Receptor-recognized forms of α₂ -macroglobulin (α₂ M) bind to cell surface-associated GRP78 and initiate pro-proliferative and anti-apoptotic signaling. Ligation of GRP78 with α₂ M also upregulates TFII-I, which binds to the GRP78 promoter and enhances GRP78 synthesis. In addition to its transcriptional functions, cytosolic TFII-I regulates agonist-induced Ca(2+) entry. In this study we show that down regulation of TFII-I gene expression by RNAi profoundly impairs its cell surface expression and anti-apoptotic signaling as measured by significant reduction of GRP78, Bcl-2, and cyclin D1 in 1-Ln and DU-145 human prostate cancer cells stimulated with α₂ M. In contrast, this treatment significantly increases levels of the pro-apoptotic proteins p53, p27, Bax, and Bak and causes DNA fragmentation. Furthermore, down regulation of TFII-I expression activates agonist-induced Ca(2+) entry. In plasma membrane lysates p-PLCγ1, TRPC3, GRP78, MTJ1, and caveolin co-immunoprecipitate with TFII-I suggesting multimeric complexes of these proteins. Consistent with this hypothesis, down regulating TFII-I, MTJ1, or GRP78 expression by RNAi greatly attenuates cell surface expression of TFII-I. In conclusion, we demonstrate that not only does cell surface GRP78 regulate apoptosis, but it also regulates Ca(2+) homeostasis by controlling cell surface localization of TFII-I.