RAT HEPATIC GLUTAMINASE - IDENTIFICATION OF THE FULL CODING SEQUENCE AND CHARACTERIZATION OF A FUNCTIONAL PROMOTER

Glutamine catabolism in mammalian liver is catalysed by a unique isoen zyme of phosphate-activated glutaminase. The full coding and 5' untran slated sequence for rat hepatic glutaminase was isolated by screening lambda ZAP cDNA libraries and a Charon 4a rat genomic library, The seq uence produces a mRNA 2225 nt in length, encoding a polypeptide of 535 amino acid residues with a calculated molecular mass of 59.2 kDa. The deduced amino acid sequence of rat liver glutaminase shows 86% simila rity to that of rat kidney glutaminase and 65% similarity to a putativ e glutaminase from Caenorhabditis elegans. A genomic clone to rat live r glutaminase was isolated that contains 3.5 kb of the gene and 7.5 kb of the 5' flanking region. The 1 kb immediately upstream of the hepat ic glutaminase gene (from -1022 to +48) showed functional promoter act ivity in HepG2 hepatoma cells. This promoter region did not respond to treatment with cAMP, but was highly responsive (10-fold stimulation) to the synthetic glucocorticoid dexamethasone. Subsequent 5' deletion analysis indicated that the promoter region between -103 and +48 was s ufficient for basal promoter activity. This region does not contain an identifiable TATA element, indicating that transcription of the gluta minase gene is driven by a TATA-less promoter. The region responsive t o glucocorticoids was mapped to -252 to -103 relative to the transcrip tion start site.