Study On Mechanisms Underlying The Mutual Regulation Between LncRNA MALAT1/NEAT1 And SMYD3 In Breast Cancer Cells

Studies have shown that LncRNA MALAT1 and NEAT1 are abnormally expressed in various cancers including breast cancer and promote tumorigenesis.SMYD3 is a histone methylase that specifically targets histone methylation of H3K4,H4K5,and H4K20,and acts as a transcription factor to regulate gene transcription and participate in signal transduction processes,it was an importance factor in cancer development and progression.Firstly,RNAi technology was used to knockdown endogenous MALAT1/NEAT1 in MCF-7 and MDA-MB-231 cells.The migration and proliferation of breast cancer cells was detected by the wound healing assay and MTT assay.The result show that interference of MALAT1/NEAT1 inhibit the migration and proliferation of breast cancer cells.At same time,Real-Time PCR and Western blot show that the transcriptional expression of SMYD3 and its downstream target genes N-cahderin,MMP9,MYL9 and CYR61 were inhibited after MALAT1/NEAT1 were interfered,indicating that MALAT1 and NEAT1 induce SMYD3 transcriptional expression.In addition,after treatment of MCF-7 cells with 0,2.5,5,10 ng/mL of TGF-β1,Real-Time PCR results showed that the expression of NEAT1,MALAT1 and SMYD3 increased with the addition of TGF-β1,further After treatment with 2.5 ng/mL of TGF-β1 and si-NEAT1,Western blot analysis showed that knockdown of NEAT1 inhibited the induction of SMYD3 expression by TGF-β1,indicating that MALAT1 and NEAT1 are involved in the regulation of TGF-β1 on SMYD3.Next,the bioinformatics website predictive analysis showed that miR-506/124 has binding sites in MALAT1/NEAT1 and SMYD3 3’UTR.Studies have shown that transfection of miR-506/124 mimics down-regulates MALAT1/NEAT 1 and SMYD3 expression,conversely,miR-506/124 inhibitor upregulate the expression of M AL AT1/NEAT1 and SMYD3.In addition,after RNAi technology inhibited MALAT1/NEAT1,miR-506/124 was up-regulated.Furthermore,MALAT1/NEAT1 siRNA and miR-506/124 inhibitors were co-transfected to detect the effect of SMYD3 expression on MCF-7 and MDA-MB-231 cells,and the results showed that the interference of MALAT1/NEAT1 attenuated the promotion of SMYD3 transcription and translation by miR-506/124 inhibitors.These results indicate that MALAT1/NEAT 1 regulates the expression of SMYD3 as a competitive endogenous RNA(ceRNA)of miR-506/124 in breast cancer.Finally,Real-Time PCR and luciferase reporter analysis also found that SMYD3 also has a certain transcriptional activation effect on MALAT1/NEAT1,indicating that there is a positive feedback circular transcriptional regulation mechanism between each other.In conclusion,LncRNA MALAT1/NEAT1 acted as a competing endogenous RNA(ceRNA)of miR-506/124 to regulate expression of SMYD3 in breast cancer,and SMYD3 can activate the transcription of MALAT1,which will affect the proliferation and migration of breast cancer cells.This study provides some new ideas for the prevention and treatment of breast cancer.