trichostatin-a and Carcinoma--Embryonal

Human Cripto-1 (CR-1) plays an important role in regulating embryonic development while also regulating various stages of tumor progression. However, mechanisms that regulate CR-1 expression during embryogenesis and tumorigenesis are still not well defined. In the present study, we investigated the effects of two nuclear receptors, liver receptor homolog (LRH)-1 and germ cell nuclear factor receptor (GCNF) and epigenetic modifications on CR-1 gene expression in NTERA-2 human embryonal carcinoma cells and in breast cancer cells. CR-1 expression in NTERA-2 cells was positively regulated by LRH-1 through direct binding to a DR0 element within the CR-1 promoter, while GCNF strongly suppressed CR-1 expression in these cells. In addition, the CR-1 promoter was unmethylated in NTERA-2 cells, while T47D, ZR75-1, and MCF7 breast cancer cells showed high levels of CR-1 promoter methylation and low CR-1 mRNA and protein expression. Treatment of breast cancer cells with a demethylating agent and histone deacetylase inhibitors reduced methylation of the CR-1 promoter and reactivated CR-1 mRNA and protein expression in these cells, promoting migration and invasion of breast cancer cells. Analysis of a breast cancer tissue array revealed that CR-1 was highly expressed in the majority of human breast tumors, suggesting that CR-1 expression in breast cancer cell lines might not be representative of in vivo expression. Collectively, these findings offer some insight into the transcriptional regulation of CR-1 gene expression and its critical role in the pathogenesis of human cancer.

Topics: Azacitidine; Binding Sites; Breast Neoplasms; Carcinoma, Ductal, Breast; Carcinoma, Embryonal; Cell Movement; Decitabine; DNA Methylation; DNA Modification Methylases; Dose-Response Relationship, Drug; Embryonal Carcinoma Stem Cells; Female; Gene Expression Regulation, Developmental; Gene Expression Regulation, Neoplastic; Genes, Reporter; GPI-Linked Proteins; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Intercellular Signaling Peptides and Proteins; Luciferases; MCF-7 Cells; Neoplasm Invasiveness; Neoplasm Proteins; Nuclear Receptor Subfamily 6, Group A, Member 1; Promoter Regions, Genetic; Receptors, Cytoplasmic and Nuclear; RNA Interference; RNA, Messenger; Time Factors; Tissue Array Analysis; Transcription, Genetic; Transfection; Tretinoin; Valproic Acid

Nestin is an intermediate filament protein expressed specifically in neural stem cells and progenitor cells of the central nervous system. DNA demethylation and histone modifications are two types of epigenetic modifications working in a coordinate or synergistic manner to regulate the expression of various genes. This study investigated and elucidated the epigenetic regulation of Nestin gene expression during embryonic differentiation along the neural cell lineage. Nestin exhibits differential DNA methylation and histone acetylation patterns in Nestin-expressing and nonexpressing cells. In P19 embryonic carcinoma cells, activation of Nestin expression is mediated by both trichostatin A and 5-aza-2'-deoxycytidine treatment, concomitant with histone acetylation, but not with DNA demethylation. Nestin transcription is also mediated by treatment with retinoic acid, again in the absence of DNA demethylation. Thus, histone acetylation is sufficient to mediate the activation of Nestin transcription. This study proposed that the regulation of Nestin gene expression can be used as a model to study the epigenetic regulation of gene expression mediated by histone acetylation, but not by DNA demethylation.

Topics: Acetylation; Animals; Azacitidine; Carcinoma, Embryonal; Cell Line; Chromatin; Chromatin Immunoprecipitation; Decitabine; DNA (Cytosine-5-)-Methyltransferase 1; DNA (Cytosine-5-)-Methyltransferases; DNA Methylation; Embryonic Stem Cells; Enhancer Elements, Genetic; Epigenesis, Genetic; Histones; Hydroxamic Acids; Intermediate Filament Proteins; Introns; Mice; Nerve Tissue Proteins; Nestin; Promoter Regions, Genetic; Transcription, Genetic; Tretinoin

Regulation of tissue homeostasis is crucial to disease prevention; cell division, cell cycle arrest, differentiation and apoptosis have to be tightly controlled in order to maintain this homeostasis. Retinoic acid (RA) and the histone deacetylase inhibitors (HDACIs) have profound effects on these processes and thus may be critical regulators of homeostasis. Consequently, RA and/or histone deacetylase inhibitors are currently being tested in clinical trials for a variety of cancers. Unfortunately, little is known of the overall affect of these compounds on cellular gene expression. Therefore, we decided to compare the effects of all-trans retinoic acid (ATRA) and a particular HDACI-Trichostatin A (TSA)-on an embryonal carcinoma (EC) cell line (F9) using gene chip analysis. We have focused particular attention on those genes that may be differentially affected by these compounds. Within the parameters established for this study, only 116 of the 12,488 genes examined were similarly regulated by ATRA and TSA: 75 positively and 41 negatively. An additional 70 genes were affected by only one of the compounds and 19 genes were actually inversely regulated. The gene set inversely regulated by ATRA and TSA includes several important patterning genes as well as the crucial tumor suppressor/promoter, transforming growth factor beta 1 (TGFbeta1). Promoter analysis suggests a motif that may regulate one set of these genes. This study provides the first comprehensive comparison of global gene expression on EC cells as affected by ATRA and a HDAC inhibitor (TSA); reveals new targets for ATRA and HDAC inhibitors; identifies a new regulatory motif; demonstrates that ATRA and HDAC inhibitors do not always act synergistically on gene expression; and examines particular questions regarding their concurrent clinical application.

Topics: Antineoplastic Agents; Biomarkers, Tumor; Carcinoma, Embryonal; Embryonal Carcinoma Stem Cells; Enzyme Inhibitors; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Neoplastic Stem Cells; Oligonucleotide Array Sequence Analysis; Tretinoin; Tumor Cells, Cultured

Histone acetylation is thought to have a role in transcription. To gain insight into the role of histone acetylation in retinoid-dependent transcription, we studied the effects of trichostatin A (TSA), a specific inhibitor of histone deacetylase, on P19 embryonal carcinoma cells. We show that coaddition of TSA and retinoic acid (RA) markedly enhances neuronal differentiation in these cells, although TSA alone does not induce differentiation but causes extensive apoptosis. Consistent with the cooperative effect of TSA and RA, coaddition of the two agents synergistically enhanced transcription from stably integrated RA-responsive promoters. The transcriptional synergy by TSA and RA required the RA-responsive element and a functional retinoid X receptor (RXR)/retinoic acid receptor (RAR) heterodimer, both obligatory for RA-dependent transcription. Furthermore, TSA led to promoter activation by an RXR-selective ligand that was otherwise inactive in transcription. In addition, TSA enhanced transcription from a minimum basal promoter, independently of the RA-responsive element. Finally, we show that TSA alone or in combination with RA increases in vivo endonuclease sensitivity within the RA-responsive promoter, suggesting that TSA treatment might alter a local chromatin environment to enhance RXR/RAR heterodimer action. Thus, these results indicate that histone acetylation influences activity of the heterodimer, which is in line with the observed interaction between the RXR/RAR heterodimer and a histone acetylase presented elsewhere.

Topics: Animals; Apoptosis; Carcinoma, Embryonal; Cell Cycle; Cell Differentiation; Dimerization; Enzyme Inhibitors; Genes, Reporter; Histone Deacetylase Inhibitors; Hydroxamic Acids; Kinetics; Luciferases; Mice; Neurons; Promoter Regions, Genetic; Receptors, Retinoic Acid; Recombinant Proteins; Retinoid X Receptors; Time Factors; Transcription Factors; Transfection; Tretinoin; Tumor Cells, Cultured