The Mechanistic Study On The Proliferation-promoting Role Of LncRNA DLGAP1-AS2 In Colorectal Cancer

Objective:This study mainly studied the expression and function of long non-coding RNA DLGAP1-AS2 in colorectal cancer(CRC)and preliminarily explored its molecular mechanism.Methods:Real time fluorescence quantitative reverse transcription PCR(qRT-PCR)was used to detect the expression of DLGAP1-AS2 in CRC tissues and cells.CCK-8 and plate clone formation experiments were used to detect the effect of DLGAP1-AS2 on proliferation of CRC cells.The effect of DLGAP1-AS2 on the migration and invasion of CRC cells was analyzed by Transwell assay.The effect of DLGAP1-AS2 on the in vivo tumor growth was studied by animal subcutaneous tumor transplantation experiment.Based on RNA pull down,mass spectrometry and literature analyses,the binding proteins of DLGAP1-AS2 in CRC cells were screened.Western Blot and RNA immunoprecipitation(RIP)techniques were perforemed to further verify the target proteins of DLGAP1-AS2.Result:The expression of DLGAP1-AS2 was up-regulated in CRC tissues compared with the adjacent non-cancerous tissues(P<0.0001)and the high expression of DLGAP1-AS2 was related to the poor prognosis of patients(P = 0.038).Knocking down DLGAP1-AS2 inhibited proliferation,migration and invasion of CRC cells whereas overexpression of DLGAP1-AS2 promoted proliferation,migration and invasion of CRC cells.Animal experiments proved that DLGAP1-AS2 knock-down inhibited tumor growth and overexpression of DLGAP1-AS2 promoted tumor growth.RNA pull down experiment proved that DLGAP1-AS2 could bind to EloginA and CSTF3 proteins.Silencing DLGAP1-AS2 expression could improve the protein expression level of EloginA,and overexpression of DLGAP1-AS2 could reduce the protein expression level of EloginA.Overexpression of DLGAP1-AS2 shortened the half-life of EloginA and reduced the protein stability of EloginA;the degradation of EloginA was inhibited after DLGAP1-AS2 was knocked out.Knocking down.Knock-down CSTF3 reduced the gene expression level of DLGAP1-AS2.Knock-down CSTF3 shortened the half-life of DLGAP1-AS2 and reduced the gene stability of DLGAP1-AS2.Conclusion:DLGAP1-AS2 plays an oncogene role in CRC;DLGAP1-AS2 can promote the proliferation,migration and invasion of CRC cells;DLGAP1-AS2 can specifically bind EloginA and CSTF3 proteins,and can regulate the protein stability of EloginA and promote its degradation.CSTF3 can regulate the stability of DLGAP1-AS2 and further affect its degradation of the downstream target protein EloginA.