FasL induces Fas/Apo1-mediated apoptosis in human embryonic kidney 293 cells routinely used to generate E1-deleted adenoviral vectors

Abstract

Human embryonic kidney 293 cells contain the E1 region E1-E3 regions, in an unpackagable form. Investigation of of adenovirus type 5, and thus sustain, through transcompthe reason for massive cell death after cotransfection led lementation, the production of recombinant E1-deleted us to determine that 293 cells express the FasL receptor, adenovirus vectors. During attempts to produce recombiFas-Apo1 (CD95), and respond with apoptosis to the nant adenovirus expressing the apoptosis-inducing molcross-linking of Fas-Apo1 with either IgM monoclonal antiecule Fas ligand (FasL) under the control of a very strong bodies or FasL. Therefore, we decided to generate adenotruncated major immediate-early human cytomegalovirus viral vectors expressing FasL under the control of tissue-(MIEhCMV) promoter, we discovered that 293 cells were specific and/or -inducible promoter elements. Our findings not surviving the initial cotransfection with a shuttle plasmid can explain difficulties several groups have had in generatencoding the mouse FasL; and pJM17, a plasmid containing recombinant adenoviral vectors expressing FasL using ing the genome of adenovirus type 5 with deletions in the 293 cells, as well as the lower titres reported.

A binary adenoviral vector system for expressing high levels of the proapoptotic gene bax

“…For example, constructing adenoviral vectors expressing the fas ligand (Fas-L) has been reported to be difficult because of the toxic effect on 293 cells. 3 Although adenoviral vectors expressing Fas-L have been constructed and proven to be effective in the treatment of established experimental tumor models, 293 cells that are resistant to Fas-L-induced apoptosis or the caspase inhibitor were used in that particular study to produce such a vector. 4 Alternatively, a stuffer DNA fragment flanked by lox P sites was placed between promoter and Fas-L cDNA to prevent transgene expression in 293 cells.…”

A novel strategy for in vivo expansion of transplanted hepatocytes using preparative hepatic irradiation and FasL-induced hepatocellular apoptosis

“…This system was required because the adenovirus generating 293 cells express Fas constitutively and are killed upon expression of FasL. 20,21 In our experiments, the desired level of Fas ligand expression was obtained by adjusting the dose of the Cre-recombinase-expressing vector, AxCANCre. Pilot experiments were performed to determine the optimum dose of AxCALNCre.…”

Development of an inducible suicide gene system based on human caspase 8

“…Some preliminary tests were performed by transient transfection in 293T cells, a transformed human kidney cell line extensively used in apoptosis studies. [24][25][26] We transferred either a CC8 or a CD8 encoding plasmid, together with a GFP tracer, and observed that cells cotransfected with the CD8 and the GFP expression vectors did not display any phenotypic differences as compared to cells transfected with GFP alone (data not shown). This indicates that the CD8 transmembrane domain used to bring the caspase 8 domain to the membrane does not affect the transfected cells.…”