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 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 versus those that measure fl ux through the autophagy pathway (i.e., the complete process 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, defi ned as increased autophagy 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 fi eld 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. 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 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. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so 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 autophagy assays, we hope to encourage technical innovation in the field

The role of micro-organisms in the ecological connectivity of running waters

Pusch M, Fiebig D, Brettar I, et al. The role of micro-organisms in the ecological connectivity of running waters. Freshwater Biology. 1998;40(3):453-495

The physics behind the fizz in champagne and sparkling wines

Bubbles in a glass of champagne may seem like the acme of frivolity to most of people, but in fact they may rather be considered as a fantastic playground for any physicist. Actually, the so-called effervescence process, which enlivens champagne and sparkling wines tasting, is the result of the fine interplay between CO2 dissolved gas molecules, tiny air pockets trapped within microscopic particles during the pouring process, and some both glass and liquid properties. Results obtained concerning the various steps where the CO2 molecule plays a role (from its ingestion in the liquid phase during the fermentation process to its progressive release in the headspace above the tasting glass as bubbles collapse) are gathered and synthesized to propose a self-consistent and global overview of how gaseous and dissolved CO2 impact champagne and sparkling wine science. Physicochemical processes behind the nucleation, rise, and burst of gaseous CO2 bubbles found in glasses poured with champagne and sparkling wines are depicted. Those phenomena observed in close-up through high-speed photography are often visually appealing. I hope that your enjoyment of champagne will be enhanced after reading this fully illustrated review dedicated to the science hidden right under your nose each time you enjoy a glass of champagne. Gérard Liger-Belair: He received his PhD in physical sciences in 2001 from the University of Reims, in France. He received an associate professor position at the University of Reims in 2002, and a full professor position, in 2007, in the same University. He has been researching the physics and chemistry behind the bubbling properties of champagne and sparkling wines for several years. His current interests include the science of bubbles, foams and thin films, and their broad interdisciplinary applications. He is the author of several academic and popular science books. His first book, Uncorked: the science of champagne, published by Princeton University Press, won the 2004 award for the Best Professional/Scholarly Book in Physics from the Association of American Publishers