Publications by authors named "Adrien Pasquier"

9 Publications

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Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition).

Autophagy 2021 Jan 8;17(1):1-382. Epub 2021 Feb 8.

University of Crete, School of Medicine, Laboratory of Clinical Microbiology and Microbial Pathogenesis, Voutes, Heraklion, Crete, Greece; Foundation for Research and Technology, Institute of Molecular Biology and Biotechnology (IMBB), Heraklion, Crete, Greece.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
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http://dx.doi.org/10.1080/15548627.2020.1797280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7996087PMC
January 2021

Breaking Bad and Breaking Good: β-Cell Autophagy Pathways in Diabetes.

J Mol Biol 2020 03 2;432(5):1494-1513. Epub 2019 Aug 2.

Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch 67404, France; Centre National de la Recherche Scientifique, UMR7104, Illkirch 67404, France; Institut National de la Santé et de la Recherche Médicale, U964, Illkirch 67404, France; Université de Strasbourg, Strasbourg 67081, France; Laboratoire de Biochimie et de Biologie Moléculaire, Nouvel Hôpital Civil, Strasbourg 67091, France. Electronic address:

For many decades the lysosome has been recognized as the terminal center of cellular waste disposal. Products of lysosomal degradation are either recycled in biosynthetic pathways or are further metabolized to produce energy. As such the lysosome was attributed a rather passive role in cellular metabolism merely transforming bulk material into small metabolites. More recently, however, the emerging evidence has brought the lysosome to the center of nutrient sensing as the organelle that harbors a complex signaling machinery which dynamically and actively regulates cell metabolism. The pancreatic β cell is unique in as much as nutrient sensing is directly coupled to insulin secretion. Importantly, defects in insulin secretion constitute a hallmark in the progression of patients from a state of impaired glucose tolerance to full blown type 2 diabetes (T2D). However, mechanisms linking nutrient-dependent lysosomal function to insulin secretion and more generally to β cell health have evolved only very recently. This review discusses emerging concepts in macroautophagy and macroautophagy-independent processes of cargo delivery to lysosomes as well as nutrient-dependent lysosomal signaling specifically in the context of β cell function in health and disease. Such mechanisms may provide a novel source of therapeutic targets to be exploited in the context of β cell failure in diabetes in the near future.
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http://dx.doi.org/10.1016/j.jmb.2019.07.030DOI Listing
March 2020

Lysosomal degradation of newly formed insulin granules contributes to β cell failure in diabetes.

Nat Commun 2019 07 25;10(1):3312. Epub 2019 Jul 25.

Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404, Illkirch, France.

Compromised function of insulin-secreting pancreatic β cells is central to the development and progression of Type 2 Diabetes (T2D). However, the mechanisms underlying β cell failure remain incompletely understood. Here, we report that metabolic stress markedly enhances macroautophagy-independent lysosomal degradation of nascent insulin granules. In different model systems of diabetes including of human origin, stress-induced nascent granule degradation (SINGD) contributes to loss of insulin along with mammalian/mechanistic Target of Rapamycin (mTOR)-dependent suppression of macroautophagy. Expression of Protein Kinase D (PKD), a negative regulator of SINGD, is reduced in diabetic β cells. Pharmacological activation of PKD counters SINGD and delays the onset of T2D. Conversely, inhibition of PKD exacerbates SINGD, mitigates insulin secretion and accelerates diabetes. Finally, reduced levels of lysosomal tetraspanin CD63 prevent SINGD, leading to increased insulin secretion. Overall, our findings implicate aberrant SINGD in the pathogenesis of diabetes and suggest new therapeutic strategies to prevent β cell failure.
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http://dx.doi.org/10.1038/s41467-019-11170-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6658524PMC
July 2019

Lysosomes in nutrient signalling: A focus on pancreatic β-cells.

Diabetes Obes Metab 2018 09;20 Suppl 2:104-115

Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France.

Regulated insulin secretion from pancreatic β-cells is a major process maintaining glucose homeostasis in mammals. Enhancing insulin release in response to chronic nutrient overload and obesity-related insulin resistance (pre-diabetes) requires several adaptive cellular mechanisms maintaining β-cell health under such stresses. Once these mechanisms are overwhelmed, β-cell failure occurs leading to full-blown Type 2 Diabetes (T2D). Nutrient-dependent macroautophagy represents one such adaptive mechanism in β-cells. While macroautophagy levels are high and protective in β-cells in pre-diabetes, they decrease at later stages contributing to β-cell failure. However, mechanisms compromising macroautophagy in β-cells remain poorly understood. In this review, we discuss how recently discovered signalling cascades that emanate from the limiting membrane of lysosomes contribute to changes in macroautophagy flux in physiology and disease. In particular, these mechanisms are put into context with β-cell function highlighting most recently described links between nutrient-dependent lysosomal signalling pathways and insulin secretion. Understanding these mechanisms in response to metabolic stress might pave the way for development of more tailored treatment strategies aimed at preserving β-cell health.
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http://dx.doi.org/10.1111/dom.13389DOI Listing
September 2018

Protein kinase D at the Golgi controls NLRP3 inflammasome activation.

J Exp Med 2017 Sep 17;214(9):2671-2693. Epub 2017 Jul 17.

Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France

The inflammasomes are multiprotein complexes sensing tissue damage and infectious agents to initiate innate immune responses. Different inflammasomes containing distinct sensor molecules exist. The NLRP3 inflammasome is unique as it detects a variety of danger signals. It has been reported that NLRP3 is recruited to mitochondria-associated endoplasmic reticulum membranes (MAMs) and is activated by MAM-derived effectors. Here, we show that in response to inflammasome activators, MAMs localize adjacent to Golgi membranes. Diacylglycerol (DAG) at the Golgi rapidly increases, recruiting protein kinase D (PKD), a key effector of DAG. Upon PKD inactivation, self-oligomerized NLRP3 is retained at MAMs adjacent to Golgi, blocking assembly of the active inflammasome. Importantly, phosphorylation of NLRP3 by PKD at the Golgi is sufficient to release NLRP3 from MAMs, resulting in assembly of the active inflammasome. Moreover, PKD inhibition prevents inflammasome autoactivation in peripheral blood mononuclear cells from patients carrying NLRP3 mutations. Hence, Golgi-mediated PKD signaling is required and sufficient for NLRP3 inflammasome activation.
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http://dx.doi.org/10.1084/jem.20162040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5584123PMC
September 2017

Insulin granules. Insulin secretory granules control autophagy in pancreatic β cells.

Science 2015 Feb;347(6224):878-82

Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM, CNRS, Université de Strasbourg, 67404 Illkirch, France. Nouvel Hôpital Civil, Laboratoire de Biochimie et de Biologie Moléculaire, Université de Strasbourg, 67091 Strasbourg, France.

Pancreatic β cells lower insulin release in response to nutrient depletion. The question of whether starved β cells induce macroautophagy, a predominant mechanism maintaining energy homeostasis, remains poorly explored. We found that, in contrast to many mammalian cells, macroautophagy in pancreatic β cells was suppressed upon starvation. Instead, starved β cells induced lysosomal degradation of nascent secretory insulin granules, which was controlled by protein kinase D (PKD), a key player in secretory granule biogenesis. Starvation-induced nascent granule degradation triggered lysosomal recruitment and activation of mechanistic target of rapamycin that suppressed macroautophagy. Switching from macroautophagy to insulin granule degradation was important to keep insulin secretion low upon fasting. Thus, β cells use a PKD-dependent mechanism to adapt to nutrient availability and couple autophagy flux to secretory function.
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http://dx.doi.org/10.1126/science.aaa2628DOI Listing
February 2015

Comparison of the Oilseed rape mosaic virus and Tobacco mosaic virus movement proteins (MP) reveals common and dissimilar MP functions for tobamovirus spread.

Virology 2014 May 28;456-457:43-54. Epub 2014 Mar 28.

Institut de Biologie Moléculaire des Plantes du CNRS (UPR 2357), Université de Strasbourg, 12 rue du Général Zimmer, 67000 Strasbourg, France; Zürich-Basel Plant Science Center, Department of Environmental Sciences, Botany, University of Basel, Hebelstrasse 1, 4056 Basel, Switzerland. Electronic address:

Tobacco mosaic virus (TMV) is a longstanding model for studying virus movement and macromolecular transport through plasmodesmata (PD). Its movement protein (MP) interacts with cortical microtubule (MT)-associated ER sites (C-MERs) to facilitate the formation and transport of ER-associated viral replication complexes (VRCs) along the ER-actin network towards PD. To investigate whether this movement mechanism might be conserved between tobamoviruses, we compared the functions of Oilseed rape mosaic virus (ORMV) MP with those of MP(TMV). We show that MP(ORMV) supports TMV movement more efficiently than MP(TMV). Moreover, MP(ORMV) localizes to C-MERs like MP(TMV) but accumulates to lower levels and does not localize to larger inclusions/VRCs or along MTs, patterns regularly seen for MP(TMV). Our findings extend the role of C-MERs in viral cell-to-cell transport to a virus commonly used for functional genomics in Arabidopsis. Moreover, accumulation of tobamoviral MP in inclusions or along MTs is not required for virus movement.
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http://dx.doi.org/10.1016/j.virol.2014.03.007DOI Listing
May 2014

Analysis of Tomato spotted wilt virus NSs protein indicates the importance of the N-terminal domain for avirulence and RNA silencing suppression.

Mol Plant Pathol 2014 Feb 5;15(2):185-95. Epub 2013 Dec 5.

Laboratory of Virology, Department of Plant Sciences, Wageningen University, Droevendaalsesteeg 1, 6708 PB, Wageningen, the Netherlands.

Recently, Tomato spotted wilt virus (TSWV) nonstructural protein NSs has been identified unambiguously as an avirulence (Avr) determinant for Tomato spotted wilt (Tsw)-based resistance. The observation that NSs from two natural resistance-breaking isolates had lost RNA silencing suppressor (RSS) activity and Avr suggested a link between the two functions. To test this, a large set of NSs mutants was generated by alanine substitutions in NSs from resistance-inducing wild-type strains (NSs(RI) ), amino acid reversions in NSs from resistance-breaking strains (NSs(RB)), domain deletions and swapping. Testing these mutants for their ability to suppress green fluorescent protein (GFP) silencing and to trigger a Tsw-mediated hypersensitive response (HR) revealed that the two functions can be separated. Changes in the N-terminal domain were found to be detrimental for both activities and indicated the importance of this domain, additionally supported by domain swapping between NSs(RI) and NSs(RB). Swapping domains between the closely related Tospovirus Groundnut ringspot virus (GRSV) NSs and TSWV NSs(RI) showed that Avr functionality could not simply be transferred between species. Although deletion of the C-terminal domain rendered NSs completely dysfunctional, only a few single-amino-acid mutations in the C-terminus affected both functions. Mutation of a GW/WG motif (position 17/18) rendered NSs completely dysfunctional for RSS and Avr activity, and indicated a putative interaction between NSs and Argonaute 1 (AGO1), and its importance in TSWV virulence and viral counter defence against RNA interference.
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http://dx.doi.org/10.1111/mpp.12082DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6638762PMC
February 2014

Micro-MRI study of cerebral aging: ex vivo detection of hippocampal subfield reorganization, microhemorrhages and amyloid plaques in mouse lemur primates.

PLoS One 2013 27;8(2):e56593. Epub 2013 Feb 27.

CNRS (Centre National de la Recherche Scientifique), URA2210 (Unité de Recherche Autonome 2210), Fontenay-aux-Roses, France.

Mouse lemurs are non-human primate models of cerebral aging and neurodegeneration. Much smaller than other primates, they recapitulate numerous features of human brain aging, including progressive cerebral atrophy and correlation between regional atrophy and cognitive impairments. Characterization of brain atrophy in mouse lemurs has been done by MRI measures of regional CSF volume and by MRI measures of regional atrophy. Here, we further characterize mouse lemur brain aging using ex vivo MR microscopy (31 µm in-plane resolution). First, we performed a non-biased, direct volumetric quantification of dentate gyrus and extended Ammon's horn. We show that both dentate gyrus and Ammon's horn undergo an age-related reorganization leading to a growth of the dentate gyrus and an atrophy of the Ammon's horn, even in the absence of global hippocampal atrophy. Second, on these first MR microscopic images of the mouse lemur brain, we depicted cortical and hippocampal hypointense spots. We demonstrated that their incidence increases with aging and that they correspond either to amyloid deposits or to cerebral microhemorrhages.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0056593PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3584101PMC
September 2013
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