Study On The Regulation And Expression Mechanism Of BLM Helicase By LncRNA

RecQ-BLM helicase plays an important regulatory role in the process of cell proliferation and growth,and the deletion of BLM helicase can cause the genetic disease of human Bloom syndrome,namely facial erythema homunculus syndrome,and is susceptible to various cancers.Lnc RNAs also play an important regulatory role in the regulation of cell growth and proliferation and other processes.Lnc RNAs can play a role through the mechanism of competitive endogenous RNA(ce RNA).When they act as endogenous competitive RNAs,It can competitively bind micro RNAs(mi RNAs)to relieve or partially relieve the inhibition of mi RNAs on target genes and promote the expression of target genes,to improve cell proliferation and other abilities.As the downstream transcriptional coactivators and key components of the Hippo signaling pathway in mammals,YAP and TAZ proteins are mainly involved in a variety of physiological processes such as cell proliferation,apoptosis,and migration in cell regulation,and can be used as index genes for proliferation detection.There have been few reports on how lnc RNA regulates cancer through the BLM gene.Previous studies of our group have confirmed that mi R-27b-3p can directly regulate the 3'UTR region of BLM helicase and inhibit the expression of BLM.Therefore,in this study,mi R-27b-3p was used as the material to screen lnc RNAs with regulatory effects on BLM.The eukaryotic expression vector of lnc RNA was constructed by the molecular cloning technique.CCK8 proliferation test,scratch test,real-time fluorescence quantitative PCR test,and Western blotting test were used to verify the BLM-related lnc RNA LINC183(N183)and lnc RNA LINC212(N212).We analyzed the expression levels of N183 and N212 in 22RV1,PC3,and WPMY-1 cells and the effects of overexpression on the m RNA and protein expression levels of BLM,YAP and TAZ genes in 22RV1,and PC3 cells,as well as the effects on cell proliferation and migration.The main research results are as follows:1.Screening and expression profile analysis of BLM-related Lnc RNAsTwo Lnc RNAs,NONHSAT184603.1(referred to as Lnc RNA LINC183,N183)and NONHSAT216122.1(referred to as Lnc RNA LINC212,N212),were successfully screened.The full length of N183 was 766 bp,located on chromosome 2,and the full length of N212 was 1195 bp,located on chromosome 8.Compared with WPMY-1,the expression of BLM gene in 22RV1 and PC3 cells was extremely significantly down-regulated(P <0.01),and the expression of N183 and N212 genes in 22RV1 and PC3 cells was extremely significantly up-regulated(P <0.01).2.The effect of over-expression of N183 and N212 on the expression level of BLM geneThe eukaryotic over-expression vectors pc DNA3.1-N183 and pc DNA3.1-N212 were constructed and transfected into WPMY-1,22RV1,and PC3 cells.The m RNA and protein levels of BLM,YAP,and TAZ genes were detected by real-time fluorescence quantitative PCR and Western Blotting.The results showed that N183 and N212 could increase the expression of BLM helicase at m RNA and protein levels,and may have an effect on cell proliferation.The down-regulation of BLM protein in22RV1 cells after transfection with N212 might be since N212 mainly plays a role at the transcriptional level.3.N183 and N212 promote the proliferation and migration of prostate cancer cellsCCK8 proliferation assay was used to detect the cell activity at different time points after over-expression.The results showed that the proliferation of 22RV1 cells after over-expression of N183 was significantly different from that of the blank control group(P <0.01),and was significantly up-regulated.Compared with the blank control group,the proliferation of PC3 cells after over-expression of N212 was significantly different(P <0.01),and was significantly up-regulated.These results indicate that N183 and N212 can promote the proliferation of prostate cancer cells.Cell migration was detected by cell scratch assay.The results showed that the healing area of 22RV1 cells transfected with N183 and N212 eukaryotic over-expression vector was significantly different from that of the blank control group(P <0.01),and was significantly reduced.The healing area of PC3 cells transfected with N183 and N212 eukaryotic over-expression vector was significantly different from that of the blank control group(P <0.01),and was significantly reduced.N183 and N212 can promote the migration of prostate cancer cells.In conclusion,this study confirmed that N183 and N212 are highly expressed in prostate cancer cells,and N183 and N212 are acting through mi RNA pathways,and mi R-27b-3p positively acts on BLM,enhances BLM expression,and promotes the proliferation and migration of prostate cancer cells.