Abstract
BackgroundAberrant spliced isoforms are specifically associated with cancer progression and metastasis. The cytoplasmic adaptor CRKL (v-crk avian sarcoma virus CT10 oncogene homolog-like) is a CRK like proto-oncogene, which encodes a SH2 and SH3 (src homology) domain-containing adaptor protein. CRKL is tightly linked to leukemia via its binding partners BCR-ABL and TEL-ABL, upregulated in multiple types of human cancers, and induce cancer cell proliferation and invasion. However, it remains unclear whether signaling adaptors such as CRKL could regulate alternative splicing.MethodsWe analyzed the expression level of CRKL in 305 cervical cancer tissue samples available in TCGA database, and then selected two groups of cancer samples with CRKL differentially expressed to analyzed potential CRKL-regulated alternative splicing events (ASEs). CRKL was knocked down by shRNA to further study CRKL-regulated alternative splicing and the activity of SR protein kinases in HeLa cells using RNA-Seq and Western blot techniques. We validated 43 CRKL-regulated ASEs detected by RNA-seq in HeLa cells, using RT-qPCR analysis of HeLa cell samples and using RNA-seq data of the two group of clinical cervical samples.ResultsThe expression of CRKL was mostly up-regulated in stage I cervical cancer samples. Knock-down of CRKL led to a reduced cell proliferation. CRKL-regulated alternative splicing of a large number of genes were enriched in cancer-related functional pathways, among which DNA repair and G2/M mitotic cell cycle, GnRH signaling were shared among the top 10 enriched GO terms and KEGG pathways by results from clinical samples and HeLa cell model. We showed that CRKL-regulated ASEs revealed by computational analysis using ABLas software in HeLa cell were highly validated by RT-qPCR, and also validated by cervical cancer clinical samples.ConclusionsThis is the first report of CRKL-regulation of the alternative splicing of a number of genes critical in tumorigenesis and cancer progression, which is consistent with CRKL reported role as a signaling adaptor and a kinase. Our results underline that the signaling adaptor CRKL might integrate the external and intrinsic cellular signals and coordinate the dynamic activation of cellular signaling pathways including alternative splicing regulation.
Highlights
Aberrant spliced isoforms are associated with cancer progression and metastasis
We further showed that 94% of CRK like proto-oncogene (CRKL)-regulated alternative splicing events detected in HeLa cells could be validated by Quantitative reverse transcription polymerase chain reaction (RT-qPCR) approach
Expression of CRKL is upregulated in cervical cancer, more pronouncedly in early stages, as revealed by TCGA data Inspired by previous studies on the overexpression of CRKL in a small number of cervical cancer samples, we downloaded RNA-seq expression data for all samples available for cervical cancer in TCGA (The Cancer Genome Atlas) database, which included 305 cervical tumor and 3 normal samples
Summary
Aberrant spliced isoforms are associated with cancer progression and metastasis. CRKL is tightly linked to leukemia via its binding partners BCR-ABL and TEL-ABL, upregulated in multiple types of human cancers, and induce cancer cell proliferation and invasion. It remains unclear whether signaling adaptors such as CRKL could regulate alternative splicing. Noncanonical and cancer-specific mRNA transcripts produced by the aberrant splicing can lead to loss of function of tumor suppressors or activator of oncogenes and cancer pathways [21]. These cancer-specific isoforms may represent attractive cancer therapeutic targets. It was reported that alternative splicing regulates cervical cancer oncogenesis via miL1RAP-NF-κB-CD47 axis, indicative of an attractive therapeutic target for treatment of cervical cancer [22]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
