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.
View: Text | PDF
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

×

Figure 2

Inhibition of glioma cell growth in vitro by knockdown of PCBP2.

Options: View larger image (or click on image) Download as PowerPoint
Inhibition of glioma cell growth in vitro by knockdown of PCBP2. (A) Wes...
(A) Western blot of PCBP2 expression in 3 glioma cell lines (T98G, U87MG, and U251) transiently transfected with the control siRNA or PCBP2 siRNA for 48 to 72 hours using Lipofectamine 2000. β-Actin was used as a loading control. (B) MTT assay on the same 3 glioma cell lines after transfection with the control siRNA or PCBP2 siRNA as above. Data are presented as the mean ± SD and are representative of 3 wells. *P < 0.05 compared with the control siRNA by a 2-tailed Student’s t test. (C) Approximately 72 hours after transfection, the 3 glioma cell lines were analyzed by flow cytometry. The proportions of cells in the G1, G2, and S phases of the cell cycle are depicted in the histograms. *P < 0.05 compared with the control siRNA by a 2-tailed Student’s t test. (D and E) Representative Western blot showing P27, P21, and P16 protein levels (D) and expression levels of differentially phosphorylated pRb (E) in the transfected glioma cell lines. P16 was deleted in U87MG and U251 cells. (F) Nuclear TUNEL staining for apoptotic cells in the glioma cell lines after approximately 72 hours of transfection. The ratio of TUNEL-positive cells was calculated (n = 5) and plotted on the histogram. *P < 0.05 compared with the control siRNA by a 2-tailed Student’s t test. (G) Representative Western blot showing (cleaved) caspase-3 and its substrate (cleaved) poly (ADP-ribose) polymerase (PARP) in the transfected glioma cell lines.