2.3  Transient transfection and reporter gene assays
Transfection of U937 cells with an LPL promoter-luciferase DNA construct (− 31 to + 187) and CMV-β-galactosidase (internal control for transfection efficiency) was carried out using SuperFect™ (Qiagen) as previously described [9,15]. In experiments involving the use of dominant negative (DN) constructs, the cells were initially transfected with the DN expression plasmid or control vector and incubated for 8 h before transfection of the cells with the LPL promoter-luciferase and CMV-β galactosidase plasmids. The DN plasmids used were pcDNA3 HA-PKB AAA from Dr. B. Hemmings (Friedrich Miescher Institute for Biomedical Research, Basel) and pSG-CK2α-K68A from Drs. E. M. Chambaz and C. Cochet (INSERM, Grenoble). DN PKB specifies for an inactive form of PKBα with a mutation to alanine of lysine 179 in the ATP-binding site and threonine 308 and serine 473 that must be phosphorylated in the active kinase. DN CK2 codes for a kinase inactive mutant with a lysine to alanine change at residue 68 within the ATP-binding domain.