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Identification of Cyclin-dependent Kinase 1 Specific Phosphorylation Sites by an In Vitro Kinase Assay
Published on: May 3, 2018
MiR-31-5p acts as a tumor suppressor in renal cell carcinoma by targeting cyclin-dependent kinase 1 (CDK1)
Yawen Li1, Jing Quan1, Fangfang Chen1
1Guangdong and Shenzhen Key Laboratory of Male Reproductive Medicine and Genetics, Institute of Urology, Peking University Shenzhen Hospital, Clinical College of Anhui Medical University, Shenzhen, 518036, China.
Background:
Renal cell carcinoma (RCC) accounts for more than 90% of cancers in the kidney. RCC is often asymptomatic, as a result people with RCC generally have advanced disease by the time it is discovered and has a poor prognosis compared to other cancers. Therefore, it is necessary to explore its pathogenesis and identify some reliable prognostic biomarker of RCC. miRNAs are emerging as important players in the development and progression of RCC. miR-31-5p has been reported to act as a tumor suppressor in hepatocellular carcinoma (HCC). The aim of this study is to determine the detailed molecular mechanism of miR-31-5p in the progression of RCC and to investigate its potential clinical value.
Methods:
In this study, RT-qPCR, EdU assay, CCK-8 assay, wound scratch assay, transwell assay, flow cytometry assay and cell cycle assay were performed to detect miR-31-5p expression and its functions in RCC. Moreover, 42 formalin-fixed paraffin-embedded (FFPE) RCC samples were used to analyze the relationship between miR-31-5p expression and patients' overall survival. Finally, luciferase reporter assay, RT-qPCR assay and western blot were used to explore the association between miR-31-5p and its potential targets.
Results:
miR-31-5p was significantly down-regulated in RCC tissues and RCC cell lines compared with paired adjacent normal tissues and normal cell lines. miR-31-5p downregulation was associated with poor prognosis in RCC patients. Overexpression of miR-31-5p inhibited RCC cell proliferation, migration and invasion and cell cycle. Conversely, down-regulation of miR-31-5p promoted cell proliferation, migration and invasion. Furthermore, cyclin-dependent kinasec1 (CDK1), a key player in cell cycle regulation, was identified as a functional target of miR-31-5p.
Conclusions:
Our results suggest that miR-31-5p serves as a tumor suppressor in RCC and is expected to be a molecular biomarker for poor prognosis of RCC.
Insights
MicroRNA-31-5p acts as a tumor suppressor in renal cell carcinoma (RCC). Its reduced expression correlates with poor prognosis, inhibiting cancer progression and cell cycle, making it a potential biomarker for RCC.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Renal cell carcinoma (RCC) is the most common kidney cancer, often diagnosed at advanced stages with poor prognosis.
- MicroRNAs (miRNAs) are implicated in RCC pathogenesis, with miR-31-5p showing tumor-suppressive roles in other cancers.
- Identifying reliable prognostic biomarkers for RCC is crucial for improving patient outcomes.
Purpose of the Study:
- To elucidate the molecular mechanisms of miR-31-5p in renal cell carcinoma (RCC) progression.
- To investigate the potential clinical utility of miR-31-5p as a prognostic biomarker in RCC.
Main Methods:
- Quantitative real-time PCR (RT-qPCR) to assess miR-31-5p expression in RCC tissues and cell lines.
- In vitro assays (EdU, CCK-8, wound scratch, Transwell) to evaluate the functional impact of miR-31-5p on RCC cell proliferation, migration, and invasion.
- Luciferase reporter assays and Western blot to identify and validate miR-31-5p targets, including CDK1.
Main Results:
- miR-31-5p was significantly downregulated in RCC tissues and cell lines compared to normal controls.
- Lower miR-31-5p expression correlated with poorer overall survival in RCC patients.
- Overexpression of miR-31-5p suppressed RCC cell proliferation, migration, invasion, and cell cycle progression, while its downregulation promoted these processes.
- Cyclin-dependent kinase 1 (CDK1) was identified as a direct functional target of miR-31-5p.
Conclusions:
- miR-31-5p functions as a tumor suppressor in renal cell carcinoma (RCC).
- miR-31-5p is a potential molecular biomarker for predicting poor prognosis in RCC patients.
- Targeting miR-31-5p may offer therapeutic strategies for RCC.
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