TGF-\u03b2-mediated epithelial\u2013mesenchymal transition and tumor-promoting effects in CMT64 cells are reflected in the transcriptomic signature of human lung adenocarcinoma

Naoya Miyashita,Patrick Micke,Hirokazu Urushiyama,Takahide Nagase,Carina Strell,Kensuke Fukuda,Akira Saito,Masafumi Horie,Hans Brunnström,Takayoshi Enokido

doi:10.1038/s41598-021-01799-x

Abstract

Epithelial–mesenchymal transition (EMT) is a cellular process during which epithelial cells acquire mesenchymal phenotypes. Cancer cells undergo EMT to acquire malignant features and TGF-β is a key regulator of EMT. Here, we demonstrate for the first time that TGF-β could elicit EMT in a mouse lung adenocarcinoma cell line. TGF-β signaling activation led to cell morphological changes corresponding to EMT and enhanced the expression of mesenchymal markers and EMT-associated transcription factors in CMT64 lung cancer cells. RNA-sequencing analyses revealed that TGF-β increases expression of Tead transcription factors and an array of Tead2 target genes. TGF-β stimulation also resulted in alternative splicing of several genes including Cd44, tight junction protein 1 (Tjp1), and Cortactin (Cttn). In parallel with EMT, TGF-β enhanced cell growth of CMT64 cells and promoted tumor formation in a syngeneic transplantation model. Of clinical importance, the expression of TGF-β-induced genes identified in CMT64 cells correlated with EMT gene signatures in human lung adenocarcinoma tissue samples. Furthermore, TGF-β-induced gene enrichment was related to poor prognosis, underscoring the tumor-promoting role of TGF-β signaling in lung adenocarcinoma. Our cellular and syngeneic transplantation model would provide a simple and useful experimental tool to study the significance of TGF-β signaling and EMT.

Highlights

Epithelial–mesenchymal transition (EMT) is a cellular process during which epithelial cells acquire mesenchymal phenotypes
These results indicated that the intracellular Smad pathway is activated in CMT64 cells in response to TGF-β
We revealed that TGF-β elicits EMT in CMT64 mouse lung adenocarcinoma cells and TGF-β-treated CMT64 cells form larger tumors in vivo

Summary

Introduction

Epithelial–mesenchymal transition (EMT) is a cellular process during which epithelial cells acquire mesenchymal phenotypes. Most studies have utilized xenograft models where human cancer cells are transplanted into immunodeficient mice This implies a major limitation, considering the importance of tumor immune response. Transplantation of Lewis lung cancer cells in syngeneic immunocompetent mice has frequently been used in previous studies[4] This cell type is highly aggressive and lacks epithelial features, questioning its relevance as an Scientific Reports | (2021) 11:22380. A549 human lung adenocarcinoma cell line was most frequently used to study EMT. These cell culture studies have demonstrated that EMT endows cells with invasive phenotypes and molecular processes related to EMT have been characterized in detail[7]. Based on the limitation of this model, the pathological relevance and clinical significance of EMT in lung cancer remains inconclusive

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Conclusion