The calcium-activated chloride channel anoctamin 1 (ANO1) is located within the 11q13 amplicon, one of the most frequently amplified chromosomal regions in human cancer, but its functional role in tumorigenesis has remained unclear. The 11q13 region is amplified in ∼15% of breast cancers. Whether ANO1 is amplified in breast tumors, the extent to which gene amplification contributes to ANO1 overexpression, and whether overexpression of ANO1 is important for tumor maintenance have remained unknown. We have found that ANO1 is amplified and highly expressed in breast cancer cell lines and primary tumors. Amplification of ANO1 correlated with disease grade and poor prognosis. Knockdown of ANO1 in ANO1-amplified breast cancer cell lines and other cancers bearing 11q13 amplification inhibited proliferation, induced apoptosis, and reduced tumor growth in established cancer xenografts. Moreover, ANO1 chloride channel activity was important for cell viability. Mechanistically, ANO1 knockdown or pharmacological inhibition of its chloride-channel activity reduced EGF receptor (EGFR) and calmodulin-dependent protein kinase II (CAMKII) signaling, which subsequently attenuated AKT, v-src sarcoma viral oncogene homolog (SRC), and extracellular signal-regulated kinase (ERK) activation in vitro and in vivo. Our results highlight the involvement of the ANO1 chloride channel in tumor progression and provide insights into oncogenic signaling in human cancers with 11q13 amplification, thereby establishing ANO1 as a promising target for therapy in these highly prevalent tumor types.ion channel | CaCCinh-A01 | TMEM16A | HNSCC | ESCC

The Zap1p transcription factor senses cellular zinc status and increases expression of its target genes in response to zinc deficiency. Previously known Zap1p-regulated genes encode the Zrt1p, Zrt2p, and Zrt3p zinc transporter genes and Zap1p itself. To allow the characterization of additional genes in yeast important for zinc homeostasis, a systematic study of gene expression on the genome-wide scale was used to identify other Zap1p target genes. Using a combination of DNA microarrays and a computer-assisted analysis of shared motifs in the promoters of similarly regulated genes, we identified 46 genes that are potentially regulated by Zap1p. Zap1p-regulated expression of seven of these newly identified target genes was confirmed independently by using lacZ reporter fusions, suggesting that many of the remaining candidate genes are also Zap1p targets. Our studies demonstrate the efficacy of this combined approach to define the regulon of a specific eukaryotic transcription factor.

All cells regulate their intracellular zinc levels. In yeast, zinc uptake is mediated by Zrt1p and Zrt2p, which belong to the ZIP family of metal transporters. Under zinc limitation, ZRT1 and ZRT2 transcription is induced by the Zap1p transcriptional activator. We describe here a new component of zinc homeostasis, vacuolar zinc storage, that is also regulated by Zap1p. Zinc-replete cells accumulate zinc in the vacuole via the Zrc1p and Cot1p transporters. Our results indicate that another zinc transporter, Zrt3p, mobilizes this stored zinc in zinc-limited cells. ZRT3 is a Zap1p-regulated gene whose transcription increases in low zinc. Zrt3p is also a member of the ZIP family and it localizes to the vacuolar membrane. The effects of ZRT3 mutation and overexpression on cell growth, cellular zinc accumulation and intracellular labile zinc pools are all consistent with its proposed role. Furthermore, we demonstrate that zrt3 mutants inef®-ciently mobilize stored zinc to offset de®ciency. Thus, our studies de®ne a system of zinc in¯ux and ef¯ux transporters in the vacuole that play important roles in zinc homeostasis.