Cytochrome P450 Transcriptional Regulation by Testis‐Specific Y‐Encoded‐Like Protein: Identification of Novel Upstream Transcription Factors

Cytochrome P450s (CYPs) display significant inter-individual variation in expression, much of which remains unexplained by known CYPsingle-nucleotide polymorphisms (SNPs). Testis‐specific Y‐encoded‐like proteins (TSPYLs) are transcriptional regulators for several drug-metabolizing CYPsincluding CYP3A4. However, transcription factors (TFs) that might influence CYPexpression through an effect on TSPYLexpression are unknown. Therefore, we studied regulators of TSPYLexpression in hepatic cell lines and their possible SNP-dependent variation. Specifically, we identified candidate TFs that might influence TSPYLexpression using the ENCODE ChIPseq database. Subsequently, the expression of TSPYL1/2/4as well as that of selected CYP targets for TSPYLregulation were assayed in hepatic cell lines before and after knockdown of TFs that might influence CYPexpression through TSPYL-dependent mechanisms. Those results were confirmed by studies of TF binding to TSPYL1/2/4gene promoter regions. In hepatic cell lines, knockdown of the REST and ZBTB7A TFs resulted in decreased TSPYL1and TSPYL4expression and increased CYP3A4expression, changes reversed by TSPYL1/4overexpression. Potential binding sites for RESTand ZBTB7Aon the promoters of TSPYL1and TSPYL4were confirmed by chromatin immunoprecipitation. Finally, common SNP variants in upstream binding sites on the TSPYL1/4promoters were identified and luciferase reporter constructs confirmed SNP-dependent modulation of TSPYL1/4gene transcription. In summary, we identified REST and ZBTB7A as regulators of the expression of TSPYL genes which themselves can contribute to regulation of CYPexpression and—potentially—of drug metabolism. SNP-dependent modulation of TSPYL transcription may contribute to individual variation in both CYPexpression and—downstream–drug response phenotypes.SIGNIFICANCE STATEMENTTestis-specific Y‐encoded‐like proteins (TSPYLs) are transcriptional regulators of cytochrome P450 (CYP) gene expression. Here, we report that variation in TSPYLexpression as a result of the effects of genetically regulated TSPYL transcription factors is an additional factor that could result in downstream variation in CYPexpression and potentially, as a result, variation in drug biotransformation.