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ScienceWeek
DEVELOPMENTAL BIOLOGY: AUTOPHAGY AND NEONATAL DEVELOPMENT
The following points are made by Nathaniel Heintz (Nature 2004 432:963):
1) All organisms must adapt to environmental stresses. One of the most cogent examples of a response to stress is autophagy -- a process first described in electron microscopic studies in the kidneys of newborn mice[1], and now known to be one of the major pathways for the degradation of long-lived proteins and cellular organelles[2,3]. It has become clear that autophagy is activated in a variety of circumstances in multicellular organisms, and thus must be regulated by diverse stimuli in different cell types. But our knowledge of the biological roles of autophagy in mammals is quite primitive. Kuma et al[4] have provided an insight into the mammalian response to birth and the biological benefits of autophagy.
2) In cells, nutrient deprivation and other stresses elicit a complex series of orchestrated steps that result in the cells enveloping portions of their cytoplasm, delivering them to enzyme-packed organelles known as lysosomes for degradation and recycling (hence the name autophagy, or self-eating)[2,3]. Over the past decade, genetic analysis in yeasts and morphological studies in a wide variety of species have defined the molecular mechanisms of autophagy and shown its induction under many developmental and pathological situations. The logic for the pathway that has been revealed by genetic and molecular dissection: in response to nutrient deprivation, the autophagy pathway scavenges intracellular substituents to supply vital components until conditions improve.
3) To investigate the importance of this pathway in mammals, Mizushima et al[5] made use of a fusion of green fluorescent protein (GFP) and LC3, the mammalian equivalent of Atg8 -- an essential protein in the yeast autophagy pathway. With this fusion protein they could reveal the formation of autophagosomes, vesicular structures that are hallmarks of autophagy. Having introduced GFP-LC3 into young mice, the authors were able to map the induction of autophagy in response to food withdrawal in many tissues, by studying the formation of small fluorescent vesicles carrying the fluorescent fusion protein.
4) The same group, Kuma et al[4], turned their attention to the developmental roles of autophagy. To do so, they first used GFP-LC3 to study the formation of autophagosomes in newborn mice. They observed massive induction of autophagy in the heart muscle, the diaphragm, the lungs and the skin -- all tissues that undergo sudden increases in energy expenditure or environmental exposure immediately following birth. The induction of autophagy was immediate and transient, reaching maximal levels only 3 to 6 hours after birth and declining to basal levels within a day or two. To explain this phenomenon, Kuma et al[4] postulated that the induction of autophagy following birth is required to provide energy before nursing begins.
5) To test this idea, the authors generated mice that lack Atg5 -- another protein required during an early step in the autophagy program. The Atg5-knockout animals were born in normal mendelian ratios but died during the first day after birth. Measurements of autophagy with the GFP-LC3 assay demonstrated that, as expected, the formation of autophagosomes was blocked in the mutant mice. The Atg5-deficient mice also died earlier than wild-type mice when not being suckled; their survival could be extended by hand feeding; and plasma and tissue amino-acid concentrations in non-suckling mutant mice were normal at birth but very much reduced relative to controls 10 hours after delivery. Furthermore, measurements of energy status in the knockout pups indicated that energy production was severely depressed but could be restored by re-feeding.
6) These data show that autophagy is required to produce amino acids and to maintain energy levels in neonatal mice. Taken together with the severely low glucose and lipid levels that are generally evident in newborn mice, the death of the Atg5-knockout mice after birth provides strong support for the hypothesis that autophagy is essential for survival during the unique period experienced by mammals as they make the transition from transplacental nutrition to milk.
References (abridged):
1. Clark, S. L. J. Biophys. Biochem. Cytol. 3, 349-362 (1957)
2. Levine, B. & Klionsky, D. J. Dev. Cell 6, 463-477 (2004)
3. Kamada, Y., Sekito, T. & Ohsumi, Y. Curr. Top. Microbiol. Immunol. 279, 73-84 (2004)
4. Kuma, A. et al. Nature 432, 1032-1036 (2004)
5. Mizushima, N., Yamamoto, A., Matsui, M., Yoshimori, T. & Ohsumi, Y. Mol. Cell. Biol. 15, 1101-1111 (2003)
Nature http://www.nature.com/nature
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Related Material:
CELL BIOLOGY: ON AUTOPHAGY
The following points are made by T. Shintani and D.J. Klionsky (Science 2004 306:990):
1) Cellular homeostasis requires a constant balance between biosynthetic and catabolic processes. Eukaryotic cells primarily use two distinct mechanisms for large-scale degradation, the proteasome and autophagy; but only autophagy has the capacity to degrade entire organelles. The three types of autophagy are macroautophagy, microautophagy, and chaperone-mediated autophagy (1).
2) Macroautophagy (hereafter called autophagy) plays an important physiological role in human health. In autophagy, a double- or multi-membrane-bound structure, called the autophagosome or autophagic vacuole, is formed de novo to sequester cytoplasm. Then, the vacuole membrane fuses with the lysosome to deliver the contents into the organelle lumen, where the contents are degraded and the resulting macromolecules recycled.
3) Autophagy occurs at basal levels in most tissues and contributes to the routine turnover of cytoplasmic components. However, autophagy can be induced by a change of environmental conditions such as nutrient depletion. In addition to turnover of cellular components, autophagy is involved in development, differentiation, and tissue remodeling in various organisms (2). Autophagy is also implicated in certain human diseases. Paradoxically, autophagy can serve to protect cells but may also contribute to cell damage.
4) Autophagy is involved in programmed cell death (PCD). Type I PCD, apoptosis, is characterized by condensation of cytoplasm and chromatin, DNA fragmentation, and cell fragmentation into apoptotic bodies, followed by removal and degradation of the dying cells by phagocytosis. Type II PCD (autophagic) is characterized by the accumulation of autophagic vesicles (autophagosomes and autophagolysosomes) and is often observed when massive cell elimination is demanded or when phagocytes do not have easy access to the dying cells. One feature that distinguishes apoptosis from autophagic cell death is the source of the lysosomal enzymes used for most of the dying-cell degradation. Apoptotic cells use phagocytic cell lysosomes for this process, whereas cells with autophagic morphology use the endogenous lysosomal machinery of dying cells. It has been unclear whether autophagy directly executes cell death or is the secondary effect of apoptosis. A recent study, however, suggests that autophagy might cause cell death (3). Caspase inhibitor-induced autophagic cell death is severely affected by RNA interference (RNAi) with ATG7 and beclin 1 expression, two genes whose products are essential for autophagy (3).
5) In summary: Autophagy, the process by which cells recycle cytoplasm and dispose of excess or defective organelles, has entered the research spotlight largely owing to the discovery of the protein components that drive this process. Identifying the autophagy genes in yeast and finding orthologs in other organisms reveals the conservation of the mechanism of autophagy in eukaryotes and allows the use of molecular genetics and molecular biology in different model systems to study this process. By mostly morphological studies, autophagy has been linked to disease processes. Whether autophagy protects from or causes disease is unclear.(4,5)
References (abridged):
1. D. J. Klionsky, Ed., Autophagy (Landes Bioscience, Georgetown, TX, 2004), pp. 1-303
2. B. Levine, D. J. Klionsky, Dev. Cell 6, 463 (2004)
3. L. Yu et al., Science 304, 1500 (2004)
4. T. Raveh, G. Droguett, M. S. Horwitz, R. A. DePinho, A. Kimchi, Nature Cell Biol. 3, 1 (2001)
5. B. Inbal, S. Bialik, I. Sabanay, G. Shani, A. Kimchi, J. Cell Biol. 157, 455 (2002)
Science http://www.sciencemag.org
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Related Material:
CELL BIOLOGY: ON AUTOPHAGY IN BIOLOGICAL CELLS
The following points are made by Daniel J. Klionsky (Nature 2004 431:31):
1) When you are fasting or are otherwise deprived of food, your body begins to break down stored nutrients to keep essential processes and organs (such as your brain) supplied with fuel. Similarly, when a cell is deprived of nutrients it will degrade some of its own constituents to stay alive. It does this by the process of autophagy -- literally, "self-eating".
2) Autophagy is marked by the formation of autophagosomes --large vesicles, bounded by a double membrane, that sequester cytosol and organelles such as mitochondria. Fusion of an autophagosome with a lysosome releases the inner vesicle into the lysosome, where enzymes break the vesicle down. Degradation and recycling of the vesicle's constituents enable the cell to continue to carry out essential processes.
3) Because autophagy has the capacity to degrade entire organelles, it could be harmful if it occurred randomly. Autophagy must therefore be tightly regulated. The question is: how is this regulation accomplished? How does a cell or organism sense its environment and trigger an appropriate signal to induce or suppress autophagy? The process occurs in organisms ranging from yeast to worms to humans(3).
4) Drosophila larvae have a storage organ called the fat body, which has analogous functions to those of the liver and adipose (fat) tissue in vertebrates: it is a source of large amounts of potential energy. If an animal has just eaten, of course, there is no need to tap into these energy stores. In response to food, the hormone insulin is produced; this binds a receptor on the surface of cells in the fat body and throughout the organism, and triggers a signalling cascade. An important part of this cascade is an enzyme termed class I phosphatidylinositol-3-OH kinase (PI(3)K). This adds a phosphate group to a particular position on the lipid phosphatidylinositol, which is part of the cell membrane. Various proteins in turn bind the phosphorylated lipid and become activated, thus transmitting the external signal (in this case, insulin) into the cell. A central player in this pathway is the enzyme Tor, which is involved in many regulatory events connected with energy metabolism, and which suppresses autophagy(4).
5) So this pathway provides a mechanism by which cells can block self-eating if the organism has just fed. On the other hand, the pathway also provides a means of inducing autophagy when the organism is starved: a lack of food (particularly of the sugar glucose) leads to a lack of insulin, leaving the pathway inactive and enabling the cell to tap into its storage reserves. But how does the cell regulate the degree of autophagy so that it does not get out of hand? An explanation for this additional level of control comes from Scott et al(1). The activity of the enzyme p70 S6 kinase has been shown to increase when Tor is turned on --that is, when nutrients are abundant(5). This led to the proposal that p70 S6 kinase is itself a negative regulator of autophagy. However, Scott et al(1). now show that this enzyme must be active for autophagy to be maximally induced.(2)
References (abridged):
1. Scott, R. C., Schuldiner, O. & Neufeld, T. P. Dev. Cell 7, 167-178 (2004)
2. Rusten, T. E. et al. Dev. Cell 7, 179-192 (2004)
3. Reggiori, F. & Klionsky, D. J. Eukaryot. Cell 1, 11-21 (2002)
4. Jacinto, E. & Hall, M. N. Nature Rev. Mol. Cell Biol. 4, 117-126 (2003) (Erratum: 4, 249 (2003))
5. Chung, J., Kuo, C. J., Crabtree, G. R. & Blenis, J. Cell 69, 1227-1236 (1992)
Nature http://www.nature.com/nature
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