Loss of the metastasis suppressor gene, KiSS-1 has been strongly correlated to the progression of metastases in numerous types of cancers. The mechanism through which KiSS-1 is lost during metastasis, however, is still not completely known. Previous studies have shown that genetic material on human chromosome 6q16.3-q23 is essential for KiSS-1 expression in normal tissues. Additionally, microcell-mediated transfer of this chromosome in cancerous tissue results in rescued expression of KiSS-1 and reduced metastatic phenotype. Here, we show that loss of Sp1-coactivator protein DRIP-130, which is encoded by human chromosome 6q16.3-q23, results in reduced KiSS-1 promoter activation in highly malignant melanoma cells. Co-expression of Sp1 and DRIP-130 not only rescues KiSS-1 expression, but also induces an inhibition of the invasive and migratory behavior in highly metastatic melanoma cells, similar to the overexpression of KiSS-1 metastasis suppressor gene in those cells. Furthermore, we demonstrate that KiSS-1 expression is regulated by Sp1 elements within the first 100-bp region of the KiSS-1 promoter and that targeted deletion of a single GC-rich region spanning -93 to -58 interrupts Sp1- and DRIP-130-modulated transcriptional control of KiSS-1 expression. Our results thus suggest that DRIP-130 is a key regulator in KiSS-1 transactivation in normal tissue, and that the loss of DRIP-130 expression, as a result of the gross loss of human chromosome 6q16.3-q23, provokes increased tumor metastasis.