Construction of mutants of MHC class II molecule transactivator gene capable of suppressing HLA-DR/DQ expression and exploration on their mechanism

• Published in: Xi bao yu fen zi mian yi xue za zhi Vol. 19; no. 5; p. 417
• Main Authors: Ou, Qi-shui, Lin, Lin, Huang, Li-dong, Lu, Pei-hua, Zhou, Guan-yan
• Format: Journal Article
• Language: Chinese
• Published: China 01.09.2003
• Subjects: Gene Expression Regulation; HeLa Cells; HLA-DQ Antigens - genetics; HLA-DR Antigens - genetics; Humans; Interferon-gamma - biosynthesis; Mutation; Nuclear Proteins - genetics; Trans-Activators - genetics; Transfection
• ISSN: 1007-8738

To construct MHC class II transactivator (CIITA) gene mutants which could suppress the expression of MHC class II molecule and explore its mechanism. Restrictive enzymes digestion, PCR and synthesized oligonucleotide strands were used to construct three mutants including pcDNA3mCIITA2, pcDNA3mCIITA3 and pcDNA3mCIITA4. These mutants and pcDNA3 vector were transfected into Hela and Raji cell lines by lipofectamine. RT-PCR and flow cytometry were used to determine the changes of the inducible/constitutive expression of MHC class II-molecule. The mCIITA4 mutant was transferred to the tetracycline dose-dependent plasmid pUHD10-3. By adjusting the concentration of Dox in cultural circumstances, and regulating expressed level of external mutant C II TA, the relationship between the expression level of mC II TA4 mutant and that of MHC class II molecule was observed. pcDNA3mCIITA3 and pcDNA3mCIITA4 could significantly suppress the expression of HLA-DR/DQ gene in Hela and Raji cells, while pcDNA3 empty vector and pcDNA