期刊
JOURNAL OF HEMATOLOGY & ONCOLOGY
卷 14, 期 1, 页码 -出版社
BMC
DOI: 10.1186/s13045-021-01072-8
关键词
Pancreatic cancer; Alternative splicing; CLK1; SRSF5; M6A Modification; METTL14; Cyclin L2
资金
- National Natural Science Foundation of China [81772560]
- Natural Science Foundation for Distinguished Young Scholars of Fujian Province [2018J06020]
- Joint Funds for the innovation of science and technology, Fujian Province [2018Y9098]
- Fujian Provincial Hospital [2019HSJJ13]
- Education and Scientific Research Foundation of Fujian Province [2060402]
- Fujian Provincial Health and Family Planning Research Medical Inovation Project [2019-cx-3]
The study indicates that the CLK1/SRSF5 pathway induces abnormal exon skipping of METTL14 and Cyclin L2, promoting growth and metastasis of PDAC cells while regulating m6A methylation. This research suggests the potential prognostic value and therapeutic targeting of this pathway in PDAC patients.
Background Both aberrant alternative splicing and m6A methylation play complicated roles in the development of pancreatic cancer (PC), while the relationship between these two RNA modifications remains unclear. Methods RNA sequencing (RNA-seq) was performed using 15 pairs of pancreatic ductal adenocarcinoma (PDAC) tissues and corresponding normal tissues, and Cdc2-like kinases 1 (CLK1) was identified as a significantly upregulated alternative splicing related gene. Real-time quantitative PCR (qPCR) and western blotting were applied to determine the CLK1 levels. The prognostic value of CLK1 was elucidated by Immunohistochemistry (IHC) analyses in two independent PDAC cohorts. The functional characterizations and mechanistic insights of CLK1 in PDAC growth and metastasis were evaluated with PDAC cell lines and nude mice. SR-like splicing factors5(250-Ser) (SRSF5(250-Ser)) was identified as an important target phosphorylation site by phosphorylation mass spectrometry. Through transcriptome sequencing, Methyltransferase-like 14(exon10) (METTL14(exon10)) and Cyclin L2(exon6.3) skipping were identified as key alternative splicing events regulated by the CLK1-SRSF5 axis. RIP assays, RNA-pulldown and CLIP-qPCR were performed to confirm molecular interactions and the precise binding sites. The roles of the shift of METTL14(exon 10) and Cyclin L2(exon6.3) skipping were surveyed. Results CLK1 expression was significantly increased in PDAC tissues at both the mRNA and protein levels. High CLK1 expression was associated with poor prognosis. Elevated CLK1 expression promoted growth and metastasis of PC cells in vitro and in vivo. Mechanistically, CLK1 enhanced phosphorylation on SRSF5(250-Ser), which inhibited METTL14(exon10) skipping while promoted Cyclin L2(exon6.3) skipping. In addition, aberrant METTL14(exon 10) skipping enhanced the N6-methyladenosine modification level and metastasis, while aberrant Cyclin L2(exon6.3) promoted proliferation of PDAC cells. Conclusions The CLK1/SRSF5 pathway induces aberrant exon skipping of METTL14 and Cyclin L2, which promotes growth and metastasis and regulates m6A methylation of PDAC cells. This study suggests the potential prognostic value and therapeutic targeting of this pathway in PDAC patients.
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