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Mechanisms of Non-Conventional Cell Death in Brain Tumor Cells

Kaul, Aparna
http://rave.ohiolink.edu/etdc/view?acc_num=mco1243364096

Abstract Details

2009, Doctor of Philosophy in Biomedical Sciences (Ph.D.), University of Toledo, College of Medicine.
The concept of programmed cell death has evolved over the years to include bothapoptotic and non-apoptotic death mechanisms. This study describes a novel form of non-apoptotic cell death induced as a result of dysregulated macropinocytosis. We have named this cell death “methuosis”. Methuosis is observed when the activated form of Ras GTPase is over-expressed in glioblastoma cells. It is accompanied by the accumulation of large phase-lucent cytoplasmic vacuoles, followed by rounding up, detachment, and disintegration of the cells. The vacuoles quickly take up extracellular fluid-phase tracers, a hallmark of macropinosomes. Our studies also show that the Ras-induced vacuoles are not acidic and are negative for LC3-II (a marker for autophagosomes), transferrin and EEA1 (endosomal markers). These observations rule out the vacuoles originating from autophagosomal, endosomal or lysosomal compartments. Even though caspase activation is observed in dying cells, death is not prevented by zVAD-fmk, a pan caspase inhibitor. Electron microscopy revealed that the dying cells did not show classical signs of apoptosis, like chromatin condensation. These findings indicate that caspase activation is not required for methuosis to occur. Studies performed to decipher the signaling pathway(s) stimulated by Ras revealed that methuosis does not depend on the activation of Raf kinase, PI-3K or RalGDS, the most well-studied Ras signaling intermediates. Interestingly, constitutively active Rac1 induces an identical vacuolar phenotype in glioblastoma cells. Rac1-induced vacuoles are also derived from macropinosomes. We postulate that activated Ras is stimulating Rac GTPase via a unique downstream effector to initiate methuosis in glioblastoma cells. ER stress-initiated apoptosis has recently gained attention as an effective death inducer in cancer cells. This work shows for the first time that the mechanism by which calphostin-C, a photoactivatable inhibitor of protein kinase C, induces apoptosis in cancer cells involves ER stress. Calphostin-C potently reduces the viability of a number of cancer cell lines, including glioblastomas. The cell death induced by cal-C involves accumulation of vacuoles derived from the ER with a concomitant block in the protein trafficking from ER to Golgi. There is a rapid activation of ER stress markers, JNK, PERK, and the induction of pro-apoptotic protein CHOP. Activation of caspases-9 and 7, along with PARP cleavage, is observed following the activation of ER stress signaling. Our studies indicate that apoptosis induced by cal-C has a strong ER-stress component and that this compound has a potential of being exploited as a chemotherapeutic agent for photodynamic therapy.
William Maltese, Ph.D. (Committee Chair)
Ronald Mellgren, Ph.D. (Committee Member)
Ana Maria Oyarce, Ph.D. (Committee Member)
Kam Yeung, Ph.D. (Committee Member)
Jean Overmeyer, Ph.D. (Committee Member)
Kathryn Eisenmann, Ph.D. (Committee Member)
234 p.
Cellular Biology; Molecular Biology
Apoptosis; methuosis; endoplasmic reticulum-stress; macropinocytosis; non-apoptotic; glioblastoma

Recommended Citations

Citations

  • Kaul, A. (2009). Mechanisms of Non-Conventional Cell Death in Brain Tumor Cells [Doctoral dissertation, University of Toledo]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=mco1243364096

    APA Style (7th edition)

  • Kaul, Aparna. Mechanisms of Non-Conventional Cell Death in Brain Tumor Cells. 2009. University of Toledo, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=mco1243364096.

    MLA Style (8th edition)

  • Kaul, Aparna. "Mechanisms of Non-Conventional Cell Death in Brain Tumor Cells." Doctoral dissertation, University of Toledo, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=mco1243364096

    Chicago Manual of Style (17th edition)

mco1243364096
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© 2009, all rights reserved.
This open access ETD is published by University of Toledo Health Science Campus and OhioLINK.