Effects of HTLV-1 tax and CREB/ATF family of transcription factors on CMV MIE promoter in HEK293 cells.

• Main Author: Ko, Chen Wei.
• Other Authors: Chao Shu Ting Edlyn
• Format: Final Year Project
• Language: English
• Published: 2009
• Subjects: DRNTU::Science
• Online Access: http://hdl.handle.net/10356/18035
• Institution: Nanyang Technological University

The Human T-Lymphotropic virus type 1 (HTLV-1) genome codes for a 40 kD viral protein Tax which can trans activate the human cytomegalovirus (CMV) major immediate-early (MIE) enhancer in Jurkat T cells. The CMV MIE promoter is one of the strongest RNA polymerase II identified and is used industrially for recombinant protein production. It is envisaged that a Tax expressing cell line could be used to increase CMV MIE promoter activity for recombinant protein production. Tax was cloned into pcDNA6/myc-His with a selection marker blasticidin-resistance gene for the generation of Tax stable mammalian cell lines. The plasmid was expressed in Human Embroyonic Kidney 293 (HEK293) cells and induced a 10-fold and 50-fold increase in CMV-Luciferase (CMV-Luc) and HTLV-Long Terminal Repeat (LTR)-Luciferase (HTLV-LTR-Luc) activity respectively. The effects of Tax interacting transcription factors which may increase CMV MIE promoter activation were also investigated. 4 members of the cyclic-AMP-responsive element binding protein/activating transcription factors (CREB/ATF) family of transcription factors, namely the X-box binding protein 1 (XBP-1), its spliced form XBP-1S, CREB1 and CREB2 were overexpressed in HEK293 cells in the presence and absence of Tax. CREB1 was found to inhibit CMV MIE promoter activity by 2 fold.