Martina Wirth

London Research Institute, Londinium, England, United Kingdom

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Publications (3)22.04 Total impact

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    ABSTRACT: Autophagy (self-eating) is a highly conserved, vesicular pathway that cells use to eat pieces of themselves, including damaged organelles, protein aggregates or invading pathogens, for self-preservation and survival (Choi et al., N Engl J Med 368:651-662, 2013; Lamb et al., Nat Rev Mol Cell Biol 14:759-774, 2013). Autophagy can be delineated into three major vesicular compartments (the phagophore, autophagosome, autolysosome, see Fig. 1). The initial stages of the pathway involve the formation of phagophores (also called isolation membranes), which are open, cup-shaped membranes that expand and sequester the cytosolic components, including organelles and aggregated proteins or intracellular pathogens. Closure of the phagophore creates an autophagosome, which is a double-membrane vesicle. Fusion of the autophagosome with the lysosome, to form an autolysosome, delivers the content of the autophagosome into the lysosomal lumen and allows degradation to occur.Autophagy is a dynamic process that is initiated within 15 min of amino acid starvation in cell culture systems (Köchl et al., Traffic 7:129-145, 2006) and is likely to occur as rapidly in vivo (Mizushima et al., J Cell Biol 152:657-668, 2001). To initiate studies on the formation of the autophagosomes, and trafficking to and from the autophagic pathway, an ideal starting approach is to do a morphological analysis in fixed cells. Additional validation of the morphological data can be obtained using simple Western blot analysis. Here we describe the most commonly used morphological technique to study autophagy, in particular, using the most reliable marker, microtubule-associated protein 1A/1B-light chain 3 (LC3). In addition, we describe a second immunofluorescence assay to determine if autophagy is being induced, using an antibody to WD repeat domain, phosphoinositide interacting 2 (WIPI2), an effector of the phosphatidylinositol (3)-phosphate (PI3P) produced during autophagosome formation.
    Methods in molecular biology (Clifton, N.J.) 01/2015; 1270:155-65. DOI:10.1007/978-1-4939-2309-0_12 · 1.29 Impact Factor
  • Martina Wirth, Justin Joachim, Sharon A Tooze
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    ABSTRACT: Autophagy is a conserved and highly regulated degradative membrane trafficking pathway, maintaining energy homeostasis and protein synthesis during nutrient stress. Our understanding of how the autophagy machinery is regulated has expanded greatly over recent years. The ULK and Beclin1-PI3KC3 complexes are key signaling complexes required for autophagosome formation. The nutrient and energy sensors mTORC1 and AMPK signal directly to the ULK complex and affect its activity. Formation and activation of distinct Beclin1-PI3KC3 complexes produces PI3P, a signaling lipid required for the recruitment of autophagy effectors. In this review we discuss how the mammalian ULK1 and Beclin1 complexes are controlled by post-translational modifications and protein-protein interactions and we highlight data linking these complexes together.
    Seminars in Cancer Biology 05/2013; 23(5). DOI:10.1016/j.semcancer.2013.05.007 · 9.33 Impact Factor
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    ABSTRACT: Autophagy is a conserved and highly regulated catabolic pathway, transferring cytoplasmic components in autophagosomes to lysosomes for degradation and providing amino acids during starvation. In multicellular organisms autophagy plays an important role for tissue homeostasis, and deregulation of autophagy has been implicated in a broad range of diseases, including cancer and neurodegenerative disorders. In mammals, many aspects of autophagy still need to be fully elucidated: what is the exact hierarchy and relationship between ATG proteins and other factors that lead to the formation and expansion of phagophores? Where does the membrane source for autophagosome formation originate? Which signaling events trigger amino acid starvation-induced autophagy? How are the activities of ULK1/2 and the class III PtdIns3K regulated and linked to each other? To develop therapeutic strategies to manipulate autophagy in human disease, a comprehensive understanding of the molecular protein machinery mediating and regulating autophagy is required.
    Autophagy 09/2012; 8(9):1397-400. DOI:10.4161/auto.21043 · 11.42 Impact Factor