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A CREB3-regulated ER–Golgi trafficking signature promotes metastatic progression in breast cancer

Abstract

In order to better understand the process of breast cancer metastasis, we have generated a mammary epithelial progression series of increasingly aggressive cell lines that metastasize to lung. Here we demonstrate that upregulation of an endoplasmic reticulum (ER) to Golgi trafficking gene signature in metastatic cells enhances transport kinetics, which promotes malignant progression. We observe increased ER–Golgi trafficking, an altered secretome and sensitivity to the retrograde transport inhibitor brefeldin A (BFA) in cells that metastasize to lung. CREB3 was identified as a transcriptional regulator of upregulated ER–Golgi trafficking genes ARF4, COPB1, and USO1, and silencing of these genes attenuated the metastatic phenotype in vitro and lung colonization in vivo. Furthermore, high trafficking gene expression significantly correlated with increased risk of distant metastasis and reduced relapse-free and overall survival in breast cancer patients, suggesting that modulation of ER–Golgi trafficking plays an important role in metastatic progression.

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Fig. 1: Characterization of in vivo selected mammary epithelial cells.
Fig. 2: Identification of a metastasis-associated gene signature through the in vivo selection of mammary epithelial cells.
Fig. 3: Upregulation of ER–Golgi trafficking and sensitization to brefeldin A in L1P and L2P cells.
Fig. 4: CREB3-mediated regulation of ER–Golgi trafficking genes.
Fig. 5: ER–Golgi trafficking genes promote cell adhesion, migration, and invasion.
Fig. 6: ER–Golgi trafficking genes promote metastatic progression.
Fig. 7: ER–Golgi trafficking genes associated with metastatic risk and survival in human breast cancer.

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Acknowledgements

This work was supported by Grants CA055536 and CA154663 from the National Cancer Institute to PHH. BVH was supported by funding from the Abney Foundation. This study used the services of the MUSC Proteogenomics Facility supported by NIH/NIGMS (GM103342 and GM103499), the Cell & Molecular Imaging Shared Resource of MUSC supported by P30 CA138313, the MUSC Center for Oral Health Research (COHR), which is partially supported by P30 GM103331, the MUSC Flow Cytometry Facility supported by NIH/NIGMS GM103342, the Biorepository & Tissue Analysis Shared Resource, Hollings Cancer Center, MUSC, and the Gene Targeting and Knockout Shared Resource at MUSC. We thank Annamarie Dalton for assisting us with xenograft experiments and also thank members of the Howe laboratory for providing helpful feedback during the preparation of this manuscript.

Author contributions

BVH designed experiments, performed research and analyzed the data. LAL generated in vivo selected cell lines, performed research, and analyzed data. SG performed research and analyzed data. ME-S performed xenograft experiments and analyzed data. PHH directed the research and analyzed data. BVH and PHH wrote the manuscript.

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Correspondence to Philip H. Howe.

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Howley, B.V., Link, L.A., Grelet, S. et al. A CREB3-regulated ER–Golgi trafficking signature promotes metastatic progression in breast cancer. Oncogene 37, 1308–1325 (2018). https://doi.org/10.1038/s41388-017-0023-0

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