Departmental Affiliation and City, State, Zip for All Authors

Department of Biomedical Engineering - University of Texas Austin, TX 78705; Department of Microbiology, Immunology and Genetics - University of North Texas Health Science Center Fort Worth, Texas 76107

Scientific Abstract

Regulation of Autophagy by PKCε in Breast Cancer Cells Samuel Razmi1, Alakananda Basu, Ph. D.2 1Department of Biomedical Engineering, University of Texas, Austin, TX 2Department of Microbiology, Immunology and Genetics, University of North Texas Health Science Center Background: The Protein kinase C (PKC) family plays a critical role in signal transduction and cell regulation. The PKC family is categorized into three subgroups: conventional, novel, and atypical PKCs. PKCε is a member of the novel PKC subgroup. PKCε is considered an oncoprotein. It promotes cell survival by inhibiting apoptosis. Autophagy is a process of self-killing in which a portion of cellular components are degraded and recycled to provide nutrients during stressful or nutrient-deprived conditions. Cancer cells may use autophagy as a form of cell survival. The goal of this study is to determine if PKCε regulates autophagy. Methods: T47D breast cancer cells, in which PKCε was depleted by siRNA or stably overexpressed PKCε or mCherry-GFP-LC3, were used in this study. The levels of PKCε and autophagy indicators p62 and LC3 were detected by Western blot analysis. Cells were starved by culturing in EBSS (Earle’s Balanced Salt Solution) and processed for either Western blot analysis or confocal microscopy. To monitor autophagy flux the experiments were performed in the presence and absence of lysosomotropic agent bafilomycin A (BFA). Autophagy puncta were monitored by confocal microscopy. Results: Knockdown of PKCε decreased LC3II and increased p62 while overexpression of PKCε increased LC3II and decreased p62. PKCε knockdown also decreased starvation-induced autophagy. Knockdown of PKCε decreased and overexpression of PKCε increased autophagy puncta. Conclusion: Our results suggest that PKCε promotes autophagy in breast cancer cells. Thus, PKCε may contribute to breast cancer cell survival not only by inhibiting apoptosis but also by increasing autophagy. Research reported in this publication was supported by the Department of Health and Human Services, National Institute of Health, National Heart, Lung and Blood Institute, SMART Award Number 4R25HL007786-25 to Harlan Jones, Ph.D. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

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Regulation of Autophagy by PKCε in Breast Cancer Cells

Regulation of Autophagy by PKCε in Breast Cancer Cells Samuel Razmi1, Alakananda Basu, Ph. D.2 1Department of Biomedical Engineering, University of Texas, Austin, TX 2Department of Microbiology, Immunology and Genetics, University of North Texas Health Science Center Background: The Protein kinase C (PKC) family plays a critical role in signal transduction and cell regulation. The PKC family is categorized into three subgroups: conventional, novel, and atypical PKCs. PKCε is a member of the novel PKC subgroup. PKCε is considered an oncoprotein. It promotes cell survival by inhibiting apoptosis. Autophagy is a process of self-killing in which a portion of cellular components are degraded and recycled to provide nutrients during stressful or nutrient-deprived conditions. Cancer cells may use autophagy as a form of cell survival. The goal of this study is to determine if PKCε regulates autophagy. Methods: T47D breast cancer cells, in which PKCε was depleted by siRNA or stably overexpressed PKCε or mCherry-GFP-LC3, were used in this study. The levels of PKCε and autophagy indicators p62 and LC3 were detected by Western blot analysis. Cells were starved by culturing in EBSS (Earle’s Balanced Salt Solution) and processed for either Western blot analysis or confocal microscopy. To monitor autophagy flux the experiments were performed in the presence and absence of lysosomotropic agent bafilomycin A (BFA). Autophagy puncta were monitored by confocal microscopy. Results: Knockdown of PKCε decreased LC3II and increased p62 while overexpression of PKCε increased LC3II and decreased p62. PKCε knockdown also decreased starvation-induced autophagy. Knockdown of PKCε decreased and overexpression of PKCε increased autophagy puncta. Conclusion: Our results suggest that PKCε promotes autophagy in breast cancer cells. Thus, PKCε may contribute to breast cancer cell survival not only by inhibiting apoptosis but also by increasing autophagy. Research reported in this publication was supported by the Department of Health and Human Services, National Institute of Health, National Heart, Lung and Blood Institute, SMART Award Number 4R25HL007786-25 to Harlan Jones, Ph.D. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Manuscript Number

1029