No abstract

Mouse vas deferens protein (MVDP) is a member of the aldo-keto reductase superfamily. The regulation of MVDP gene expression by activators of the protein kinase A signalling pathway was investigated in human (H295-R) and murine (Y1) adrenocortical carcinoma cells. Immunoblotting with polyclonal antibodies showed that MVDP is expressed in adrenal glands from mouse, rat, rabbit and guinea-pig, probably under the control of ACTH. In both adrenocortical cell lines used, MVDP is constitutively synthesized and its accumulation is increased by treatment with cAMP or forskolin. MVDP mRNA steady-state levels were up-regulated by forskolin in adrenocortical cells by a process that does not require de novo protein synthesis. The results suggest that cAMP is at least one of the key regulators of adrenal MVDP expression and that this effect is direct.

Studies of ACTH functions in adrenal steroidogenesis have been facilitated by the availability of immortalized mouse adrenocortical Y1 cells. In order to obtain alternative cell lines with a more differentiated zona fasciculata (ZF) phenotype we used targeted tumorigenesis strategy. We have generated transgenic mice expressing the SV40 T antigen under the control of the ACTH-dependent promoter for the AKR1B7/MVDP gene (aldo-keto reductase 1B7/mouse vas deferens protein), which encodes an enzyme responsible for detoxifying isocaproaldehyde, the product of side-chain cleavage of cholesterol generated by steroidogenesis. Our previous data indicated that in the mouse adrenal, AKR1B7 expression was restricted to the ZF and that a 0.5-kb promoter region was able to target specific adrenal expression in transgenic mice. In situ hybridization analyses indicate that AKR1B7 expression during fetal and post-natal periods paralleled the onset of glucocorticoid synthesis and the development of ZF. In transgenic mice, ACTH control and developmental programming of the CAT gene driven by the 0.5-kb promoter followed endogenous gene regulation. Then transgenic mice harboring the 0.5-kb/SV40 T antigen construct were generated and two founders out of three developed adrenal tumors. Cells derived from the tumor of founder 1 (ATC1) were grown in presence of forskolin to maintain ACTH receptor expression and were tested for ACTH responsiveness by immunocychemistry and northern blot analyses. Even after several passages, the ACTH induced AKR1B7 and P450c11beta mRNAs accumulations were similar to that observed in mouse primary adrenocortical cell cultures. Our findings suggest that ATC1 cells have conserved essential features of ZF cells. In order to achieve complete characterization of these cells further analyses are currently performed to investigate their steroidogenic activity.