Effect of Bcl‐x_L overexpression on apoptosis and autophagy in recombinant Chinese hamster ovary cells under nutrient‐deprived condition

Abstract

Upon nutrient deprivation during culture, recombinant Chinese hamster ovary (rCHO) cells are subjected to two types of programmed cell death (PCD), apoptosis and autophagy. To investigate the effect of Bcl-x(L) overexpression on apoptosis and autophagy in rCHO cells, an erythropoietin (EPO)-producing rCHO cell line with regulated Bcl-x(L) overexpression (EPO-off-Bcl-x(L)) was established using the Tet-off system. The expression level of Bcl-x(L) in EPO-off-Bcl-x(L) cells was tightly regulated by doxycycline in a dose-dependent manner. Bcl-x(L) overexpression enhanced cell viability and extended culture longevity in batch culture. Upon nutrient depletion in the later stage of batch culture, Bcl-x(L) overexpression suppressed apoptosis by inhibiting the activation of caspase-3 and -7. Simultaneously, Bcl-x(L) overexpression also delayed autophagy, characterized by LC3-II accumulation. Immunoprecipitation analysis with a Flag-tagged Bcl-x(L) revealed that Bcl-x(L) interacts with Bax and Bak, essential mediators of caspase-dependent apoptosis, as well as with Beclin-1, an essential mediator of autophagy, and may inhibit their pro-cell death function. Taken together, it was found that Bcl-x(L) overexpression inhibits both apoptosis and autophagy in rCHO cell culture.

Autophagy is a cell survival program for female germ cells in the murine ovary

“…In vitro abrogation of the BECN1-BCL2/BCL2L1 interaction with the pharmacological mimetic ABT737 triggers autophagy by reducing available BCL2/BCL2L1 (Maiuri et al 2007). Similarly, Bcl2 siRNA in MCF-7 cells induces autophagy, whereas overexpression of BCL2L1 in CHO cells suppressed both apoptosis and autophagy (Akar et al 2008, Kim et al 2009). However, BECN1 and BCL2 co-expression in HeLa cells did not demonstrate an effect of BECN1 on inhibiting the anti-apoptotic effects of BCL2 (Ciechomska et al 2009).…”

Hyperosmotic stress induces autophagy and apoptosis in recombinant Chinese hamster ovary cell culture

“…The CHO-EPO was previously established in our laboratory (Kim et al, 2009). The CHO-Ab was provided by GenexelSein (Daejeon, Korea).…”

Overexpression of survivin and cyclin D1 in CHO cells confers apoptosis resistance and enhances growth in serum-free suspension culture

“…Metabolically-engineered CHO cells may, though, have enhanced proliferation and antiapoptosis performance and a decreased serum dependency (Fussenegger and Betenbaugh 2002). For instance, overexpression of bcl-2 (Lee and Lee 2003) and bcl-x L (Kim et al 2009) and the constitutive activation of Akt (Hwang and Lee 2009), efficiently inhibited apoptosis in CHO cells, and bcl-x L and Akt inhibited cell autophagy (Kim et al 2009;Hwang and Lee 2009). In addition, overexpression of hTERT reduced serum dependency and conferred resistance to serum deprivation-induced apoptosis (Crea et al 2006).…”