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GULP1 regulates the NRF2-KEAP1 signaling axis in urothelial carcinoma
Masamichi Hayashi1,2, Elisa Guida1, Yoshikuni Inokawa1,2
1Department of Otolaryngology-Head and Neck Surgery, Johns Hopkins University School of Medicine, Baltimore, MD 21231, USA.
Abstract:
Disruption of the KEAP1-NRF2 pathway results in the transactivation of NRF2 target genes, consequently inducing cell proliferation and other phenotypic changes in cancer cells. Here, we demonstrated that GULP1 was a KEAP1-binding protein that maintained actin cytoskeleton architecture and helped KEAP1 to sequester NRF2 in the cytoplasm. In urothelial carcinoma of the bladder (UCB), silencing of GULP1 facilitated the nuclear accumulation of NRF2, led to constitutive activation of NRF2 signaling, and conferred resistance to the platinum drug cisplatin. Knockdown of GULP1 in UCB cells promoted tumor cell proliferation in vitro and enhanced tumor growth in vivo. In primary UCB, GULP1 silencing was more prevalent in muscle-invasive UCB compared to nonmuscle-invasive UCB. GULP1 knockdown cells showed resistance to cisplatin treatment. In parallel with decreased GULP1 expression, we observed increased expression of NRF2, HMOX1, and other candidate antioxidant genes in cisplatin-resistant cells. Furthermore, low or no expression of GULP1 was observed in most cisplatin nonresponder cases. Silencing of GULP1 was associated with GULP1 promoter hypermethylation in cell lines and primary tumors, and a high frequency of GULP1 promoter methylation was observed in multiple sets of primary clinical UCB samples. Together, our findings demonstrate that GULP1 is a KEAP1-binding protein that regulates KEAP1-NRF2 signaling in UCB and that promoter hypermethylation of GULP1 is a potential mechanism of GULP1 silencing.
Insights
GULP1 protein loss in bladder cancer promotes tumor growth and cisplatin resistance by activating the KEAP1-NRF2 pathway. GULP1 silencing, often due to promoter hypermethylation, is linked to advanced disease and poor treatment response.
Area of Science:
- Oncology
- Molecular Biology
- Cell Biology
Background:
- The KEAP1-NRF2 pathway regulates cellular responses to stress and is frequently disrupted in cancer.
- NRF2 (Nuclear factor erythroid 2-related factor 2) target genes promote cell proliferation and survival.
- Urothelial carcinoma of the bladder (UCB) is a significant health concern with limited treatment options.
Purpose of the Study:
- To investigate the role of GULP1 (Gastrokine, Liver, and Intestinal Protein 1) in the KEAP1-NRF2 pathway in UCB.
- To determine the association between GULP1 expression, tumor progression, and response to cisplatin in UCB.
- To elucidate the mechanism underlying GULP1 silencing in UCB.
Main Methods:
- Investigated GULP1 as a KEAP1-binding protein.
- Assessed the impact of GULP1 knockdown on NRF2 localization and signaling in UCB cells.
- Evaluated tumor cell proliferation, growth, and cisplatin resistance in vitro and in vivo.
- Analyzed GULP1 expression and promoter methylation in clinical UCB samples.
Main Results:
- GULP1 binds to KEAP1 and sequesters NRF2 in the cytoplasm, maintaining actin cytoskeleton integrity.
- GULP1 silencing in UCB cells leads to NRF2 nuclear accumulation, constitutive NRF2 signaling, and cisplatin resistance.
- GULP1 knockdown promotes UCB cell proliferation and tumor growth.
- GULP1 silencing is more frequent in muscle-invasive UCB and associated with cisplatin resistance and poor response.
- GULP1 silencing correlates with GULP1 promoter hypermethylation in cell lines and primary UCB tumors.
Conclusions:
- GULP1 acts as a tumor suppressor in UCB by regulating the KEAP1-NRF2 pathway.
- GULP1 silencing, driven by promoter hypermethylation, contributes to UCB progression and chemoresistance.
- GULP1 expression status may serve as a predictive biomarker for cisplatin treatment response in UCB.
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