The Role of Threonyl-tRNA Synthetase in Regulating Autophagy in Ovarian Cancer

Conference Year

January 2019

Abstract

Autophagy is the process by which lysosomes degrade and recycle damaged proteins and organelles in order to promote cell survival under situations of nutrient deprivation and homeostatic stress. The process of autophagy is initially preventative for cancer, as lysosomal degradation deters the accumulation of damaged organelles and the proliferation of mutated DNA. However, once a tumor has established itself within the tissue, upregulation of autophagy allows the cancer to survive and proliferate under extreme cellular stressors such as chemotherapy and radiation. Studies in the Lounsbury Lab have identified threonyl tRNA synthetase (TARS) as a potential regulator of autophagy in human ovarian cancer cells. This study tests the hypotheses that TARS independently inhibits the process of autophagy within human ovarian cancer cells and that blocking TARS signaling increases markers of autophagy within tumors in a mouse model of ovarian cancer. To assess effects on autophagy, ovarian cancer cells were cultured and transfected with siRNA in order to reduce TARS levels. Quantitative Western blotting was employed to test the effects of TARS knockdown during amino acid starvation, autophagosome stabilization, or mTOR inhibition. Changes were detected using antibodies against TARS and the positive autophagy marker p-AMPK. Initial results indicate that TARS was effectively knocked down by siRNA (% knockdown, pin vivoeffects of TARS activity on autophagy, mice were injected subcutaneously with ID-8 ovarian cancer cells. After three weeks, the animals were treated three times per week with either vehicle control or the TARS inhibitor BC194. Tumor sections were analyzed by immunohistochemistry to detect TARS, autophagy, and inflammatory markers. Results will determine whether tumors from mice treated with BC194 have changes in autophagy markers and inflammatory response. Preliminary results suggest that inflammation is increased in TARS-deficient tumors. The results of this study have strong implications for the potential to identify a novel therapeutic target in ovarian cancer treatment.

Primary Faculty Mentor Name

Dr. Karen Lounsbury

Secondary Mentor Name

Dr. Christopher Francklyn

Faculty/Staff Collaborators

Jake Bellotte, Diana Grinberg, Theresa Wellman

Status

Undergraduate

Student College

College of Arts and Sciences

Program/Major

Biological Sciences, Integrated

Primary Research Category

Health Sciences

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The Role of Threonyl-tRNA Synthetase in Regulating Autophagy in Ovarian Cancer

Autophagy is the process by which lysosomes degrade and recycle damaged proteins and organelles in order to promote cell survival under situations of nutrient deprivation and homeostatic stress. The process of autophagy is initially preventative for cancer, as lysosomal degradation deters the accumulation of damaged organelles and the proliferation of mutated DNA. However, once a tumor has established itself within the tissue, upregulation of autophagy allows the cancer to survive and proliferate under extreme cellular stressors such as chemotherapy and radiation. Studies in the Lounsbury Lab have identified threonyl tRNA synthetase (TARS) as a potential regulator of autophagy in human ovarian cancer cells. This study tests the hypotheses that TARS independently inhibits the process of autophagy within human ovarian cancer cells and that blocking TARS signaling increases markers of autophagy within tumors in a mouse model of ovarian cancer. To assess effects on autophagy, ovarian cancer cells were cultured and transfected with siRNA in order to reduce TARS levels. Quantitative Western blotting was employed to test the effects of TARS knockdown during amino acid starvation, autophagosome stabilization, or mTOR inhibition. Changes were detected using antibodies against TARS and the positive autophagy marker p-AMPK. Initial results indicate that TARS was effectively knocked down by siRNA (% knockdown, pin vivoeffects of TARS activity on autophagy, mice were injected subcutaneously with ID-8 ovarian cancer cells. After three weeks, the animals were treated three times per week with either vehicle control or the TARS inhibitor BC194. Tumor sections were analyzed by immunohistochemistry to detect TARS, autophagy, and inflammatory markers. Results will determine whether tumors from mice treated with BC194 have changes in autophagy markers and inflammatory response. Preliminary results suggest that inflammation is increased in TARS-deficient tumors. The results of this study have strong implications for the potential to identify a novel therapeutic target in ovarian cancer treatment.