Establishment And Identification Of Human Tudor-SN Knock-down Breast Cancer Cell Line MDA-MB-231

Objective:The human Tudor-SN protein is a vital transcription regulator that increases gene transcription by forming a physical bridge between promoter-specific activators and the basal transcription machinery. Recently, it was reported that the expression level of Tudor-SN was higher in breast cancer tissues than the normal breast tissues. Therefore, we used breast cancer cell line MDA-MB-231to further investigate the effect of Tudor-SN on bio-logical behaviors of breast cancer cells and its molecular mechanism.Methods:1. Transfect separately the constructed plasmids pGenesil-shRNA-hTudor-SNI in to MDA-MB-231cells, Screen the stable cell lines.2. Transwell model was applied for invasion of pGenesil-shRNA-hTudor-SNI、 untreated human breast cancer cells in vitro.3. Wound-healing experiment was applied for migration of pGenesil-shRN A--hTudor-SNI、untreated human breast cancer cells in vitro.4. PCR and Western blot were applied for detecting the level of mRNA and protein of Tudor-SN in different group:Tudor-SN stable knockdown and blank control group in both cells and tissues.6. PCR and Western blot were applied for detecting the level of mRNA of MMP-2and MMP-9in different group:Tudor-SN stable knock-down and untransfected group in both cells and tissues.Results:②Tudor-SN mRNA and protein expression levels in stable knock-down cell lines were obviously down-regulated, as compared with that in negative control group and untransfected group. The stable MDA-MB-231-Tudor-SNI cell lines were screened and indentified successfully.②Tudor-SN protein could promote the invasion of MDA-MB-231cells.③Tudor-SN protein could promote the migration of MDA-MB-231cells.④MMP-2mRNA levels in both stable knock-down cell lines and mice Tudor-SN stable knock-down breast cancer tissues were obviously down-regulated compared with untransfected groups. Interestingly, MMP-9mRNA levels in both Tudor-SN stable knock-down cell lines and mice Tudor-SN stable knock-down breast cancer tissues were obviously up-regulated compared with untransfected groups.Conclusion:①The stable MDA-MB-231-Tudor-SNI cell lines were screened and identified successfully.②Tudor-SN protein could promote the invasion of MDA-MB-231cells.③Tudor-SN protein could promote the migration of MDA-MB-231cells.④Tudor-SN protein improved mRNA levels of MMP-2,but decreased mRNA levels of MMP-9both in breast cancer cells and mice breast cancer tissues.