Stress protein GRP78 prevents apoptosis induced by calcium ionophore, ionomycin, but not by glycosylation inhibitor, tunicamycin, in human prostate cancer cells

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Abstract
GRP78 induction has recently been shown to play a critical role in maintaining cell viability against several kinds of stress, including depletion of endoplasmic reticulum Ca2+ and accumulation of unglycosylated proteins, under specific experimental conditions. However, the functional significance of GRP78 induction after stressful treatment has not been well defined. This article characterizes the different biological features associated with GRP78 induction by two kinds of stress agents, calcium ionophore, ionomycin (IM), and glycosylation inhibitor, tunicamycin (TM), focusing on the association with apoptosis in human prostate cancer cells. Both IM and TM treatment resulted in marked induction of GRP78 transcription in androgen‐dependent prostate cancer LNCaP cells maintained in medium without androgen, but not in medium containing androgen, as measured by Northern blotting and nuclear run‐off assays. After pretreatment with tumor necrosis factor‐α, which has potent cytotoxic effects on LNCaP cells, both IM and TM could induce substantial increases in GRP78 transcription in LNCaP cells, even in medium containing androgen. Under both experimental conditions described, DNA fragmentation assays showed a direct correlation between the onset of apoptosis in LNCaP cells after IM treatment and the initiation of GRP78 transcript induction, while induction of GRP78 expression preceded TM‐induced apoptosis. To elucidate the functional differences of GRP78 induction by IM and TM, an antisense oligodeoxynucleotide (ODN) targeted against the grp78 gene was designed to reduce GRP78 expression in a sequence‐specific and dose‐dependent manner. Antisense GRP78 ODN treatment substantially enhanced apoptosis of LNCaP cells induced by IM compared with mismatch control ODN treatment, whereas no marked differences were observed in apoptotic features induced by TM with antisense GRP78 and mismatch control ODN treatment. Studies of additional androgen‐independent prostate cancer PC3 cells also demonstrated a correlation between GRP78 induction and resistance to apoptosis after IM treatment, but not after TM treatment. These findings suggest that there are at least two GRP78 signaling pathways, which play different roles in resistance against stress‐induced apoptosis. J. Cell. Biochem. 77:396–408, 2000.