Abstract
Cysteine conjugate β-lyase (CS-lyase) [glutamine transaminase K (GTK)/kynurenine aminotransferase (KAT)] is involved in the toxicity of certain halogenated alkenes by metabolising their corresponding cysteine S-conjugates into reactive metabolites. These are known to produce marked nephrotoxicity and neurotoxicity in experimental animals. The KAT activity has been linked to neuromodulatory pathways which are the target of therapeutic strategies for the treatment of neurodegenerative disorders. There is substantial evidence to suggest that CS-lyase/GTK/KAT is hormonally modulated following reports of differences in the levels of activity of this enzyme between male and female rodents, and between young and mature animals. Therefore, to understand in greater depth the regulation of CS-lyase/GTK/KAT gene expression, the ability of hormones to modulate this enzyme was studied.The ability of the glucocorticoid and sex hormones to modulate CS-lyase/GTK/KAT activity in male rat brain and kidney, during post-natal development, was studied. Corticosterone treatment significantly increased GTK specific activity in both kidney and brain tissue. This induction was observed at post-natal days 19,21,23,25 and 30. Following administration of dexamethasone, induction was also observed, however, this was restricted to post-natal day 21 in both tissues. Progesterone was the only sex hormone found to induce GTK activity in post-natal day 21 male rat kidney and brain. The pattern of induction was comparable to the effect produced by dexamethasone treatment. GTK activity was induced at post-natal day 21 in the kidney and post-natal days 21,23 and 25 in the brain. This study clearly demonstrated an age-related influence on the susceptibility of GTK to induction by these hormones which supports the proposal that CS-lyase/GTK/KAT is developmentally modulated in the post-natal male rat.Long distance PCR techniques have been employed to isolate and sequence the upstream regulatory region from the rat CS-lyase/GTK/KAT gene and to identify hormone response elements. Using gene-specific primers, designed against the non-coding region of the known rat renal CS-lyase cDNA sequence, 'genomewalking' experiments yielded a 1kb sequence. Following, cloning into a pCR®2.1-TOPO TA vector and DNA sequencing of the 1kb insert, no hormone response elements or other regulatory motifs were identified. TATA and CAAT boxes were present in the sequence, indicating that a region of the upstream sequence had been isolated. This suggests that the regulatory sequence motifs are located further upstream.