RNA-binding Protein UNR Promotes Glioma Cell Migration and Regulates the Expression of Ribosomal Protein L9

Tian Ningyu; Qi Yingjiao; Hu Yan; Yin Bin; Yuan Jiangang; Qiang Boqin; Peng Xiaozhong; Han Wei

doi:10.24920/11815

Your Location：

Home >

Browse articles >

RNA-binding Protein UNR Promotes Glioma Cell Migration and Regulates the Expression of Ribosomal Protein L9

Chinese Medical Sciences Journal Vol. 33, Issue 3, Pages: 143-151(2018)

ORIGINAL ARTICLE | Updated：2024-04-10

- RNA-binding Protein UNR Promotes Glioma Cell Migration and Regulates the Expression of Ribosomal Protein L9
- Affiliations：
  
  中国医学科学院基础医学研究所北京协和医学院基础学院,国家医学分子生物重点实验室,北京 100005
- Author bio：
- Funds：
- DOI：10.24920/11815
  CLC：
- Published Online：20 September 2018，
  
  Published：20 September 2018
- Accepted：
Scan QR Code
Tian Ningyu, Qi Yingjiao, Hu Yan, et al. RNA-binding Protein UNR Promotes Glioma Cell Migration and Regulates the Expression of Ribosomal Protein L9[J]. Chinese medical sciences journal, 2018, 33(3): 143-151.
DOI：

Tian Ningyu, Qi Yingjiao, Hu Yan, et al. RNA-binding Protein UNR Promotes Glioma Cell Migration and Regulates the Expression of Ribosomal Protein L9[J]. Chinese medical sciences journal, 2018, 33(3): 143-151. DOI： 10.24920/11815.

摘要

目的

研究RNA结合蛋白UNR在神经胶质瘤发生发展中的作用以及其发挥作用的分子机制。

方法

首先

利用生物信息学方法分析CGGA数据库中UNR的表达水平及其表达水平高低与病人预后的关系。采用western blot和real-time PCR方法分析胶质瘤病人组织样本和细胞株中UNR的表达水平。其次

在胶质瘤细胞株A172

HS683

LN18中转染siRNA敲低UNR

采用MTS和划痕实验检测肿瘤细胞增殖能力以及迁移能力的变化。再次

通过分析已发布的黑色素瘤UNR iCLIP-seq、RNA-seq 和核糖体印迹测序数据库筛选出UNR在胶质瘤细胞中可能结合的靶mRNA。利用RNA免疫沉淀和生物素标记的RNA pull down实验确认UNR可结合核糖体蛋白L9（RPL9）的mRNA。最后

通过western blot检测RPL9在胶质瘤细胞株中的表达水平

并且检测敲低UNR以后RPL9蛋白水平的变化。在神经胶质瘤细胞株A172

T98G中转染siRNA敲低RPL9

检测和肿瘤侵袭迁移有关的上皮向间充质转化有关的标志物—波形蛋白的表达变化。

结果

生物信息学分析UNR mRNA表达水平

发现其在恶性程度更高的胶质瘤样本中表达较高(P

0.001)。高表达UNR的患者预后较差（P=0.0177）。与正常细胞株以及正常脑组织样本相比

UNR在9株胶质瘤细胞以及患者组织样本中具有较高的表达水平（P

0.01)。敲低UNR抑制肿瘤细胞的迁移能力(P

0.05)

但不影响其增殖能力。UNR可以结合在PTEN mRNA和RPL9 mRNA的3’非翻译区。敲低UNR以后RPL9的表达水平下调。RPL9在9株胶质瘤细胞中具有高表达的趋势

RPL9可以正向调控波形蛋白的表达。

结论

本研究揭示了RNA结合蛋白UNR在胶质瘤细胞迁移方面的功能

并且发现UNR可以结合PTEN和RPL9 mRNA的3’非翻译区并调控RPL9的表达。这为我们更近一步研究胶质瘤的发生发展提供了新思路。

Abstract

Objective

To investigate the role of RNA binding protein—upstream-of-N-Ras (UNR) in the development of glioma and its molecular mechanism.

Methods

First

bioinformatics analysis of CGGA database was performed to detect UNR expression level and prognosis of patients with glioma. Western blot and real-time PCR were used to detect UNR expression level in glioma cell lines and tissues. Next

UNR siRNAs were transfected in glioma cells

and MTS assay and scratch wound-healing assay were used to detect changes in cell proliferation and migration. Then

the candidate UNR target mRNAs were identified by analyzing the sequencing data of UNR iCLIP-seq

RNA sequencing and ribosome profiling databases of human melanoma. RNA immunoprecipitation and biotin pull-down assays were used to identify the UNR target mRNAs in glioma cells. Finally

western blot was used to detect the effect of UNR knockdown on ribosomal protein L9 (RPL9) and RPL9 protein expression level in glioma cell lines. RPL9 siRNA was transfected in A172 and T98G and the expression of vimentin in the cells was detected with western blot.

Results

Bioinformatics analysis showed that UNR mRNA expression level was significantly higher in high-grade glioma [Grade Ⅱ (

=126)

Grade Ⅲ (

=51)

Grade Ⅳ (

=128)

0.001]. UNR high expression levels were associated with poor prognosis (

=0.0177). UNR had high expression level in glioma cell lines and patient samples compared with normal cell lines and normal brain samples (

0.01). Knockdown of UNR inhibited glioma cells migration (

0.05)

but did not inhibit glioma cells growth in three glioma cell lines. UNR binded the 3’ untranslated region (UTR) of PTEN and RPL9 mRNAs. RPL9 protein was significantly highly expressed in most glioma cell lines (

=9) and knockdown of UNR resulted in a downregulation of RPL9 protein expression. Epithelial-mesenchymal transition (EMT)-related marker—vimentin was positively regulated by RPL9.

Conclusions

UNR could bind to the 3’UTR of PTEN and RPL9 in glioma cell lines

therefore promoting glioma cell migration and regulating the expression of RPL9. Here

we establish a link between UNR and RPL9 protein

which will provide new ideas for the further study of glioma.

关键词

Keywords

references

Marchese D , de Groot NS , Lorenzo Gotor N , et al . Advances in the characterization of RNA-binding proteins . Wiley Interdiscip Rev RNA 2016 ; 7 ( 6 ): 793 - 810 . doi: 10.1002/wrna.1378 10.1002/wrna.1378 . DOI: 10.1002/wrna.1378 http://doi.org/10.1002/wrna.1378 http://doi.wiley.com/10.1002/wrna.1378 http://doi.wiley.com/10.1002/wrna.1378

Campos-Melo D , Droppelmann CA , Volkening K , et al . RNA-binding proteins as molecular links between cancer and neurodegeneration . Biogerontology 2014 ; 15 ( 6 ): 587 - 610 . doi: 10.1007/s10522-014-9531-2 10.1007/s10522-014-9531-2 . DOI: 10.1007/s10522-014-9531-2 http://doi.org/10.1007/s10522-014-9531-2 http://link.springer.com/10.1007/s10522-014-9531-2 http://link.springer.com/10.1007/s10522-014-9531-2

Han W , Xin Z , Zhao Z , et al . RNA-binding protein PCBP2 modulates glioma growth by regulating FHL3 . J Clin Invest 2013 ; 123 ( 5 ): 2103 - 18 . doi: 10.1172/JCI61820 10.1172/JCI61820 . DOI: 10.1172/JCI61820 http://doi.org/10.1172/JCI61820 http://www.jci.org/articles/view/61820 http://www.jci.org/articles/view/61820

Jeffers M , Paciucci R , Pellicer A . Characterization of UNR: a gene closely linked to N-ras . Nucleic Acids Res 1990 ; 18 ( 16 ): 4891 - 9 . DOI: 10.1093/nar/18.16.4883 http://doi.org/10.1093/nar/18.16.4883 http://www.ncbi.nlm.nih.gov/pubmed/2204029 http://www.ncbi.nlm.nih.gov/pubmed/2204029

Hunt SL , Hsuan JJ , Totty N , et al . UNR, a cellular cytoplasmic RNA-binding protein with five cold-shock domains, is required for internal initiation of translation of human rhinovirus RNA . Genes Dev 1999 ; 13 ( 4 ): 437 - 48 . DOI: 10.1101/gad.13.4.437 http://doi.org/10.1101/gad.13.4.437 http://www.genesdev.org/cgi/doi/10.1101/gad.13.4.437 http://www.genesdev.org/cgi/doi/10.1101/gad.13.4.437

Boussadia O , Niepmann M , Creancier L , et al . UNR is required in vivo for efficient initiation of translation from the internal ribosome entry sites of both rhinovirus and poliovirus . J Virol 2003 ; 77 ( 6 ): 3353 - 9 . DOI: 10.1128/JVI.77.6.3353-3359.2003 http://doi.org/10.1128/JVI.77.6.3353-3359.2003 http://jvi.asm.org/cgi/doi/10.1128/JVI.77.6.3353-3359.2003 http://jvi.asm.org/cgi/doi/10.1128/JVI.77.6.3353-3359.2003

Cornelis S , Tinton SA , Schepens B , et al . UNR translation can be driven by an IRES element that is negatively regulated by polypyrimidine tract binding protein . Nucleic Acids Res 2005 ; 33 ( 10 ): 3095 - 108 . DOI: 10.1093/nar/gki611 http://doi.org/10.1093/nar/gki611 https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gki611 https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gki611

Dormoy-Raclet V , Markovits J , Jacquemin-Sablon A , et al . Regulation of UNR expression by 5’- and 3’-untranslated regions of its mRNA through modulation of stability and IRES mediated translation . RNA Biol 2005 ; 2 ( 3 ): e27 - 35 .

Patel GP , Ma S , Bag J . The autoregulatory translational control element of poly(A)-binding protein mRNA forms a heteromeric ribonucleoprotein complex . Nucleic Acids Res 2005 ; 33 ( 22 ): 7074 - 89 . DOI: 10.1093/nar/gki1014 http://doi.org/10.1093/nar/gki1014 https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gki1014 https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gki1014

Elatmani H , Dormoy-Raclet V , Dubus P , et al . The RNA-binding protein UNR prevents mouse embryonic stem cells differentiation toward the primitive endoderm lineage . Stem Cells 2011 ; 29 ( 10 ): 1504 - 16 . doi: 10.1002/stem.712 10.1002/stem.712 . DOI: 10.1002/stem.712 http://doi.org/10.1002/stem.712 http://doi.wiley.com/10.1002/stem.712 http://doi.wiley.com/10.1002/stem.712

Dinur M , Kilav R , Sela-Brown A , et al . In vitro evidence that upstream of N-ras participates in the regulation of parathyroid hormone messenger ribonucleic acid stability . Mol Endocrinol 2006 ; 20 ( 7 ): 1652 - 60 . DOI: 10.1210/me.2005-0333 http://doi.org/10.1210/me.2005-0333 https://academic.oup.com/mend/article-lookup/doi/10.1210/me.2005-0333 https://academic.oup.com/mend/article-lookup/doi/10.1210/me.2005-0333

Kobayashi H , Kawauchi D , Hashimoto Y , et al . The control of precerebellar neuron migration by RNA-binding protein Csde1 . Neuroscience 2013 ; 253 : 292 - 303 . doi: 10.1016/j.neuroscience.2013.08.055 10.1016/j.neuroscience.2013.08.055 . DOI: 10.1016/j.neuroscience.2013.08.055 http://doi.org/10.1016/j.neuroscience.2013.08.055 http://linkinghub.elsevier.com/retrieve/pii/S0306452213007513 http://linkinghub.elsevier.com/retrieve/pii/S0306452213007513

Ju Lee H , Bartsch D , Xiao C , et al . A post-transcriptional program coordinated by CSDE1 prevents intrinsic neural differentiation of human embryonic stem cells . Nat Commun 2017 ; 8 ( 1 ): 1456 . doi: 10.1038/s41467-017-01744-5 10.1038/s41467-017-01744-5 . DOI: 10.1038/s41467-017-01744-5 http://doi.org/10.1038/s41467-017-01744-5 http://www.nature.com/articles/s41467-017-01744-5 http://www.nature.com/articles/s41467-017-01744-5

Horos R , Ijspeert H , Pospisilova D , et al . Ribosomal deficiencies in Diamond-Blackfan anemia impair translation of transcripts essential for differentiation of murine and human erythroblasts . Blood 2012 ; 119 ( 1 ): 262 - 72 . doi: 10.1182/blood-2011-06-358200 10.1182/blood-2011-06-358200 . DOI: 10.1182/blood-2011-06-358200 http://doi.org/10.1182/blood-2011-06-358200 http://www.bloodjournal.org/cgi/doi/10.1182/blood-2011-06-358200 http://www.bloodjournal.org/cgi/doi/10.1182/blood-2011-06-358200

Xia K , Guo H , Hu Z , et al . Common genetic variants on 1p13.2 associate with risk of autism . Mol Psychiatry 2014 ; 19 ( 11 ): 1212 - 9 . doi: 10.1038/mp.2013.146 10.1038/mp.2013.146 . DOI: 10.1038/mp.2013.146 http://doi.org/10.1038/mp.2013.146 http://www.nature.com/articles/mp2013146 http://www.nature.com/articles/mp2013146

Wurth L , Papasaikas P , Olmeda D , et al . UNR/CSDE1 drives a post-transcriptional program to promote melanoma invasion and metastasis . Cancer Cell 2016 ; 30 ( 5 ): 694 - 707 . doi: 10.1016/j.ccell.2016.10.004 10.1016/j.ccell.2016.10.004 . DOI: 10.1016/j.ccell.2016.10.004 http://doi.org/10.1016/j.ccell.2016.10.004 https://linkinghub.elsevier.com/retrieve/pii/S1535610816304913 https://linkinghub.elsevier.com/retrieve/pii/S1535610816304913

Martinez-Useros J , Georgiev-Hristov T , Fernandez-Acenero MJ , et al . UNR/CDSE1 expression as prognosis biomarker in resectable pancreatic ductal adenocarcinoma patients: a proof-of-concept . PLoS One 2017 ; 12 ( 8 ): e0182044 . doi: 10.1371/journal.pone.0182044 10.1371/journal.pone.0182044 . DOI: 10.1371/journal.pone.0182044 http://doi.org/10.1371/journal.pone.0182044 http://dx.plos.org/10.1371/journal.pone.0182044 http://dx.plos.org/10.1371/journal.pone.0182044

Hu PS , Xia QS , Wu F , et al . NSPc1 promotes cancer stem cell self-renewal by repressing the synthesis of all-trans retinoic acid via targeting RDH16 in malignant glioma . Oncogene 2017 ; 36 ( 33 ): 4706 - 18 . doi: 10.1038/onc.2017.34 10.1038/onc.2017.34 . DOI: 10.1038/onc.2017.34 http://doi.org/10.1038/onc.2017.34 http://www.nature.com/doifinder/10.1038/onc.2017.34 http://www.nature.com/doifinder/10.1038/onc.2017.34

Ray S , Anderson EC . Stimulation of translation by human Unr requires cold shock domains 2 and 4, and correlates with poly(A) binding protein interaction . Sci Rep 2016 ; 6 : 22461 . doi: 10.1038/srep22461 10.1038/srep22461 . DOI: 10.1038/srep22461 http://doi.org/10.1038/srep22461 http://www.nature.com/articles/srep22461 http://www.nature.com/articles/srep22461

Dormoy-Raclet V , Markovits J , Malato Y , et al . UNR, a cytoplasmic RNA-binding protein with cold-shock domains, is involved in control of apoptosis in ES and HuH7 cells . Oncogene 2007 ; 26 ( 18 ): 2595 - 605 . DOI: 10.1038/sj.onc.1210068 http://doi.org/10.1038/sj.onc.1210068 http://www.nature.com/articles/1210068 http://www.nature.com/articles/1210068

Tang Z , Li C , Kang B , et al . GEPIA: a web server for cancer and normal gene expression profiling and interactive analyses . Nucleic Acids Res 2017 ; 45 ( W1 ): W98 - W102 . doi: 10.1093/nar/gkx247 10.1093/nar/gkx247 . DOI: 10.1093/nar/gkx247 http://doi.org/10.1093/nar/gkx247 https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gkx247 https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gkx247

Zhang Y , Chao T , Li R , et al . MicroRNA-128 inhibits glioma cells proliferation by targeting transcription factor E2F3a . J Mol Med (Berl) 2009 ; 87 ( 1 ): 43 - 51 . doi: 10.1007/s00109-008-0403-6 10.1007/s00109-008-0403-6 . DOI: 10.1007/s00109-008-0403-6 http://doi.org/10.1007/s00109-008-0403-6 http://link.springer.com/10.1007/s00109-008-0403-6 http://link.springer.com/10.1007/s00109-008-0403-6

Legler JM , Ries LA , Smith MA , et al . Cancer surveillance series [corrected]: brain and other central nervous system cancers: recent trends in incidence and mortality . J Natl Cancer Inst 1999 ; 91 ( 16 ): 1382 - 90 . DOI: 10.1093/jnci/91.16.1382 http://doi.org/10.1093/jnci/91.16.1382 https://academic.oup.com/jnci/article-lookup/doi/10.1093/jnci/91.16.1382 https://academic.oup.com/jnci/article-lookup/doi/10.1093/jnci/91.16.1382

Views

1321

Downloads

CSCD

Alert me when the article has been cited

Submit

Publicity Resources

No data

Related Author

No data

Related Institution

No data

⁰