The neuro-steroid, 5-androstene 3beta,17alpha diol; induces endoplasmic reticulum stress and autophagy through PERK/eIF2alpha signaling in malignant glioma cells and transformed fibroblasts.

摘要

In this study, we identified a mechanism by which the neuro-steroid, 5-androstene 3beta,17alpha diol (17alpha-AED) induces autophagy in human malignant glioma cells and transformed fibroblasts. 17alpha-AED treatment induced endoplasmic reticulum (ER) stress, identified by the partial activation of an unfolded protein response in T98G, U87MG, U251MG, LN-18, LN-229 and LN-Z308 glioma cell lines. In this regard, there were increased levels of CCAAT/enhancer-binding protein homologous protein (CHOP) and glucose-regulated protein of 78kDa transcripts but no splicing of X-box-binding protein 1 mRNA or processing of activating transcription factor-6 in glioma cells treated with the neuro-steroid. 17alpha-AED induced eukaryotic translational initiation factor 2alpha (eIF2alpha) phosphorylation in glioma cells which correlated with microtubule-associated protein-light chain 3 (LC3) conversion from LC3-I to -II. In transformed murine embryonic fibroblasts (MEFs) that are deficient of eIF2alpha function or T98G glioma cells transfected with a dominant-negative eIF2alpha construct, 17alpha-AED induced LC3 conversion was significantly reduced as compared to control cells. Neuro-steroid treatment caused the activation of the eIF2alpha kinase, protein kinase-like ER kinase (PERK) but not other eIF2alpha kinases in glioma cells. Moreover, eIF2alpha phosphorylation and LC3 conversion, in response to 17alpha-AED treatment, was blocked in MEFs that lacked PERK activity. T98G cells transfected with a dominant-negative PERK construct exhibited an attenuated response to neuro-steroid treatment in terms of decreases in: eIF2alpha activation; CHOP expression; the incidence of autophagy; and cytotoxicity. These results demonstrate that ER stress is linked to 17alpha-AED induced autophagy by PERK/eIF2alpha signaling in human malignant glioma cells and transformed fibroblasts.