Transcriptional regulation of the alpha 4 integrin subunit gene in the metastatic spread of uveal melanoma.
Recently, expression of the alpha 4 integrin subunit has been shown to be inversely correlated with the invasive potential of B16 mouse epidermal melanoma. The purpose of this study was to establish whether expression of the human alpha 4 integrin subunit gene might be similarly regulated in human uveal melanoma which has varying degrees of invasiveness, and whether such modifications are determined by alterations in the transcriptional activity directed by the alpha 4 gene promoter. Two metastatic variants (MH5 and MH10) derived from a human uveal melanoma (SP6.5) were used. Expression studies were performed by transiently transfecting each of these cell lines with recombinant plasmids bearing various lengths of the alpha 4 promoter fused to the CAT reporter gene, and were further validated by Northern blot analyses of the alpha 4 transcript. Both transient transfection and mRNA analyses provided evidence that the transcriptional activity directed by the alpha 4 promoter sequences extending up to position -76 and -120 was indeed inversely correlated to the potential of uveal melanoma to yield metastasis. Experiments in electrophoretic mobility shift assay (EMSA) demonstrated that binding of the nuclear proteins that likely account for transcription of the alpha 4 gene to alpha 4.1 (namely Bp1, Bp2, Bp4, and Bp5) was dramatically reduced in uveal melanoma, but not in normal uveal melanocytes. These results highlight the fundamental function the alpha 4 integrin subunit may play in the ability of tumor cells to evade the primary tumor and form metastasis.