Knockdown Of PRR11Expression In Lung Cancer: A Critical Role For Cell Proliferation And Metastasis

Proline rich11(PRR11) is a novel cancer-associated gene identifiedin our previous study. As an evolutionary conserved gene, PRR11probablyplay an important role in cell proliferation and metastasis.In this paper, we conducted the following parts to investigate theeffect of PRR11depletion on cell proliferation and metastasis, and explorethe underlying molecular mechanism.(1) siRNA-mediated PRR11depletion affected the cell cycleprogression in HeLa cells. The knockdown of PRR11expression in Helacells was performed by siRNA transfection and then confirmed byqRT-PCR and Western blot.Cell proliferation assay showed that the cellproliferation was inhibited after PRR11depletion. Furthermore, flowcytometry assay revealed that silencing PRR11expression led to cell cyclearrest in S pahse. Additionally,Cell cycle associated genes (CCNA1,RRM1, etc.) were concertedly downregulated responding to the PRR11depletion.(2) siRNA-mediated PRR11depletion affected the cell proliferation and metastasis in lung cancer cells.The PRR11-targeted siRNAs weretransfected into lung cancer cells, including H1299, A549, and HCC827cells. The silencing efficiency of PRR11expression was confirmed byqRT-PCR and Western blot afterwards. Cell proliferation assay exhibitedthat PRR11depletion suppressed cell proliferation in lung cancer cells.Transwell migration/invasion assay revealed that knockdown of PRR11also inhibited cell migration and invasion ability in H1299Cells.（3）Prognostic analysis of PRR11in tumor tissues of lung cancer.ThemRNA expression profiles of lung cancer with prognosis information wereretrieved from Gene Expression Omnibus. The univariate survival analysesand multivariate Cox regression analysis showed that the PRR11mRNAlevel was an independent prognostic factor for the overall survival of lungcancer patients, with a association of high levels of PRR11with poorprognosis.(4) Analysis of gene expression changes responding tosiRNA-mediated PRR11depletion in lung cancer H1299Cells. siRNAswere used to inhibit the PRR11expression, total RNAs were then preparedand subjected to microarray analysis. The differentially expressed geneswere enriched for GO and pathway analysis. qRT-PCR and Western blotwas used to verify several important differentially expressed genes.Microarray analysis revealed that siRNA-mediated PRR11depletionresulted into the expression changes of550genes including139 upregulated and411downregulated. Bioinformatic analysis,qRT-PCR andWestern blot analysis verified that siRNA-mediated PRR11depletion led tothe expression changes of several cell cycle-and cancer-related genesincluding DHRS2, EPB41L3, CCNA1, MAP4K4, RRM1and NFIB. Takentogether, the present study suggested that PRR11might be implicated inlung cancer development via regulating the aforementioned genes and/orpathways.Taken together, our study demostrated that PRR11probably promotedthe proliferation and metastasis of lung cancer cells, via dysregulatingvarious genes involved in cell cycle and metastasis (e.g. DHRS2、EPB41L3、CCNA1、MAP4K4、RRM1、NFIB). In addition, PRR11mightserve as an independent predictor for survival of lung cancer patients.