Smad4 regulates TGF-β1-mediated hedgehog activation to promote epithelial-to-mesenchymal transition in pancreatic cancer cells by suppressing Gli1 activity

  • Comput Struct Biotechnol J. 2024 Mar 13:23:1189-1200. doi: 10.1016/j.csbj.2024.03.010.
Hangcheng Guo  1  2 ,  Zujian Hu  1 ,  Xuejia Yang  1 ,  Ziwei Yuan  1 ,  Mengsi Wang  1 ,  Chaoyue Chen  1 ,  Lili Xie  1 ,  Yuanyuan Gao  1 ,  Wangjian Li  3 ,  Yongheng Bai  1  4 ,  Chunjing Lin  5  6
Affiliations
  • 1. Zhejiang Key Laboratory of Intelligent Cancer Biomarker Discovery and Translation, First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325035, China.
  • 2. Sichuan Mianyang 404 Hospital, Mianyang 621000, China.
  • 3. Department of Urology, The Central Hospital Affiliated to Shaoxing University, Shaoxing 312030, China.
  • 4. National Key Clinical Specialty (General Surgery), The First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, China.
  • 5. Department of Gastroenterology, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, China.
  • 6. Medicine and Health Care Center, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, China.
PMID: 38525105 DOI: 10.1016/j.csbj.2024.03.010
Abstract

Pancreatic Cancer (PC) is an aggressive and metastatic gastrointestinal tumor with a poor prognosis. Persistent activation of the TGF-β/Smad signaling induces PC cell (PCC) invasion and infiltration via epithelial-to-mesenchymal transition (EMT). Hedgehog signaling is a crucial pathway for the development of PC via the transcription factors Gli1/2/3. This study aimed to investigate the underlying molecular mechanisms of action of Hedgehog activation in TGF-β1-triggered EMT in PCCs (PANC-1 and BxPc-3). In addition, overexpression and shRNA techniques were used to evaluate the role of SMAD4 in TGF-β1-treated PCCs. Our data showed that TGF-β1 promoted PCC invasion and infiltration via SMAD2/3-dependent EMT. Hedgehog-Gli signaling axis in PCCs was activated upon TGF-β1 stimulation. Inhibition of Hedgehog with cyclopamine effectively antagonized TGF-β1-induced EMT, thereby suggesting that the Hedgehog signaling may act as a downstream cascade signaling of TGF-β1. As a key protein that assists the nuclear translocation of SMAD2/3, SMAD4 was highly expressed in PANC-1 cells, but not in BxPc-3 cells. Conversely, Gli1 expression was low in PANC-1 cells, but high in BxPc-3 cells. Furthermore, knockdown of SMAD4 in PANC-1 cells by shRNA inhibited TGF-β1-mediated EMT and Collagen deposition. Overexpression of SMAD4 did not affect TGF-β1-mediated EMT due to the lack of significant increase in nuclear expression of SMAD4. Importantly, Gli1 activity was upregulated by SMAD4 knockdown in PANC-1 cells and downregulated by SMAD4 overexpression in BxPc-3 cells, indicating that Gli1 may be a negative target protein downstream of SMAD4. Thus, SMAD4 regulates TGF-β1-mediated Hedgehog activation to promote EMT in PCCs by suppressing Gli1 activity.

Keywords
Epithelial-mesenchymal transition (EMT); Gli1; Hedgehog signaling; Pancreatic cancer; Smad4; TGF-β1.
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