The ubiquitin ligase gp78 promotes sarcoma metastasis by targeting KAI1 for degradation

Abstract

Metastasis is the primary cause of mortality from cancer, but the mechanisms leading to metastasis are poorly understood. In particular, relatively little is known about metastasis in cancers of mesenchymal origins, which are known as sarcomas. Approximately ten proteins have been characterized as 'metastasis suppressors', but how these proteins function and are regulated is, in general, not well understood. Gp78 (also known as AMFR or RNF45) is a RING finger E3 ubiquitin ligase that is integral to the endoplasmic reticulum (ER) and involved in ER-associated degradation (ERAD) of diverse substrates1,2,3,4,5,6. Here we report that expression of gp78 has a causal role in the metastasis of an aggressive human sarcoma and that this prometastatic activity requires the E3 activity of gp78. Further, gp78 associates with and targets the transmembrane metastasis suppressor, KAI1 (also known as CD82), for degradation. Suppression of gp78 increases KAI1 abundance and reduces the metastatic potential of tumor cells, an effect that is largely blocked by concomitant suppression of KAI1. An inverse relationship between these proteins was confirmed in a human sarcoma tissue microarray. Whereas most previous efforts have focused on genetic mechanisms for the loss of metastasis suppressor genes, our results provide new evidence for post-translational downregulation of a metastasis suppressor by its ubiquitin ligase, resulting in abrogation of its metastasis-suppressing effects.

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Figure 1: The ubiquitin-protein ligase activity of gp78 is required for metastasis of HT1080 sarcoma cells.
Figure 2: Suppression of gp78 increases tumor cell apoptosis during early experimental metastasis.
Figure 3: The metastasis suppressor KAI1 is a substrate of gp78.
Figure 4: Suppression of gp78 resulted in accumulation of KAI1 and reduced metastatic potential.

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Acknowledgements

We thank members of the Laboratory of Protein Dynamics and Signaling for helpful discussions, and we thank S.H. Hong for assistance in the caspase activation and imaging assays. Some experiments were performed with the help A.H. Chan and E. Sum. We also thank the following people for materials and reagents: K. Baird and P. Meltzer (NCI) for the human sarcoma tissue microarray, J.C. Barrett (Novartis) and M. Custer (NCI) for the KAI1 cDNA and antibody, P. Cresswell (Yale University) for the MaP.CD82 antibody, R. Gemmill (University of Colorado) for the Trc8 cDNA, M. Hochstrasser and L. Wang (Yale University) for the TEB4 cDNA, R. Kopito (Stanford University) for GFPu, and G. Melillo (NCI) for 786-O cells. We are indebted to members of the Tissue Array Research Program and to S. Lockett and members of the Image Analysis Laboratory for their assistance. This research is supported by the Intramural Research Program of the US National Institutes of Health, National Cancer Institute, Center for Cancer Research. Y.C.T. and A.M.W. dedicate this study to a wonderful mentor and colleague, C. Pickart, who was an inspiration for this research.

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Y.C.T. designed and carried out experiments, interpreted data and wrote the paper. A.M. performed animal studies. J.M.M., Z.K and B.C. developed reagents and performed experiments. M.Z. and T.V. carried out mass spectrometric analysis. S.M.H. helped to analyze the histopathology and tissue microarray data. L.J.H. helped to initiate this project and contributed to experimental design. C.K. contributed to directing the project, oversaw animal studies and wrote the paper. A.M.W. initiated and directed the project and wrote the paper.

Corresponding authors

Correspondence to Chand Khanna or Allan M Weissman.

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Tsai, Y., Mendoza, A., Mariano, J. et al. The ubiquitin ligase gp78 promotes sarcoma metastasis by targeting KAI1 for degradation. Nat Med 13, 1504–1509 (2007). https://doi.org/10.1038/nm1686

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