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ABSTRACT: Macroautophagy (hereafter autophagy) is a degradative cellular pathway that protects eukaryotic cells from stress, starvation and microbial infection. This process must be tightly controlled because too little or too much autophagy can be deleterious to cellular physiology. The phosphatidylinositol (PtdIns) 3-kinase Vps34 is a lipid kinase that regulates autophagy, but the role of other PtdIns kinases has not been examined. Here, we demonstrate a role for PtdIns 4-kinases and PtdIns4P 5-kinases in selective and nonselective types of autophagy in yeast. The PtdIns 4-kinase Pik1 is involved in Atg9 trafficking through the Golgi, and is involved in both nonselective and selective types of autophagy, whereas the PtdIns4P 5-kinase Mss4 is specifically involved in mitophagy but not nonselective autophagy. Our data indicate that phosphoinositide kinases have multiple roles in the regulation of autophagic pathways.
Journal of Biological Chemistry 09/2012; · 4.77 Impact Factor
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Daniel J Klionsky,
Fabio C Abdalla,
Hagai Abeliovich,
Robert T Abraham,
Abraham Acevedo-Arozena,
Khosrow Adeli,
Lotta Agholme,
Maria Agnello,
Patrizia Agostinis,
Julio A Aguirre-Ghiso, [......],
Changlian Zhu,
Wei-Guo Zhu,
Xiao-Feng Zhu,
Xiongwei Zhu,
Yuangang Zhu,
Teresa Zoladek,
Wei-Xing Zong,
Antonio Zorzano,
Jürgen Zschocke,
Brian Zuckerbraun
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ABSTRACT: In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
Autophagy 04/2012; 8(4):445-544. · 7.45 Impact Factor
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ABSTRACT: Great progress has been made toward understanding the pathogenesis of Parkinson's disease (PD) during the past two decades, mainly as a consequence of the discovery of specific gene mutations contributing to the onset of PD. Recently, dysregulation of the autophagy pathway has been observed in the brains of PD patients and in animal models of PD, indicating the emerging role of autophagy in this disease. Indeed, autophagy is increasingly implicated in a number of pathophysiologies, including various neurodegenerative diseases. This article will lead you through the connection between autophagy and PD by introducing the concept and physiological function of autophagy, and the proteins related to autosomal dominant and autosomal recessive PD, particularly α-synuclein and PINK1-PARKIN, as they pertain to autophagy.
Cold Spring Harbor perspectives in medicine. 04/2012; 2(4):a009357.
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ABSTRACT: The autophagy-dependent selective degradation of mitochondria (mitophagy) plays an important role in removing excessive, damaged and dysfunctional mitochondria to maintain a proper cellular homeostasis. Relative to its significance in cell physiology, very little is known about the molecular machinery and regulatory mechanism of mitophagy in mammalian cells or yeast. We found that two mitogen-activated protein kinases (MAPKs), Slt2 and Hog1, are required for mitophagy in Saccharomyces cerevisiae. Slt2 is involved in both mitophagy and pexophagy (the selective degradation of peroxisomes through autophagy), whereas Hog1 functions specifically in mitophagy.
Autophagy 12/2011; 7(12):1564-5. · 7.45 Impact Factor
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ABSTRACT: Macroautophagy (hereafter referred to simply as autophagy) is a catabolic pathway that mediates the degradation of long-lived proteins and organelles in eukaryotic cells. The regulation of mitochondrial degradation through autophagy plays an essential role in the maintenance and quality control of this organelle. Compared with our understanding of the essential function of mitochondria in many aspects of cellular metabolism such as energy production and of the role of dysfunctional mitochondria in cell death, little is known regarding their degradation and especially how upstream signaling pathways control this process. Here, we report that two mitogen-activated protein kinases (MAPKs), Slt2 and Hog1, are required for mitophagy in Saccharomyces cerevisiae. Slt2 is required for the degradation of both mitochondria and peroxisomes (via pexophagy), whereas Hog1 functions specifically in mitophagy. Slt2 also affects the recruitment of mitochondria to the phagophore assembly site (PAS), a critical step in the packaging of cargo for selective degradation.
The Journal of Cell Biology 05/2011; 193(4):755-67. · 10.26 Impact Factor
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Circulation Research 04/2011; 108(7):787-8. · 9.49 Impact Factor
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Tomotake Kanki,
Ke Wang,
Misuzu Baba,
Clinton R Bartholomew,
Melinda A Lynch-Day,
Zhou Du,
Jiefei Geng, Kai Mao,
Zhifen Yang,
Wei-Lien Yen,
Daniel J Klionsky
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ABSTRACT: Mitophagy is the process of selective mitochondrial degradation via autophagy, which has an important role in mitochondrial quality control. Very little is known, however, about the molecular mechanism of mitophagy. A genome-wide yeast mutant screen for mitophagy-defective strains identified 32 mutants with a block in mitophagy, in addition to the known autophagy-related (ATG) gene mutants. We further characterized one of these mutants, ylr356wDelta that corresponds to a gene whose function has not been identified. YLR356W is a mitophagy-specific gene that was not required for other types of selective autophagy or macroautophagy. The deletion of YLR356W partially inhibited mitophagy during starvation, whereas there was an almost complete inhibition at post-log phase. Accordingly, we have named this gene ATG33. The new mutants identified in this analysis will provide a useful foundation for researchers interested in the study of mitochondrial homeostasis and quality control.
Molecular biology of the cell 09/2009; 20(22):4730-8. · 5.98 Impact Factor
