Activated α2-Macroglobulin Regulates Transcriptional Activation of c-MYC Target Genes through Cell Surface GRP78 Protein

Abstract

Activated ␣ 2 -macroglobulin (␣ 2 M*) signals predominantly through cell surface GRP78 (CS-GRP78) to promote proliferation and survival of cancer cells; however, the molecular mechanism remains obscure. c-MYC is an essential transcriptional regulator that controls cell proliferation. We hypothesize that ␣ 2 M*/CS-GRP78-evoked key signaling events are required for transcriptional activation of c-MYC target genes. Activation of CS-GRP78 by ␣ 2 M* requires ligation of the GRP78 primary amino acid sequence (Leu 98 -Leu 115 ). After stimulation with ␣ 2 M*, CS-GRP78 signaling activates 3-phosphoinositide-dependent protein kinase-1 (PDK1) to induce phosphorylation of PLK1, which in turn induces c-MYC transcription. We demonstrate that PLK1 binds directly to c-MYC and promotes its transcriptional activity by phosphorylating Ser 62 . Moreover, activated c-MYC is recruited to the E-boxes of target genes FOSL1 and ID2 by phosphorylating histone H3 at Ser 10 . In addition, targeting the carboxyl-terminal domain of CS-GRP78 with a mAb suppresses transcriptional activation of c-MYC target genes and impairs cell proliferation. This work demonstrates that ␣ 2 M*/CS-GRP78 acts as an upstream regulator of the PDK1/PLK1 signaling axis to modulate c-MYC transcription and its target genes, suggesting a therapeutic strategy for targeting c-MYC-associated malignant progression.␣ 2 -Macroglobulin (␣ 2 M) 2 is a plasma protein that interacts with and entraps virtually all proteinases, thereby blocking access to their substrates (1). In prostate cancer patients, ␣ 2 M is proteolytically activated (␣ 2 M*) and signals predominantly through interaction with cell surface GRP78 (K d ϳ 50 -100 pM), promoting proliferation and survival of cancer cells (2, 3). GRP78 is a stress-inducible, prosurvival, endoplasmic reticulum chaperone belonging to the HSP70 family. It is composed of an ATPase domain, a peptide binding domain, and a COOHterminal domain of unknown function (4 -6). Several different cell types, including proliferating endothelial cells and tumor cells, express GRP78 on their surface (7-15). GRP78 expression at the cell surface and its ligation by ␣ 2 M* are clearly implicated in the development of metastatic prostate cancer (2, 9, 16 -19). However, the mechanism by which ␣ 2 M*/cell surface GRP78 (CS-GRP78) signaling regulates gene transcription and their responses in cell proliferation is unknown.CS-GRP78 is a multifunctional receptor that forms complexes with phosphatidylinositol 3-kinase (PI3K) and enhances phosphatidylinositol 3,4,5-trisphosphate production, consistent with its novel role as a regulator of the PI3K/Akt signaling pathway. Thus it promotes cell proliferation, survival, metastasis, and chemoresistance (9, 20 -22). CS-GRP78, through its NH 2 -terminal domain, drives PI3K/Akt activity (2), whereas targeting the COOH-terminal domain with antibody promotes apoptotic signaling (21,23). Recently, we demonstrated in vivo that targeting the GRP78 COOH-terminal domain with monoclonal antibody C38 (C38 mAb) delays tumor gro...

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