RNA-binding protein PCBP2 modulates glioma growth by regulating FHL3
Wei Han, … , Boqin Qiang, Xiaozhong Peng
Wei Han, … , Boqin Qiang, Xiaozhong Peng
Published April 15, 2013
Citation Information: J Clin Invest. 2013;123(5):2103-2118. https://doi.org/10.1172/JCI61820.
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Research Article Oncology

RNA-binding protein PCBP2 modulates glioma growth by regulating FHL3

Abstract

PCBP2 is a member of the poly(C)-binding protein (PCBP) family, which plays an important role in posttranscriptional and translational regulation by interacting with single-stranded poly(C) motifs in target mRNAs. Several PCBP family members have been reported to be involved in human malignancies. Here, we show that PCBP2 is upregulated in human glioma tissues and cell lines. Knockdown of PCBP2 inhibited glioma growth in vitro and in vivo through inhibition of cell-cycle progression and induction of caspase-3–mediated apoptosis. Thirty-five mRNAs were identified as putative PCBP2 targets/interactors using RIP-ChIP protein-RNA interaction arrays in a human glioma cell line, T98G. Four-and-a-half LIM domain 3 (FHL3) mRNA was downregulated in human gliomas and was identified as a PCBP2 target. Knockdown of PCBP2 enhanced the expression of FHL3 by stabilizing its mRNA. Overexpression of FHL3 attenuated cell growth and induced apoptosis. This study establishes a link between PCBP2 and FHL3 proteins and identifies a new pathway for regulating glioma progression.

Authors

Wei Han, Zhongshuai Xin, Zhiqiang Zhao, Wen Bao, Xihua Lin, Bin Yin, Jizong Zhao, Jiangang Yuan, Boqin Qiang, Xiaozhong Peng

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Figure 7

Identification of the FHL3 core recognition sequence; PCBP2 regulates FHL3 expression by stabilizing its mRNA.

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Identification of the FHL3 core recognition sequence; PCBP2 regulates FH...
(A) Schematic of FHL3 mRNA depicting the 3′ UTR-A in light gray rectangles. The templates for the synthesis of biotin-labeled 3′A-1 to 4 RNAs are indicated by dark lines and CU-rich patches are shown in red. (B) Detection of different binding abilities to PCBP2 by biotin pull-down analysis using the biotin-labeled 3′A-1 to 4 RNA. FHL3 mRNA 3′A and 3′B were included as positive and negative controls. (C) Nucleotide sequence of the identified FHL3 mRNA target binding site, with the core recognition sequence shown in red. (D) T98G cells were transiently transfected with the control siRNA or PCBP2 siRNA and either an empty pLuc reporter plasmid or a pLuc-FHL3 3′A-1,-2,-3,-4 reporter plasmid. *P < 0.001. (E) FHL3 mRNA stability assay in T98G cells transiently transfected with the control siRNA or PCBP2 siRNA. Transfectants were treated with the transcription inhibitor ActD for 0, 2, 4, and 6 hours; the whole-cell RNA was analyzed for FHL3 mRNA levels by an RNase protection assay. Representative blots are shown in the top panel. FHL3 mRNA levels were quantified by densitometric scanning. For each set of siRNA transfections, the band intensity at the 0 hour time point was set to 100%, and the percentage of mRNA remaining at the 2-, 4-, and 6-hour time points is plotted as shown in the bottom panel.