Cloning and expression of rat cDNA encoding corticosteroid 11 beta-dehydrogenase

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• Corticosteroid 11 beta-dehydrogenase (11-DH) inactivates cortisol to cortisone.
• Deficiency in 11-DH activity causes apparent mineralocorticoid excess, a severe childhood hypertension.
• Understanding the molecular basis of 11-DH is critical for this disorder.

Purpose of the Study:

• To isolate and characterize a rat cDNA clone encoding 11-DH.
• To investigate the molecular basis of apparent mineralocorticoid excess.
• To identify potential new gene families related to dehydrogenases.

Main Methods:

• Isolation and characterization of a rat cDNA clone for 11-DH.
• Northern blot analysis to detect mRNA expression in various tissues.
• DNA sequence analysis and comparison with known dehydrogenases.
• Expression of the 11-DH cDNA in Chinese hamster ovary cells.
• Southern blot analysis of rat and human DNA.

Main Results:

• A 1265 bp rat cDNA clone encoding 11-DH (287 amino acids) was isolated.
• 11-DH mRNA was detected in liver, kidney, and testis, but not heart.
• Sequence analysis revealed 11-DH belongs to a new dehydrogenase superfamily, sharing homology with bacterial dehydrogenases.
• Expressed 11-DH exhibited both 11 beta-dehydrogenation and 11-oxoreduction activity.
• Southern blots suggest related genes exist in both rat and human genomes.

Conclusions:

• The study successfully isolated and characterized the rat 11-DH cDNA.
• The findings establish 11-DH as a member of a novel dehydrogenase superfamily.
• This work provides a foundation for understanding apparent mineralocorticoid excess and related genetic disorders.