Activation and Cross-talk between Akt, NF-κB, and Unfolded Protein Response Signaling in 1-LN Prostate Cancer Cells Consequent to Ligation of Cell Surface-associated GRP78*

Binding of activated forms of the proteinase inhibitor α2-macroglobulin (α2M*) to cell surface-associated GRP78 on 1-LN human prostate cancer cells causes their proliferation. We have now examined the interplay between Akt activation, regulation of apoptosis, the unfolded protein response, and activation of NF-κBin α2M*-induced proliferation of 1-LN cells. Exposure of cells to α2M* (50 pm) induced phosphatidylinositol 3-kinase-dependent activation of Akt by phosphorylation at Thr-308 and Ser-473 with a concomitant 60-80% increase in Akt-associated kinase activity. ERK1/2 and p38 MAPK were also activated, but there was only a marginal effect on JNK activation. Treatment of 1-LN cells with α2M* down-regulated apoptosis and promoted NF-κB activation as shown by increases of Bcl-2, p-BadSer-136, p-FOXO1Ser-253, p-GSK3βSer-9, XIAP, NF-κB, cyclin D1, GADD45β, p-ASK1Ser-83, and TRAF2 in a time of incubation-dependent manner. α2M* treatment of 1-LN cells, however, showed no increase in the activation of caspase -3, -9, or -12. Under these conditions, we observed increased unfolded protein response signaling as evidenced by elevated levels of GRP78, IRE1α, XBP-1, ATF4, ATF6, p-PERK, p-eIF2α, and GADD34 and reduced levels of GADD153. Silencing of GRP78 gene expression by RNAi suppressed activation of AktThr-308, AktSer-473, and IκB kinase α kinase. The effects of α2M* on the NF-κB activation, antiapoptotic signaling, unfolded protein response signaling, and proapoptotic signaling were also reversed by this treatment. In conclusion, α2M* promotes cellular proliferation of 1-LN prostate cancer cells by activating MAPK and Akt-dependent signaling, down-regulating apoptotic signaling, and activating unfolded protein response signaling.