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- Klionsky, Daniel J., et al.
(author)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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In: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Research review (peer-reviewed)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.
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| 4. |
- O'Gorman, Eoin J., et al.
(author)
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Impacts of Warming on the Structure and Functioning of Aquatic Communities : Individual-to Ecosystem-Level Responses
- 2012
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In: Advances in Ecological Research, Vol 47. - : Elsevier. - 9780123983152 ; , s. 81-176
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Book chapter (peer-reviewed)abstract
- Environmental warming is predicted to rise dramatically over the next century, yet few studies have investigated its effects in natural, multi-species systems. We present data collated over an 8-year period from a catchment of geothermally heated streams in Iceland, which acts as a natural experiment on the effects of warming across different organisational levels and spatiotemporal scales. Body sizes and population biomasses of individual species responded strongly to temperature, with some providing evidence to support temperature size rules. Macroinvertebrate and meiofaunal community composition also changed dramatically across the thermal gradient. Interactions within the warm streams in particular were characterised by food chains linking algae to snails to the apex predator, brown trout These chains were missing from the colder systems, where snails were replaced by much smaller herbivores and invertebrate omnivores were the top predators. Trout were also subsidised by terrestrial invertebrate prey, which could have an effect analogous to apparent competition within the aquatic prey assemblage. Top-down effects by snails on diatoms were stronger in the warmer streams, which could account for a shallowing of mass-abundance slopes across the community. This may indicate reduced energy transfer efficiency from resources to consumers in the warmer systems and/or a change in predator-prey mass ratios. All the ecosystem process rates investigated increased with temperature, but with differing thermal sensitivities, with important implications for overall ecosystem functioning (e.g. creating potential imbalances in elemental fluxes). Ecosystem respiration rose rapidly with temperature, leading to increased heterotrophy. There were also indications that food web stability may be lower in the warmer streams.
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| 5. |
- Lopes, Miguel, et al.
(author)
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Temporal profiling of cytokine-induced genes in pancreatic beta-cells by meta-analysis and network inference
- 2014
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In: Genomics. - : Elsevier BV. - 0888-7543 .- 1089-8646. ; 103:4, s. 264-275
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Journal article (peer-reviewed)abstract
- Type I Diabetes (T1D) is an autoimmune disease where local release of cytokines such as IL-1 beta and IFN-gamma contributes to beta-cell apoptosis. To identify relevant genes regulating this process we performed a meta-analysis of 8 datasets of beta-cell gene expression after exposure to IL-1 beta and IFN-gamma. Two of these datasets are novel and contain time-series expressions in human islet cells and rat INS-1E cells. Genes were ranked according to their differential expression within and after 24 h from exposure, and characterized by function and prior knowledge in the literature. A regulatory network was then inferred from the human time expression datasets, using a time-series extension of a network inference method. The two most differentially expressed genes previously unknown in T1D literature (RIPK2 and ELF3) were found to modulate cytokine-induced apoptosis. The inferred regulatory network is thus supported by the experimental validation, providing a proof-of-concept for the proposed statistical inference approach.
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