Cloning and analysis of MART-1/Melan-A human melanoma antigen promoter regions.

The MART-1/Melan-A human melanoma tumor antigen can be recognized by T lymphocytes and appears to be involved in tumor regression. To study the transcriptional regulation of this important gene, the 5' untranslated (UT) region of the MART-1/Melan-A gene was cloned and sequenced. Human melanoma cell lines were screened for MART-1/Melan-A mRNA expression. Primer extension and northern analysis were performed to confirm the mRNA size and start site. Several overlapping fragments of 5'UT were isolated from genomic DNA by polymerase chain reaction (PCR) from the previously described sequence for an additional 700 bp upstream. The fragments isolated (ranging from 838 bp to 160 bp in length) were used to drive luciferase reporter gene expression in melanoma and non-melanoma cell lines. Tissue-specific promoter activity was found in a 233-bp fragment of 5' UT with an average index of induction of 35 fold. The 233-bp MART-1/Melan-A promoter does not appear to have cytokine (IL-2, IL-4, IL-7, GM-CSF, TNF-alpha or IFN-gamma) responsive elements when studied in transient transfection assays.