Abstract B3: Potential new mechanism whereby IGFBP-5 contributes to cancer progression: Binding of GADD34 and enhancement of eIF2α dephosphorylation

Abstract

Abstract Insulin-like Growth Factor Binding Protein (IGFBP-5) has a well-defined role in normal mammary gland physiology and has recently been shown to play an important role in the biology of breast cancer; however the IGF-independent mechanisms of action remain obscure. Over-expression of IGFBP-5, but not the addition of the exogenous protein, has been shown to inhibit the growth of breast cancer cells in vitro. This suggests novel intracellular mechanisms where IGFBP-5 interacts with unidentified binding partners and thereby modulates the proliferation and survival of breast cancer cells. To address this issue, we performed a yeast two-hybrid screen utilizing a normal human mammary gland cDNA library and full-length IGFBP-5 as the bait. The pre-eminent binding partner identified was Growth Arrest and DNA Damage-inducible protein 34 (GADD34), which assembles an eukaryotic initiation factor 2 alpha subunit (eIF2α) dephosphorylation complex at the endoplasmic reticulum (ER). We have confirmed that IGFBP-5 and GADD34 interact through coimmunoprecipitation experiments using ectopically expressed IGFBP-5 and GADD34 in mammalian cells. We have also shown that IGFBP-3, which belongs to the same protein family as IGFBP-5, interacts with GADD34 through coimmunoprecipitation studies. Furthermore, IGFBP-5 or IGFBP-3 expression was found to dramatically increase GADD34 protein levels as demonstrated through immunoblot analysis. However, IGFBP-2, another protein related to IGFBP-5, did not interact with GADD34 or increase its protein expression. Because GADD34-mediated eIF2α dephosphorylation is required to promote resumption of protein translation after periods of ER stress, we next examined eIF2α phosphorylation levels in the presence and absence of IGFBPs. Expression of IGFBP-5 or IGFBP-3 decreased the levels of phosphorylated eIF2α, suggesting that the interactions diminish the magnitude and/or duration of ER stress-induced inhibition of translation. Our results demonstrate not only a physical interaction between IGFBP-5 or IGFBP-3 and GADD34, but also show that these IGFBPs appear to increase the stability and activity of GADD34. This discovery suggests a novel role for IGFBP-5 and IGFBP-3 in the regulation of a protective mechanism induced by ER stress. Since ER stress is one of the hallmarks of solid tumor progression, by modifying the tumor response to hostile microenvironments, it is likely that these IGFBPs can have significant impact on cancer growth, survival, and apoptosis. Citation Information: Cancer Res 2009;69(23 Suppl):B3.