at 12 h and 6 days, after parturition. The transcription of the enzyme gene followed a similar pattern, with mRNA peaks preceding those of activity and amount of enzyme. However, in kidney, pyruvate carboxylase activity, amount and mRNA remain low until weaning. These results confirm the limited role of renal gluconeogenesis during the perinatal development. Since all carboxylases contain biotin as prosthetic group, the biotinylation of pyruvate carboxylase during the perinatal period was investigated by western-blot using streptavidin-biotin peroxidase. In the mitochondrial samples from liver and kidney, all the pyruvate carboxylase detected was fully biotinylated, indicating an early development of the holocarboxylase synthetase activity in the perinatal period. This Western-blot technique also allowed us the detection of other biotin-enzymes based on their molecular weight. In liver, during the perinatal development propionyl-coA and 3-methyl-crotonyl-coA carboxylases followed a pattern of induction similar to pyruvate carboxylase. In kidney, the expression of mitochondrial carboxylases was lower compared to liver and propionyl-coA carboxylase was not detected during the studied period.