Abstract
Transcribed-ultraconserved regions (T-UCRs), which contain conserved sequences with 100% identity across human, rat and mouse species, are a novel category of functional RNAs. The human transformer 2β gene (TRA2B) encodes a UCR that spans exon 2 (276 bp) and its neighboring introns. Among five spliced RNA variants (TRA2β1-5) transcribed from the TRA2B gene, only TRA2β4 contains the conserved exon 2. TRA2β4 is overexpressed in colon cancer cells and accelerates cell growth by blocking the transcription of CDKN1A. However, the mechanisms underlying the overexpression of TRA2β4 in colon cancer cells are unknown. Using biotinylated RNA pull-down assays followed by liquid chromatography-mass spectrometric analysis, we identified nucleolin as a TRA2β4-binding protein. Knockdown of nucleolin reduced the nuclear retention of TRA2β4 and accelerated its degradation in the cytoplasm, whereas nucleolin overexpression increased TRA2β4 levels and its mitogenic activity. Nucleolin directly bound to TRA2β4 exon 2 via the glycine/arginine-rich (GAR) domain. Overexpression of GAR-deficient nucleolin failed to increase TRA2β4 expression and growth of colon cancer cells. RNA fluorescence in situ hybridization showed that TRA2β4 co-localized with nucleolin in nuclei but not with the mutant lacking GAR. Our results suggest that specific interactions between nucleolin and UCR-containing TRA2β4 may be associated with abnormal growth of colon cancer cells.
Highlights
Transformer 2β (Tra2β) belongs to the serine/ arginine-rich (SR)-like protein splicing factor family
Our results suggest that specific interactions between nucleolin and UCR-containing TRA2β4 may be associated with abnormal growth of colon cancer cells
To reveal the mechanism supporting the expression of TRA2β4 in colon cancer cells, we first searched for nuclear RNA-binding protein(s) that interacted with and retained TRA2β4 in nuclei
Summary
Transformer 2β (Tra2β) belongs to the serine/ arginine-rich (SR)-like protein splicing factor family It contains two SR domains and an RNA recognition motif (RRM), and it functions as a sequence-specific pre-mRNA splicing enhancer [1, 2]. Overexpression of Tra2β protein is associated with several types of cancer, including those in breast, ovary, and colon. It appears to accelerate tumor growth and metastasis, and induces drug resistance [3,4,5]. We previously reported that Hu antigen R regulated alternative splicing of TRA2β pre-mRNA to selectively produce TRA2β4 in human colon cancer cells under oxidative stress [11]. The accumulated TRA2β4 in the nucleus regulates gene expression (including CDKN1A) by occupying transcriptional factor Sp1, resulting in the promotion of cell growth by interrupting p21-related cellular senescence [12]
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