Cancer-associated fibroblasts (CAFs) play a crucial role in cancer progression, drug resistance and tumor recurrence. We have recently shown that the Notch pathway determines the tumor-regulatory role of experimentally created 'CAFs'. Here, we examined the status of Notch signaling in human melanoma-associated fibroblasts (MAFs) versus their normal counterparts and tested whether manipulation of the Notch pathway activity in MAFs alters their tumor-regulatory function. Using tissue microarrays, we found that MAFs exhibit decreased Notch pathway activity compared with normal fibroblasts in adjacent and non-adjacent skin. Consistently, MAFs isolated from human metastatic melanoma exhibited lower Notch activity than did normal human fibroblasts, demonstrating that Notch pathway activity is low in MAFs. We then investigated the effect of increasing Notch pathway activity in MAF on melanoma growth in co-cultures and in a mouse co-graft model. We found that activation of the Notch pathway in MAFs significantly restricted melanoma cell growth in vitro and suppressed melanoma skin growth and tumor angiogenesis in vivo. Our study demonstrates that the Notch signaling is inhibited in MAFs. Increase of Notch pathway activity can confer tumor-suppressive function on MAFs. Thus, targeting melanoma by activating Notch signaling in MAF may represent a novel therapeutic approach.