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Macquarie University
AU
2 researchers0 verified2 linked papers6,206 indexed citations
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2016 · Autophagy · 6,021 citations
Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)
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. For example, a key point that needs to be emphasized is thatthere 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 versus those that measure flux through the autophagy pathway (i.e., the completeprocess including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defined as increasedautophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost 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 manycases, 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. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. 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 forreviewers 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 multipleassays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation, it is imperative to target by gene knockout or RNA interference more than one autophagyrelated protein. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways implying that not all Atg proteins can be used as a specific marker for an autophagic process. 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 assays, we hope to encourage technical innovation in the field.
2011 · Diversity and Distributions · 185 citations
Reproductive biology of Australian acacias: important mediator of invasiveness?
Abstract Aim Reproductive traits are important mediators of establishment and spread of introduced species, both directly and through interactions with other life‐history traits and extrinsic factors. We identify features of the reproductive biology of Australian acacias associated with invasiveness. Location Global. Methods We reviewed the pollination biology, seed biology and alternative modes of reproduction of Australian acacias using primary literature, online searches and unpublished data. We used comparative analyses incorporating an Acacia phylogeny to test for associations between invasiveness and eight reproductive traits in a group of introduced and invasive (23) and non‐invasive (129) species. We also explore the distribution of groups of trait ‘syndromes’ between invasive and non‐invasive species. Results Reproductive trait data were only available for 126 of 152 introduced species in our data set, representing 23/23 invasive and 103/129 non‐invasive species. These data suggest that invasives reach reproductive maturity earlier (10/13 within 2 years vs. 7/26 for non‐invasives) and are more commonly able to resprout (11/21 vs. 13/54), although only time to reproductive maturity was significant when phylogenetic relationships were controlled for. Our qualitative survey of the literature suggests that invasive species in general tend to have generalist pollination systems, prolific seed production, efficient seed dispersal and the accumulation of large and persistent seed banks that often have fire‐, heat‐ or disturbance‐triggered germination cues. Conclusions Invasive species respond quicker to disturbance than non‐invasive taxa. Traits found to be significant in our study require more in‐depth analysis involving data for a broader array of species given how little is known of the reproductive biology of so many taxa in this species‐rich genus. Sets of reproductive traits characteristic of invasive species and a general ability to reproduce effectively in new locations are widespread in Australian acacias. Unless there is substantial evidence to the contrary, care should be taken with all introductions.