Abstract
Death receptor 4 (DR4) is a cell surface receptor for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and triggers apoptosis upon ligation with TRAIL or aggregation. MEK/ERK signaling is a well known and the best-studied effector pathway downstream of Ras and Raf. This study focuses on determining the impact of pharmacological MEK inhibition on DR4 expression and elucidating the underlying mechanism. We found that several MEK inhibitors including MEK162, AZD6244, and PD0325901 effectively decreased DR4 protein levels including cell surface DR4 in different cancer cell lines. Accordingly, pre-treatment of TRAIL-sensitive cancer cell lines with a MEK inhibitor desensitized them to TRAIL-induced apoptosis. These results indicate that MEK inhibition negatively regulates DR4 expression and cell response to TRAIL-induced apoptosis. MEK inhibitors did not alter DR4 protein stability, rather decreased its mRNA levels, suggesting a transcriptional regulation. In contrast, enforced activation of MEK/ERK signaling by expressing ectopic B-Raf (V600E) or constitutively activated MEK1 (MEK1-CA) or MEK2 (MEK2-CA) activated ERK and increased DR4 expression; these effects were inhibited when a MEK inhibitor was present. Promoter analysis through deletion and mutation identified the AP-1 binding site as an essential response element for enhancing DR4 transactivation by MEK1-CA. Furthermore, inhibition of AP-1 by c-Jun knockdown abrogated the ability of MEK1-CA to increase DR4 promoter activity and DR4 expression. These results suggest an essential role of AP-1 in mediating MEK/ERK activation-induced DR4 expression. Our findings together highlight a previously undiscovered mechanism that positively regulates DR4 expression through activation of the MEK/ERK/AP-1 signaling pathway.
Highlights
Death receptor 4 (DR4),4 known as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor 1 (TRAIL-R1) or tumor necrosis factor receptor superfamily member 10A (TNFRSF10A), is a cell surface receptor that binds TRAIL and induces apoptosis
MEK Inhibition with MEK Inhibitors Substantially Decreases DR4 Levels in Cancer Cells—While working with the MEK inhibitor, MEK162, we found that at the tested concentration ranges (1 and 3 M), it effectively decreased the levels of p-ERK1/2 in several lung cancer cell lines, indicating the potent inactivation of ERK1/2
Our previous studies have demonstrated that death receptor 5 (DR5) expression is positively regulated by MEK/ERK signaling through increasing CHOP/Elk-dependent gene transcription [25, 26]
Summary
Death receptor 4 (DR4), known as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor 1 (TRAIL-R1) or tumor necrosis factor receptor superfamily member 10A (TNFRSF10A), is a cell surface receptor that binds TRAIL and induces apoptosis. Many agents, including some anticancer drugs, sensitize cancer cells to TRAIL-induced apoptosis through increasing the expression of DR4 and/or DR5 [7, 8]. DR4 does display distinct functions from DR5, such as in mediating apoptosis induced by certain stimuli [9, 10] and in the regulation of cancer cell invasion and metastasis [11], the underlying mechanisms are largely unknown. The MEK/ERK kinase cascade is a well known and the bestcharacterized effector pathway downstream of oncogenic RAS and RAF. Given our reported findings that RAS/RAF/MEK/ERK signaling positively regulates DR5 expression through enhancing CHOP/Elk1-mediated gene transcription [11, 25, 26], we explored in this study whether the MEK/ERK signaling pathway regulates DR4 expression and investigated the underlying mechanism
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