Presentation Type
Poster
Faculty Mentor’s Full Name
Jesse Hay
Faculty Mentor’s Department
Division of Biological Sciences
Abstract / Artist's Statement
Regulatory mechanisms of protein trafficking from the endoplasmic reticulum (ER) are critical to
understand since neurodegenerative diseases involve defects in this process leading to chronic
ER stress and cell death. This study aimed to better understand the calcium regulatory mechanisms of ER-to-Golgi body complex trafficking in hybrid neuroglioblastoma cells (NG108). Specifically, we asked whether proteosomal degradation of transport machinery was involved in the previously demonstrated upregulation of ER-to-Golgi transport evoked by calcium signaling. This was accomplished by NG108 transfection with a fluorescent cargo protein engineered to allow inducible, synchronous trafficking from the ER. Cargo transport was completed for a fixed time and the cells were subsequently fixed and immunolabeled for detection of the Golgi complex. Cells were imaged on a widefield microscope and the images were analyzed using ImageJ software to quantitate the transport index, a measure of the speed of cargo transport. Prior to transport cells were treated with a calcium agonist and/or the proteasome inhibitor mg132. Results indicated that Ca2+ signaling caused an increase in cargo transport, as expected. Importantly, inhibition of the proteasome caused a decrease in cargo transport, but treatment with both a Ca2+ agonist and the proteasome inhibitor caused an increase compared to treatment with just the proteasome inhibitor. Based upon previous literature, we believe that the Ca2+ induced increase in transport is due to the activation of the Ca2+ sensitive regulatory protein apoptosis linked gene 2 (ALG-2) and its interactions with the inhibitory regulatory protein peflin. A potential mechanism is that after Ca2+ induced ALG-2 dissociation from peflin, peflin is degraded by the proteasome and the removal of its inhibitory effects cause the increase in transport. Our new results show that proteasomal activity is required for optimum transport but is not involved in the calcium stimulation of transport per se. This suggests that other, yet unknown mechanisms drive the Ca2+ and ALG-2-mediated regulation of transport.
Category
Life Sciences
Calcium regulation of ER to Golgi protein transport in neuronal cells
UC South Ballroom
Regulatory mechanisms of protein trafficking from the endoplasmic reticulum (ER) are critical to
understand since neurodegenerative diseases involve defects in this process leading to chronic
ER stress and cell death. This study aimed to better understand the calcium regulatory mechanisms of ER-to-Golgi body complex trafficking in hybrid neuroglioblastoma cells (NG108). Specifically, we asked whether proteosomal degradation of transport machinery was involved in the previously demonstrated upregulation of ER-to-Golgi transport evoked by calcium signaling. This was accomplished by NG108 transfection with a fluorescent cargo protein engineered to allow inducible, synchronous trafficking from the ER. Cargo transport was completed for a fixed time and the cells were subsequently fixed and immunolabeled for detection of the Golgi complex. Cells were imaged on a widefield microscope and the images were analyzed using ImageJ software to quantitate the transport index, a measure of the speed of cargo transport. Prior to transport cells were treated with a calcium agonist and/or the proteasome inhibitor mg132. Results indicated that Ca2+ signaling caused an increase in cargo transport, as expected. Importantly, inhibition of the proteasome caused a decrease in cargo transport, but treatment with both a Ca2+ agonist and the proteasome inhibitor caused an increase compared to treatment with just the proteasome inhibitor. Based upon previous literature, we believe that the Ca2+ induced increase in transport is due to the activation of the Ca2+ sensitive regulatory protein apoptosis linked gene 2 (ALG-2) and its interactions with the inhibitory regulatory protein peflin. A potential mechanism is that after Ca2+ induced ALG-2 dissociation from peflin, peflin is degraded by the proteasome and the removal of its inhibitory effects cause the increase in transport. Our new results show that proteasomal activity is required for optimum transport but is not involved in the calcium stimulation of transport per se. This suggests that other, yet unknown mechanisms drive the Ca2+ and ALG-2-mediated regulation of transport.