Abstract
Melanoma is the most aggressive skin malignancy with increasing incidence worldwide. Pannexin1 (PANX1), a member of the pannexin family of channel-forming glycoproteins, regulates cellular processes in melanoma cells including proliferation, migration, and invasion/metastasis. However, the mechanisms responsible for coordinating and regulating PANX1 function remain unclear. Here, we demonstrated a direct interaction between the C-terminal region of PANX1 and the N-terminal portion of β-catenin, a key transcription factor in the Wnt pathway. At the protein level, β-catenin was significantly decreased when PANX1 was either knocked down or inhibited by two PANX1 blockers, Probenecid and Spironolactone. Immunofluorescence imaging showed a disrupted pattern of β-catenin localization at the cell membrane in PANX1-deficient cells, and transcription of several Wnt target genes, including MITF, was suppressed. In addition, a mitochondrial stress test revealed that the metabolism of PANX1-deficient cells was impaired, indicating a role for PANX1 in the regulation of the melanoma cell metabolic profile. Taken together, our data show that PANX1 directly interacts with β-catenin to modulate growth and metabolism in melanoma cells. These findings provide mechanistic insight into PANX1-mediated melanoma progression and may be applicable to other contexts where PANX1 and β-catenin interact as a potential new component of the Wnt signaling pathway.
Highlights
Endoplasmic reticulum [6, 7]
To determine whether there is an interaction between PANX1 and β-catenin in melanoma cells, we first analyzed a panel of melanoma biopsies from the Cancer Genome Atlas (TCGA) and found a modest, yet significant, correlation between PANX1 and β-catenin mRNA (CTNNB1) in 471 patients with malignant melanoma (Fig. 1A, left panel)
Given the similarity of connexin and pannexin structure, we investigated whether calmodulin interacts with PANX1 in melanoma cells
Summary
Endoplasmic reticulum [6, 7]. PANX1 plays an important role in normal physiological processes, including skin development and wound healing as well as in pathophysiological conditions and metabolic disorders, such as Alzheimer’s disease, diabetes, inflammation, and cancer [8, 9]. Our recent findings indicate that knocking down (KD) PANX1 with shRNA in aggressive BL6 mouse melanoma cells, as well as in human melanoma cell lines, reduces the abundance of β-catenin [11, 12], a key transcription factor in the Wnt signaling pathway implicated in melanoma tumorigenesis [13]. Wnt/β-catenin signaling regulates proliferation, migration, and invasion of melanoma cells [14,15,16]. We observed that PANX1 is expressed in all cell lines tested but most abundant in 131/4-5B1 cells at both the protein and mRNA levels (Fig. 1, B and C).
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