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SKP1 promotes YAP-mediated colorectal cancer stemness via suppressing RASSF1
Cong Tian1,2, Tingyuan Lang3, Jiangfeng Qiu4
1Department of Medical Oncology, Shanghai University of Medicine & Health Sciences Affiliated Sixth People's Hospital East Campus, No. 222 Huan Hu Xi San Road, Pudong New Area, Shanghai, 201306, People's Republic of China.
Background:
Cancer stem cells (CSCs) have been recognized as an important drug target, however, the underlying mechanisms have not been fully understood. SKP1 is a traditional drug target for cancer therapy, while, whether SKP1 promotes colorectal cancer (CRC) stem cells (CRC-SCs) and the underlying mechanisms have remained elusive.
Methods:
Human CRC cell lines and primary human CRC cells were used in this study. Gene manipulation was performed by lentivirus system. The mRNA and protein levels of target genes were examined by qRT-PCR and western blot. The sphere-forming and in vitro migration capacities were determined by sphere formation and transwell assay. The self-renewal was determined by limiting dilution assay. The tumorigenicity and metastasis of cancer cells were examined by xenograft model. The promoter activity was examined by luciferase reporter assay. Nuclear run-on and Chromatin immunoprecipitation-PCR (ChIP-PCR) assay were employed to examine the transcription and protein-DNA interaction. Co-immunoprecipitation assay was used to test protein-protein interaction. The relationship between gene expression and survival was analyzed by Kaplan-meier analysis. The correlation between two genes was analyzed by Spearman analysis. Data are represented as mean ± SD and the significance was determined by Student's t test.
Results:
SKP1 was upregulated in CRC-SCs and predicted poor prognosis of colon cancer patients. Overexpression of SKP1 promoted the stemness of CRC cells reflected by increased sphere-forming, migration and self-renewal capacities as well as the expression of CSCs markers. In contrast, SKP1 depletion produced the opposite effects. SKP1 strengthened YAP activity and knockdown of YAP abolished the effect of SKP1 on the stemness of CRC cells. SKP1 suppressed RASSF1 at both mRNA and protein level. Overexpression of RASSF1 abolished the effect of SKP1 on YAP activity and CRC stemness.
Conclusion:
Our results demonstrated that SKP1 suppresses RASSF1 at both mRNA and protein level, attenuates Hippo signaling, activates YAP, and thereby promoting the stemness of CRC cells.
Insights
SKP1 promotes colorectal cancer stemness by suppressing RASSF1 and activating YAP, highlighting SKP1 as a potential therapeutic target for colorectal cancer (CRC). This study elucidates the mechanisms driving CRC stem cell behavior.
Area of Science:
- Oncology
- Molecular Biology
- Cancer Stem Cell Research
Background:
- Cancer stem cells (CSCs) are crucial drug targets, but their mechanisms remain unclear.
- SKP1 is a known cancer target, but its role in colorectal cancer stem cells (CRC-SCs) is unknown.
Purpose of the Study:
- To investigate the role of SKP1 in promoting colorectal cancer stemness.
- To elucidate the underlying molecular mechanisms involving SKP1, RASSF1, and YAP in colorectal cancer (CRC).
Main Methods:
- Utilized human CRC cell lines and primary cells.
- Employed gene manipulation (lentivirus), qRT-PCR, western blot, sphere formation, transwell, limiting dilution, and xenograft assays.
- Investigated protein-DNA and protein-protein interactions using ChIP-PCR and co-immunoprecipitation.
Main Results:
- SKP1 was upregulated in CRC-SCs and correlated with poor prognosis.
- SKP1 overexpression enhanced CRC stemness (sphere formation, migration, self-renewal) and CSC marker expression.
- SKP1 suppressed RASSF1, attenuated Hippo signaling, and activated YAP, promoting CRC stemness.
Conclusions:
- SKP1 promotes CRC stemness by suppressing RASSF1 and activating YAP.
- SKP1 represents a potential therapeutic target for colorectal cancer treatment.
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