Effects of Pluronic F-68 on hCTLA4Ig production in transgenic rice cell suspension cultures

초록

Transgenic plant cell cultures have been used as a production system for therapeutic proteins. Human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig), a fusion protein, is an effective immunosuppressive agent. The production and secretion of hCTLA4Ig in rice (Oryza sativa L.) cell suspension cultures were investigated in this study. Rice cells were transformed to express hCTLA4Ig using RAmy3D promoter. With the RAmy3D promoter system, expression of the target protein could be strongly induced by sugar starvation. However, sugar-depleted environment can cause cell death and secretion of proteases into the medium due to cell lysis. Various stabilizing agents such as polyethylene glycol (PEG), gelatin, bovine serum albumin, and polyvinylpyrrolidone (PVP) have been used to reduce the degradation of secreted proteins in plant cell cultures. Pluronic F-68 (PF-68) and Tween 20, nonionic surfactants, are widely used not only to protect cells from shear stress in bioreactor but also to improve the secretion of the target protein in mammalian cell cultures. To enhance the production of hCTLA4Ig in transgenic rice cell suspension cultures, PF-68 was added at different concentrations (0, 1, 3, 5, 7 g/L) to the media during induction phase. With the presence of PF-68, the cell viability and hCTLA4Ig production were improved compared to those in control cultures. In conclusion, optimal supplementation of PF-68 showed protective effects against shear stress and enhanced hCTLA4Ig production in transgenic rice cell suspension cultures.