The Impact Of RBM3 Expression On The Whole Transcriptome Of Prostate Cancer Cells: An RNA Sequencing Study

Objective:The incidence of prostate cancer in male malignant tumors ranked second.With the acceleration of China's modernization and the aging of population,prostate cancer?PCa?has become one of the the major malignant tumor threatening male lives.RBM3?RNA-binding motif 3?was one of cold stress protein family.Recent years,it has been reported that the specific expression of RBM3 was involved in ovarian cancer,breast cancer,malignant melanoma,colorectal cancer,bladder cancer and prostate cancer.Therefore,it is considered to be a potential cancer related gene,and plays an important role in promoting cell proliferation and decreasing cell apoptosis,especially asscoiated with the occurrence,development,invasion and metastasis of PCa.However,the role of RBM3 in PCa remains controversial.In the study,we tried to use high-throughput sequencing to conduct preliminary exploration of the above problems at the gene level.Methods:1.We constructed a RBM3 knocking-down prostate cancer cell line,then extracted total RNA and constructed cDNA library.2.Using high-throughput sequencing technology,the cell samples were sequenced and the differentially expressed genes based on RBM3 were screened through the filtering and quality evaluation of the raw data.3.By GO analysis,pathway analysis,pathway act analysis and differentiallly expressed genes act network,we tyr to reveal the relationship between RBM3 and PCa.4.The sequencing results were verified by RT-qPCR?real time fluorescence quantitative PCR?,and the key DEGs for the progression of prostate cancer regulated by RBM3 were confirmed.Results:1.The transcriptome sequencing results revealed that there were 698 DEGs in RBM3 knocking-down prostate cancer cell line PC3-siRBM3,of which 446 genes were up-regulated and 252 genes were down-regulated respectively.2.GO and pathway analysis showed that the DEGs were enriched in lipid metabolism,synaptic transmission,membrane depolarization and G-protein coupled receptor signaling pathway.3.Pathways associated with lipid metabolism,such as ether lipid metabolism,alpha-linolenic acid metabolism and linoleic acid metabolism were the core pathways of the network,while pathways associated with neurological diseases or neurodegeneration,such as glutamatergic synapse and GABAergic synapse also occupied a prominent position in the network through pathway analysis and pathway act analysis.4.GO analysis,pathway analysis combined with differentiallly expressed genes act network analysis showed that the differentiallly expressed genes in lipidmetabolism-related pathways were closely related to PCa.Genes in the phospholipase A2family were enriched multiple times,such as PLA2G2A,PLA2G2F,PLA2G4C andPLA2G3,the expression of which were significantly obvious(p-value were PLA2G2A:0.0024,PLA2G2F:1.42×10^-5,PLA2G4C:0.046,PLA2G3:0.018,respectively).5.It was shown that GNG5,EDN1,NOS3,GAL,ACE and CD38 possessed higher node degree and closely related to each other as well as widely associated with other DEGs,and were the core genes in the network in differentiallly expressed genes act network.6.In addition,GNG5 and G-protein coupled receptor signaling pathway,EDN1 and angiogenesis pathways,and CD38 and signal transduction pathway,NOS3 and NO synthesis pathway,the relationship between CYP2B6 and drug metabolism,the state of activation of GAL and its receptor and polymorphism of ACE were also closely related to the regulation of RBM3.7.The results of RNA-seq were verified by RT-qPCR experiment.The results showed that most of the DEGs were in accordance with the RNA-seq.Conclusions:We applied to transcriptome sequencing and bioinformatics analysis of knocking-down of RBM3 in prostate cancer cell lines,and the results of RNA-seq were verified by RT-qPCR experiment.Tumor-related genes closely related to PCa,such as PLA2G2A,PLA2G2F,PLA2G4C,PLA2G3,etc.,were screened by bioinformatics analysis.Moreover,the lipid metabolism-related pathway was also found to be a core signaling pathway.These findings suggest that RBM3 might regulate the progress of PCa through the lipid metabolism pathway.And RBM3 might also have a close relationship with GABAergic synapse and glutamatergic synapse in the regulation of neurological diseases or neurodegeneration.These genes and pathways could be useful in further study of the development of PCa,providing a new perspective for our study of the regulation of RBM3 in the future.