Graduation Date

Summer 8-18-2017

Document Type


Degree Name

Doctor of Philosophy (PhD)


Genetics, Cell Biology & Anatomy

First Advisor

Dr. Band Vimla


Tumor cells are well known to exhibit ER stress as a result of their altered environment characterized by redox and calcium imbalance, deregulation of protein synthesis to meet their oncogenic demands, and decreased vascularization associated with nutrient limitation and hypoxia, all of which are conducive of ER stress. Accordingly, markers of ER stress signaling response are up-regulated in various cancers. The outcome of ER stress signaling response varies from survival and adaption to apoptosis. Although pro-apoptotic ER stress signaling molecules are up-regulated in cancer, the mechanisms by which cancer cells seem refractory to, or evade, their apoptotic signaling are not fully understood. In this study, we investigate roles of the mammalian Ecdysoneless (ECD) in ER stress and oncogenesis. Certain tumors overexpress ECD protein suggesting possible roles in oncogenesis. In fact, both normal and cancer cells require ECD for their growth and undergo cell cycle arrest when the ECD level is deficient. ECD associates with various stress response proteins such as p53, well known for genotoxic stress response such as DNA damage, and TXNIP, an oxidative stress protein recently implicated in ER stress response. Furthermore, ECD is a critical component of chaperone-like complexes such as the R2TP complex whose members have ER stress-related roles. In this study, we provide evidence that ECD is involved in ER stress.

Stress induction by multiple stimuli, including Thapsigargin, Tunicamycin, glucose starvation, H2O2, NOX4 overexpression, led to reduced ECD protein levels, but ECD mRNA increased, in a PERK-eIF2α dependent manner. Although a phospho-protein, ECD is not a target for a phosphorylation-mediated degradation by PERK but is rather targeted for a translation block via the PERK-eIF2α axis. To assess the functional connection between ECD and ER stress signaling pathways, we used cells in which ECD could be inducibly depleted or overexpressed. Depletion of ECD enhanced PERK signaling and apoptosis upon ER stress induction while overexpression of ECD produced the opposite effect by inhibiting PERK signaling and increasing cell survival. IRE1α signaling was slightly affected by these changes in the cellular level of ECD, as indicated by the slight increase or decrease of its downstream target, spliced XBP-1, upon ER stress induction in the presence or absence of ECD; on the other hand, the ATF6 pathway was minimally affected.

Based on these findings, we examined the possible mechanism by which ECD regulates ER stress signaling, particularly the PERK pathway. We found that ECD co-localized and associated with all the ER stress sensors PERK, IRE1α, ATF6 and GRP78. However, ECD does not modulate the enzymatic activity of PERK toward its substrate eIF2α. Rather, ECD enhanced chaperones’ levels, predominantly, GRP78 upon ER stress induction. Finally, disruption of this chaperone-enhancing effect of ECD abrogated the attenuating effect of ECD on PERK and impaired the pro-survival effect of overexpressed ECD upon ER stress induction. Taken together, ECD regulates the levels of chaperones, predominantly, GRP78 to enhance the folding capacity of the stressed ER and provide survival advantage in a stress condition.

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