Abstract
Flotillin-1 contributes to invasion and metastasis in triple negative breast cancer (TNBC) and is modified post-translationally through palmitoylation. Palmitoylation, the process of conjugating palmitoyl-CoA to proteins, plays an essential role in protein stability and trafficking. Thus far, there has not been any investigation into the role of flotillin-1 palmitoylation in the context of metastasis in vivo. To address the role of flotillin-1 palmitoylation in metastasis, MDA-MB-231 cells expressing palmitoylation defective flotillin-1 constructs were used as models. Compared to flotillin-1 WT expressing tumors, flotillin-1 palmitoylation defective displayed abrogated tumor progression and lung metastasis in vivo in both spontaneous and experimental models. Further mechanistic investigation led to the identification of zDHHC5 as the main palmitoyl acyltransferase responsible for palmitoylating endogenous flotillin-1. Modulation of flotillin-1 palmitoylation status through mutagenesis, zDHHC5 silencing, and 2-bromopalmitate inhibition all resulted in the proteasomal degradation of flotillin-1 protein. To assess if flotillin-1 palmitoylation can be inhibited for potential clinical relevance, we designed a competitive peptide fused to a cell penetrating peptide sequence, which displayed efficacy in blocking flotillin-1 palmitoylation in vitro without altering palmitoylation of other zDHHC5 substrates, highlighting its specificity. Additionally, TNBC xenograft tumor models expressing a doxycycline inducible flotillin-1 palmitoylation inhibiting peptide displayed attenuated tumor growth and lung metastasis. Collectively, these results reveal a novel palmitoylation dependent mechanism which is essential for the stability of flotillin-1 protein. More specifically, disruption of flotillin-1 palmitoylation through mutagenesis or competitive peptide promoted flotillin-1 protein degradation, subsequently impeding its tumor promoting and metastasis-inducing effects in TNBC tumor models.
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Acknowledgements
We would like to thank Dr. Brittany Harlow for offering her support during data collection as well as the UT Austin flow cytometry core for their support with the FACS experimentation.
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University of Texas Vice President of Research Creative Grant.
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BM designed and performed all experiments unless stated otherwise and wrote the manuscript, CW assisted in animal experiments and edited the manuscript, AB edited the manuscript and provided supervision, CJ edited the manuscript and provided supervision, LD edited the manuscript, provided supervision, and acquired funding for the study.
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McClellan, B., Wilson, C.N., Brenner, A.J. et al. Flotillin-1 palmitoylation is essential for its stability and subsequent tumor promoting capabilities. Oncogene 43, 1063–1074 (2024). https://doi.org/10.1038/s41388-024-02946-0
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DOI: https://doi.org/10.1038/s41388-024-02946-0