Lysosome dysfunction triggers Atg7-dependent neural apoptosis.
ABSTRACT Macroautophagy (autophagy) is a process wherein bulk cytosolic proteins and damaged organelles are sequestered and degraded via the lysosome. Alterations in autophagy-associated proteins have been shown to cause neural tube closure defects, neurodegeneration, and tumor formation. Normal lysosome function is critical for autophagy completion and when altered may lead to an accumulation of autophagic vacuoles (AVs) and caspase activation. The tumor suppressor p53 is highly expressed in neural precursor cells (NPCs) and has an important role in the regulation of both autophagy and apoptosis. We hypothesized that altered lysosome function would lead to NPC death via an interaction between autophagy- and apoptosis-associated proteins. To test our hypothesis, we utilized FGF2-expanded NPCs and the neural stem cell line, C17.2, in combination with the lysosomotropic agent chloroquine (CQ) and the vacuolar ATPase inhibitor bafilomycin A1 (Baf A1). Both CQ and Baf A1 caused concentration- and time-dependent AV accumulation, p53 phosphorylation, increased damage regulator autophagy modulator levels, caspase-3 activation, and cell death. Short hairpin RNA knockdown of Atg7, but not Beclin1, expression significantly inhibited CQ- and Baf A1-induced cell death, indicating that Atg7 is an upstream mediator of lysosome dysfunction-induced cell death. Cell death and/or caspase-3 activation was also attenuated by protein synthesis inhibition, p53 deficiency, or Bax deficiency, indicating involvement of the intrinsic apoptotic death pathway. In contrast to lysosome dysfunction, starvation-induced AV accumulation was inhibited by either Atg7 or Beclin1 knockdown, and Atg7 knockdown had no effect on starvation-induced death. These findings indicate that Atg7- and Beclin1-induced autophagy plays a cytoprotective role during starvation but that Atg7 has a unique pro-apoptotic function in response to lysosome dysfunction.
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ABSTRACT: Mounting evidence suggests that reactive oxygen species (ROS) are multifaceted signaling molecules implicated in a variety of cellular programs during physiological as well as pathological conditions. Recently, ROS produced endogenously, by deranged metabolism of cancer cells, or exogenously, by ROS-generating drugs, have been shown to promote macroautophagy, a lysosomal pathway of self-degradation with essential prosurvival functions. Several molecular aspects of the modulation of autophagy pathways by ROS have been revealed in the past years and it is now clear that these processes are mutually linked and play a crucial role in cancer progression and in response to cancer therapeutics. In this review we address the molecular mechanisms underlying the activation of autophagy pathways by ROS and focus on the role of autophagy in cancer cells responding to ROS-producing agents, which are utilized as a therapeutic modality to kill cancer cells.Autophagy 10/2010; 6(7):838-54. · 12.04 Impact Factor
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ABSTRACT: V-ATPases are highly conserved, ATP-driven proton pumps regulated by reversible dissociation of its cytosolic, peripheral V1 domain from the integral membrane Vo domain. Multiple stresses induce changes in V1- Vo assembly, but signaling mechanisms behind these changes are not understood. Here we show that certain stress-responsive changes in V-ATPase activity and assembly require the signaling lipid phosphatidylinositol (3,5) bis-phosphate (PI(3,5)P2). V-ATPase activation through V1-Vo assembly in response to salt stress is strongly dependent on PI(3,5)P2 synthesis. Purified Vo complexes preferentially bind to PI(3,5)P2 on lipid arrays, suggesting direct binding between the lipid and the membrane sector of the V-ATPase. Increasing PI(3,5)P2 levels in vivo recruits the N-terminal domain of Vo sector subunit Vph1p from cytosol to membranes, independent of other subunits. This Vph1p domain is critical for V1-Vo interaction, suggesting that interaction of Vph1p with PI(3,5)P2-containing membranes stabilizes V1-Vo assembly and thus increases V-ATPase activity. These results help explain the previously described vacuolar acidification defect in yeast fab1 and vac14 mutants and suggest that human disease phenotypes associated with PI(3,5)P2 loss may arise from compromised V-ATPase stability and regulation.Molecular biology of the cell 02/2014; · 5.98 Impact Factor
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ABSTRACT: The process of cellular eating, or the phagocytic swallowing of one cell by another, is an ancient manifestation of the struggle for life itself. Following the endosymbiotic origin of eukaryotic cells, increased cellular and then multicellular complexity was accompanied by the emergence of autophagic mechanisms for self-digestion. Heterophagy and autophagy function not only to protect the nutritive status of cells, but also as defensive responses against microbial pathogens externally or the ill effects of damaged proteins and organelles within. Because of the key roles played by phagocytosis and autophagy in a wide range of acute and chronic human diseases, pathologists have played similarly key roles in elucidating basic regulatory phases for both processes. Studies in diverse organ systems (including the brain, liver, kidney, lung, and muscle) have defined key roles for these lysosomal pathways in infection control, cell death, inflammation, cancer, neurodegeneration, and mitochondrial homeostasis. The literature reviewed here exemplifies the role of pathology in defining leading-edge questions for continued molecular and pathophysiological investigations into all forms of cellular digestion.American Journal Of Pathology 01/2013; · 4.60 Impact Factor