Conditional regulation of the human CYP4X1 and CYP4Z1 genes
Section snippets
Cell culture
The human mammary carcinoma cell lines T47-D and MCF-7 were obtained from the American Type Culture Collection. T47-D cells were maintained in phenol red-free RPMI supplemented with 10% fetal bovine serum (FBS), insulin (0.2 U/ml), sodium pyruvate (1 mM), and penicillin/streptomycin. MCF-7 cells were grown in phenol red-free minimal essential medium with Earle’s salts, supplemented with 10% FBS, non-essential amino acids (0.1 mM), sodium pyruvate (1 mM), insulin (0.2 U/ml), and
Tissue expression profile and cloning of full-length human CYP4Z1 cDNA
The highest numbers of ESTs encoding CYP4Z1 have been reported from the mammary gland (http://www.ncbi.nlm.nih.gov/UniGene/ESTProfileViewer.cgi?uglist=Hs.176588). Additional ESTs for CYP4Z1 have also been described from liver, kidney, lung, uterus, and brain suggesting low levels of CYP4Z1 mRNA expression in these tissues. A real-time PCR assay was developed in order to examine CYP4Z1 mRNA levels in selected tissues including breast, kidney, and liver (Fig. 1). Two breast RNA samples exhibited
Discussion
The human CYP4Z1 gene is preferentially expressed in mammary tissue and is induced by dexamethasone or progesterone in the mammary carcinoma derived T47-D cell line, but only by dexamethasone in MCF-7 cells. The induction of CYP4Z1 mRNA by dexamethasone or progesterone was suppressed by the glucocorticoid and progesterone receptor antagonist mifepristone implicating these receptors in the activation of the CYP4Z1 gene.
Two of five tested breast RNA samples exhibited >15-fold elevated CYP4Z1 mRNA
Acknowledgments
We thank Dr. J.M. Lasker at the Institute for Biomedical Research, Hackensack University Medical Center, N.J. for providing the CYP4A11 antibody. This work was supported by United States Public Health Service Grant HD 04445. Facilities for computer-assisted sequence analysis, DNA sequencing, and the synthesis of oligonucleotides are supported in part by General Clinical Research Center Grant M01 RR00833 and by the Sam and Rose Stein Charitable Trust. D.R.B. was supported by a grant from the
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