Caspase-8 gene expression in neuroblastoma.
Neuroblastoma (NB) is a solid tumor of infancy that presents a high rate of spontaneous regression, a phenomenon that likely reflects the activation of an apoptotic/differentiation program. Indeed, the level of expression of molecules involved in the regulation of apoptosis, such as p73 or survivin, is a prognostic factor in NB patients. The caspase-8 gene (CASP8) encodes a key enzyme at the top of the apoptotic cascade. Although methylation of a putative regulatory region of the CASP8 gene reportedly inhibits its transcription in some MYCN-amplified NB, our results indicate that the transcriptional inactivation of caspase-8 occurs in a subset of primary NB independently of MYCN amplification or CpG methylation. In addition, the apoptotic agent fenretinide (4HPR) and interferon-gamma (IFN-gamma) induce caspase-8 expression without modifying the methylation status of this gene. Nevertheless, the methylation level of CASP8 intragenic and promoter regions is higher in MYCN-amplified tumors as compared to nonamplified samples. This phenomenon might reflect the existence of distinct DNA methylation errors in MYCN-amplified and MYCN-single copy tumors. To gain information on the mechanisms that regulate the expression of this crucial apoptotic gene, we searched for potential CASP8 regulatory regions and cloned a DNA element at the 5' terminus of this gene that functionally acts as a promoter only in NB cell lines that express caspase-8. The retinoic acid analogue 4HPR, IFN-gamma, and the demethylating agent 5-aza-cytidine activate this promoter in NB cells that lack endogenous caspase-8, indicating that this element may regulate both constitutive and inducible CASP8 expression. These results indicate also that demethylation of the cellular genome may upregulate CASP8 through the action of trans-acting factors. Our results provide new insights to the regulation of CASP8, a gene with an essential role in a variety of physiologic and pathologic conditions.
Casciano
Ida
I
Laboratory of Tumor Genetics, Istituto Nazionale per la Ricerca sul Cancro, Largo Rosanna Benzi 10, 16132 Genova, Italy.
Banelli
Barbara
B
Croce
Michela
M
De Ambrosis
Alessandro
A
di Vinci
Angela
A
Gelvi
Ilaria
I
Pagnan
Gabriella
G
Brignole
Chiara
C
Allemanni
Giorgio
G
Ferrini
Silvano
S
Ponzoni
Mirco
M
Romani
Massimo
M
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis; D001374:Azacitidine / Q000494:pharmacology; D053181:Caspase 8; D020169:Caspases / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D018899:CpG Islands; D004247:DNA / Q000378:metabolism; D019175:DNA Methylation; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology*; D011401:Promoter Regions, Genetic; D012313:RNA / Q000378:metabolism; D014212:Tretinoin / Q000494:pharmacology
D000964:Antimetabolites, Antineoplastic; D014212:Tretinoin; D012313:RNA; D007371:Interferon-gamma; D004247:DNA; C505530:CASP8 protein, human; D053181:Caspase 8; D020169:Caspases; D001374:Azacitidine