A Notch1–neuregulin1 autocrine signaling loop contributes to melanoma growth

Abstract

The Notch pathway is an evolutionary conserved signaling cascade that has an essential role in melanoblast and melanocyte stem cell homeostasis. Notch signaling is emerging as a key player in melanoma, the most deadly form of skin cancer. In melanoma, Notch1 is inappropriately reactivated and contributes to melanoma tumorigenicity. Here, we propose a novel mechanism by which Notch1 promotes the disease. We found that Notch1 directly regulates the transcription of neuregulin1 (NRG1) by binding to its promoter region. NRG1 is the ligand for ERBB3 and 4, members of the Epidermal Growth Factor family of receptors that are involved in the genesis and progression of a number of cancers. Notch1 and NRG1 expression are associated in melanoma and inhibition of NRG1 signaling leads to melanoma cell growth inhibition and tumor growth delay. Mechanistically, these effects are associated with the inhibition of the PI3Kinase/Akt signaling pathway and with the accumulation of p27Kip1. On the other end, addition of recombinant NRG1 can partially restore melanoma cell growth that is inhibited by Notch1 ablation. Taken together, our findings underline a new, previously undescribed autocrine signaling loop between Notch1 and NRG1 that controls melanoma growth and provide experimental evidence that the targeting of Notch and ERBB signaling may represent a novel potential therapeutic approach in melanoma

Noncanonical Activation of Notch1 Protein by Membrane Type 1 Matrix Metalloproteinase (MT1-MMP) Controls Melanoma Cell Proliferation

“…5B). Expression of active Notch1 conferred growth advantages as described previously (6), in both control and shMT1-MMP-expressing cells, suggesting that reconstitution of Notch1 signaling can, at least in part, restore cell growth that is inhibited by MT1-MMP knockdown. Conversely, WM115 cells expressing active MT1-MMP were transduced with a specific shRNA against Notch1 (Fig.…”

“…Notch1 is an essential embryonic stem cell factor that plays key roles in the maintenance of melanocyte/melanoma precursor cell homeostasis (4). We have previously demonstrated that although in mature melanocytes the levels of Notch1 are low or undetectable, melanomas re-express Notch1, and active Notch1 is required for their growth and survival (5,6). However, unlike T-cell lymphoblastic leukemia, melanomas do not present activating mutations in the Notch1 gene (7), suggesting that Notch1 activation occurs via other mechanisms.…”

“…The MT1-MMP lentiviral expression vectors were constructed by inserting the cDNA sequence corresponding to full-length (MT1-FL) and mutated (MT1-Mut) human MT1-MMP that were PCR-amplified from pSG-MT1-MMP (kindly provided by Dr. Motoharu Seiki, University of Tokyo, Tokyo, Japan) into the pLM-CMV-Ha-puro-PL3 lentiviral plasmid (20). The Notch1 NIC construct was previously described (6). Viral particles were produced in 293-FT (fast growing variant of the 293 cell line stably expressing SV40 large T antigen) cells using the FuGENE 6 reagent (Roche Applied Science, Mannheim, Germany) as per the manufacturer's instructions.…”

“…Modulates Melanoma Cell Growth-Previous work by us and others highlighted a key role of Notch1 in melanoma growth and survival (5)(6)(7)(8). Therefore, we wanted to test whether MT1-MMP could affect melanoma cell growth as a consequence of activating Notch1.…”

“…Lack of Notch1 in the melanocytic lineage results in premature hair graying in the mouse caused by loss of melanoblasts in the embryo and premature differentiation of melanocyte stem cells in the adult mouse (4). We have previously shown that melanomas re-express and activate Notch1 and rely on its activity for tumor growth and survival (5,6). In addition, other groups have demonstrated that Notch1 is sufficient to confer transforming properties to primary human melanocytes (7) and to convert primary melanoma cells from noninvasive to metastatic (8,9).…”