Enhanced viability of a nervous necrosis virus-infected stable cell line over-expressing a fusion product of the zfBcl-x _L and green fluorescent protein genes

Nervous necrosis virus (NNV) infection induces host cell apoptosis by an ill-understood process. We utilized a fusion between enhanced green fluorescent protein (EGFP) and the zfBcl-xL gene in GL-av cells to select for zfBcl-xL stable cell lines and to assess the effectiveness of the anti-apoptotic protein Bcl-xL in circumventing NNV-induced cell death. Stable EGFP and EGFP-Bcl-xL-expressing clones were obtained at high purity within 2.5-3 months. In the latter, the EGFP-Bcl-xL fusion protein (approximately 58.2 kDa, as ascertained by Western blot) was predominantly targeted to mitochondria. We assayed for apoptosis in red-spotted grouper NNV Tainan no. 1 (RGNNV TN1)-infected cells with terminal deoxynucleotidyl transferase (TdT)-mediated end labelling (TUNEL) of DNA at different virus doses. NNV infection of NNV Bcl-xL GL-av cell line revealed a protective effect, with a decrease in TUNEL-positive cells of 7%, 8% and 31.8% at 24, 48 and 72 h, respectively. In addition, RGNNV infection of the Bcl-xL GL-av cell line revealed a protective effect, with an enhanced viability of 3%, 40% and 73% at 24, 48, and 72 h, respectively. We conclude that NNV-induced apoptotic cell death can be lessened in transgenic grouper fish cells.