RESUMO
Alpha-defensin-1 gene expression in promyelocytic HL-60 cells is ('delayed-late' > or =1-2 days) activated by retinoic acid (RA), lipopolysaccharide, tumor necrosis factor-alpha and elevated levels of cAMP. Using stably integrated reporter constructs, we show that this activation is directed through a proximal and distal element within a minimal (-83/+82) def1 promoter, and is mediated by phosphorylation of the associated factors, PU.1 and D1BP, in an inducer-dependent manner. Whereas binding of PU.1 to the proximal element confers cell specificity and relays the effects of most inducers, the selectively enhancing capacity of the distal element for RA- and cAMP-dependent activation is uniquely correlated with D1BP-binding. We propose that D1BP and PU.1 are the end-points of separate pathways.
Assuntos
Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regiões Promotoras Genéticas , Sequências Reguladoras de Ácido Nucleico , Retinoides/farmacologia , Transcrição Gênica/efeitos dos fármacos , alfa-Defensinas/genética , Antineoplásicos/farmacologia , Sítios de Ligação , AMP Cíclico/farmacologia , Ensaio de Desvio de Mobilidade Eletroforética , Genes Reporter , Células HL-60 , Células HeLa , Humanos , Lipopolissacarídeos/farmacologia , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas/metabolismo , Deleção de Sequência , Transativadores/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , alfa-Defensinas/metabolismoRESUMO
The elimination of malignant tumors by intratumoral virus replication is a challenging therapeutic approach but is critically dependent on the speed and efficacy of intratumoral virus spread. The expression of oncolytic transgenes in the context of a replicating virus may help to enhance the therapeutic potency of this strategy. We have established a human hepatocarcinoma-derived cell line (Huh7-E1) which stably expresses adenoviral E1-genes. Tumors derived from these cells support replication of E1-deficient adenoviruses in SCID mice. This model can be used to evaluate E1-negative viruses encoding reporter genes or oncolytic transgenes in a replicating context. Most oncolytic viruses for human use could then be re-engineered as E1-postive viruses. Moreover, Huh7-E1 tumors release human alpha-1-antitrypsin (hAAT), which allows the monitoring of occult growing tumors (i.e. liver, peritoneum) by measuring serum hAAT levels.