CD44 splice isoform switching in human and mouse epithelium is essential for epithelial-mesenchymal transition and breast cancer progression
Rhonda L. Brown, … , Jing Yang, Chonghui Cheng
Rhonda L. Brown, … , Jing Yang, Chonghui Cheng
Published February 14, 2011
Citation Information: J Clin Invest. 2011;121(3):1064-1074. https://doi.org/10.1172/JCI44540.
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Research Article Oncology

CD44 splice isoform switching in human and mouse epithelium is essential for epithelial-mesenchymal transition and breast cancer progression

Abstract

Epithelial-mesenchymal transition (EMT) is a tightly regulated process that is critical for embryogenesis but is abnormally activated during cancer metastasis and recurrence. Here we show that a switch in CD44 alternative splicing is required for EMT. Using both in vitro and in vivo systems, we have demonstrated a shift in CD44 expression from variant isoforms (CD44v) to the standard isoform (CD44s) during EMT. This isoform switch to CD44s was essential for cells to undergo EMT and was required for the formation of breast tumors that display EMT characteristics in mice. Mechanistically, the splicing factor epithelial splicing regulatory protein 1 (ESRP1) controlled the CD44 isoform switch and was critical for regulating the EMT phenotype. Additionally, the CD44s isoform activated Akt signaling, providing a mechanistic link to a key pathway that drives EMT. Finally, CD44s expression was upregulated in high-grade human breast tumors and was correlated with the level of the mesenchymal marker N-cadherin in these tumors. Together, our data suggest that regulation of CD44 alternative splicing causally contributes to EMT and breast cancer progression.

Authors

Rhonda L. Brown, Lauren M. Reinke, Marin S. Damerow, Denise Perez, Lewis A. Chodosh, Jing Yang, Chonghui Cheng

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Figure 5

CD44s potentiates Akt activation and promotes cell survival in TGF-β–induced mesenchymal MCF10A cells.

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CD44s potentiates Akt activation and promotes cell survival in TGF-β–ind...
(A) Immunoblot analysis of activated Akt (pAkt) in epithelial MCF10A cells (10A) and TGF-β–induced mesenchymal MCF10A cells (10AM), indicating an increased level of Akt phosphorylation in mesenchymal MCF10A cells. Cells were starved for 24 hours and then stimulated with 2 μg/ml insulin. (B) qRT-PCR analysis of E-cadherin levels in TGF-β–treated mesenchymal MCF10A cells treated with DMSO (control) or the PI3K inhibitor LY-294002 (LY; 50 μM) for 20 hours. Error bars indicate SD; n = 3. (C) Immunoblot analysis of pAkt in TGF-β–treated MCF10A cells expressing vector (Ctrl), CD44s, or CD44v. (D) Left: Knockdown efficiency of CD44 shRNA in TGF-β–treated MCF10A cells that predominantly express CD44s. Right: Levels of pAkt after serum starvation for 24 hours, followed by insulin (1 μg/ml) stimulation for 30 minutes, showing that silencing CD44s impairs insulin-stimulated Akt activation. (E) Results of apoptosis assays showing that ectopic expression of CD44s, but not CD44v, inhibits apoptosis in TGF-β–induced mesenchymal MCF10A cells when treated with cisplatin (100 μM) or cultured in suspension. Error bars indicate SEM; n = 3. (F) Results of apoptosis assays showing that treatment with LY-294002, but not the MEK inhibitor U0126, causes an increase in apoptosis in CD44s-expressing TGF-β–induced mesenchymal MCF10A cells. Error bars indicate SEM; n = 4.