Caleosin 1 contributes to seed lipid droplet degradation by interaction with autophagy-related protein ATG8

Triacylglycerols (TAGs) accumulate in lipid droplets (LDs) of seed tissues to provide energy and carbon for seedling establishment. In the major route of LD degradation (lipolysis), TAGs are mobilized by lipases. However, LDs may be also degraded via lipophagy, a type of selective autophagy, which mediates LDs delivery to vacuoles or lysosomes. The exact mechanism of this process in plants still remains unresolved. Here, we provide evidence that during Arabidopsis thaliana seed germination, LDs are degraded by microlipophagy and that this process requires caleosin 1 (CLO1), a LD surface protein. We show co-localization of autophagy-related protein 8b (ATG8b) and LDs during seed germination and localization of lipidated ATG8 (ATG8-PE) to the LD fraction. We further demonstrate that CLO1, CLO2 and CLO3 interact with ATG8 proteins via their ATG8-interacting motifs (AIMs). Deletion of AIM localized directly before the proline knot disrupts CLO1 interaction with ATG8b, suggesting the essential role of this region in the interaction between the two proteins. Collectively, we provide new insights into the molecular mechanisms governing the interaction of LDs with the autophagy machinery in plant cells, contributing to understanding of the role of structural LD proteins in lipid mobilization.

[1]  K. Chapman,et al.  Finding new friends and revisiting old ones - How plant lipid droplets connect with other subcellular structures. , 2022, The New phytologist.

[2]  Meng Zhang,et al.  Multiple caleosins have overlapping functions in oil accumulation and embryo development. , 2022, Journal of experimental botany.

[3]  Changcheng Xu,et al.  Links between autophagy and lipid droplet dynamics. , 2022, Journal of experimental botany.

[4]  Qingjun Xie,et al.  ATG8-Interacting Motif: Evolution and Function in Selective Autophagy of Targeting Biological Processes , 2021, Frontiers in Plant Science.

[5]  N. Sheikh,et al.  A Decade of Mighty Lipophagy: What We Know and What Facts We Need to Know? , 2021, Oxidative medicine and cellular longevity.

[6]  K. Zienkiewicz,et al.  Lipid metabolism and accumulation in oilseed crops , 2021, OCL.

[7]  T. Kuroiwa,et al.  Ultrastructural characterization of microlipophagy induced by the interaction of vacuoles and lipid bodies around generative and sperm cells in Arabidopsis pollen , 2020, Protoplasma.

[8]  T. Ischebeck,et al.  Ties between Stress and Lipid Droplets Pre-date Seeds. , 2020, Trends in plant science.

[9]  K. Zienkiewicz,et al.  Degradation of Lipid Droplets in Plants and Algae—Right Time, Many Paths, One Goal , 2020, Frontiers in Plant Science.

[10]  Sebastian Schuck.  Microautophagy – distinct molecular mechanisms handle cargoes of many sizes , 2020, Journal of Cell Science.

[11]  W. Araújo,et al.  Multifaceted Roles of Plant Autophagy in Lipid and Energy Metabolism. , 2020, Trends in plant science.

[12]  Wei Huang,et al.  Multiple Functions of ATG8 Family Proteins in Plant Autophagy , 2020, Frontiers in Cell and Developmental Biology.

[13]  S. Martens,et al.  Activation and targeting of ATG8 protein lipidation , 2020, Cell Discovery.

[14]  S. Singer,et al.  The Role of Triacylglycerol in Plant Stress Response , 2020, Plants.

[15]  M. Hamberg,et al.  Protein Profiles of Lipid Droplets During the Hypersensitive Defense Response of Arabidopsis Against Pseudomonas Infection. , 2020, Plant & cell physiology.

[16]  R. Vierstra,et al.  Reticulon proteins modulate autophagy of the endoplasmic reticulum in maize endosperm , 2020, eLife.

[17]  E. Baehrecke,et al.  Autophagy in animal development , 2020, Cell Death & Differentiation.

[18]  Meng-Xiang Sun,et al.  Autophagy-mediated compartmental cytoplasmic deletion is essential for tobacco pollen germination and male fertility , 2020, Autophagy.

[19]  T. Lamark,et al.  Selective Autophagy: ATG8 Family Proteins, LIR Motifs and Cargo Receptors. , 2020, Journal of molecular biology.

[20]  Yasin F. Dagdas,et al.  Plant Selective Autophagy - Still an uncharted territory with a lot of hidden gems. , 2020, Journal of molecular biology.

[21]  Qun Shao,et al.  New Insights Into the Role of Seed Oil Body Proteins in Metabolism and Plant Development , 2019, Front. Plant Sci..

[22]  I. Feussner,et al.  Disruption of Arabidopsis neutral ceramidases 1 and 2 results in specific sphingolipid imbalances triggering different phytohormone-dependent plant cell death programs. , 2019, The New phytologist.

[23]  E. M. Farré,et al.  The Microalga Nannochloropsis during Transition from Quiescence to Autotrophy in Response to Nitrogen Availability1[OPEN] , 2019, Plant Physiology.

[24]  Changcheng Xu,et al.  Dual Role for Autophagy in Lipid Metabolism in Arabidopsis. , 2019, The Plant cell.

[25]  R. Vierstra,et al.  ATG8-Binding UIM Proteins Define a New Class of Autophagy Adaptors and Receptors , 2019, Cell.

[26]  Sudhir Kumar,et al.  MEGA X: Molecular Evolutionary Genetics Analysis across Computing Platforms. , 2018, Molecular biology and evolution.

[27]  W. Sakamoto,et al.  Selective Elimination of Membrane-Damaged Chloroplasts via Microautophagy1[OPEN] , 2018, Plant Physiology.

[28]  R. Vierstra,et al.  Autophagy: The Master of Bulk and Selective Recycling. , 2018, Annual review of plant biology.

[29]  D. Bassham,et al.  New advances in autophagy in plants: Regulation, selectivity and function. , 2017, Seminars in cell & developmental biology.

[30]  Rebecca L. Roston,et al.  Recovery from N Deprivation Is a Transcriptionally and Functionally Distinct State in Chlamydomonas1[OPEN] , 2017, Plant Physiology.

[31]  A. Huang.  Plant Lipid Droplets and Their Associated Proteins: Potential for Rapid Advances1[OPEN] , 2017, Plant Physiology.

[32]  Ben Zhang,et al.  Mating Based Split-ubiquitin Assay for Detection of Protein Interactions. , 2017, Bio-protocol.

[33]  J. Lippincott-Schwartz,et al.  AMPK and vacuole-associated Atg14p orchestrate μ-lipophagy for energy production and long-term survival under glucose starvation , 2017, eLife.

[34]  Yasin F. Dagdas,et al.  ATG8 Expansion: A Driver of Selective Autophagy Diversification? , 2017, Trends in plant science.

[35]  K. Chapman,et al.  Two Acyltransferases Contribute Differently to Linolenic Acid Levels in Seed Oil1[OPEN] , 2017, Plant Physiology.

[36]  A. Hanano,et al.  Specific Caleosin/Peroxygenase and Lipoxygenase Activities Are Tissue-Differentially Expressed in Date Palm (Phoenix dactylifera L.) Seedlings and Are Further Induced Following Exposure to the Toxin 2,3,7,8-tetrachlorodibenzo-p-dioxin , 2017, Front. Plant Sci..

[37]  Lili Wang,et al.  GENOME-WIDE CHARACTERIZATION AND PHYLOGENETIC AND EXPRESSION ANALYSES OF THE CALEOSIN GENE FAMILY IN SOYBEAN, COMMON BEAN AND BARREL MEDIC , 2016 .

[38]  Changcheng Xu,et al.  Triacylglycerol Metabolism, Function, and Accumulation in Plant Vegetative Tissues. , 2016, Annual review of plant biology.

[39]  Elsje G. Otten,et al.  Autophagy, lipophagy and lysosomal lipid storage disorders. , 2016, Biochimica et biophysica acta.

[40]  D. Andrews,et al.  Lipid Droplet-Associated Proteins (LDAPs) Are Required for the Dynamic Regulation of Neutral Lipid Compartmentation in Plant Cells1 , 2016, Plant Physiology.

[41]  Sangeeta Khare,et al.  Guidelines for the use and interpretation of assays formonitoring autophagy (3rd edition) , 2016 .

[42]  R. Chan,et al.  Role for Lipid Droplet Biogenesis and Microlipophagy in Adaptation to Lipid Imbalance in Yeast. , 2015, Developmental cell.

[43]  T. Chardot,et al.  N-terminus of seed caleosins is essential for lipid droplet sorting but not for lipid accumulation. , 2015, Archives of biochemistry and biophysics.

[44]  I. Hara-Nishimura,et al.  Leaf oil bodies are subcellular factories producing antifungal oxylipins. , 2015, Current opinion in plant biology.

[45]  A. Fernie,et al.  Global Analysis of the Role of Autophagy in Cellular Metabolism and Energy Homeostasis in Arabidopsis Seedlings under Carbon Starvation[OPEN] , 2015, Plant Cell.

[46]  Li Zhao,et al.  Autophagy-like processes are involved in lipid droplet degradation in Auxenochlorella protothecoides during the heterotrophy-autotrophy transition , 2014, Front. Plant Sci..

[47]  I. Feussner,et al.  The Reductase Activity of the Arabidopsis Caleosin RESPONSIVE TO DESSICATION20 Mediates Gibberellin-Dependent Flowering Time, Abscisic Acid Sensitivity, and Tolerance to Oxidative Stress1[W] , 2014, Plant Physiology.

[48]  Meng Zhang,et al.  Genomic analysis and expression investigation of caleosin gene family in Arabidopsis. , 2014, Biochemical and biophysical research communications.

[49]  Kazuki Saito,et al.  OsATG7 is required for autophagy-dependent lipid metabolism in rice postmeiotic anther development , 2014, Autophagy.

[50]  V. Promponas,et al.  iLIR , 2014, Autophagy.

[51]  D. Klionsky,et al.  An overview of autophagy: morphology, mechanism, and regulation. , 2014, Antioxidants & redox signaling.

[52]  S. Kohlwein,et al.  Lipid droplet autophagy in the yeast Saccharomyces cerevisiae , 2014, Molecular biology of the cell.

[53]  Yingkao Hu,et al.  Delineation of plant caleosin residues critical for functional divergence, positive selection and coevolution , 2014, BMC Evolutionary Biology.

[54]  D. Klionsky,et al.  Autophagic Processes in Yeast: Mechanism, Machinery and Regulation , 2013, Genetics.

[55]  R. Vierstra,et al.  Autophagy: a multifaceted intracellular system for bulk and selective recycling. , 2012, Trends in plant science.

[56]  Kevin W Eliceiri,et al.  NIH Image to ImageJ: 25 years of image analysis , 2012, Nature Methods.

[57]  Robert Clarke,et al.  Guidelines for the use and interpretation of assays for monitoring autophagy , 2012 .

[58]  K. Chapman,et al.  Biogenesis and functions of lipid droplets in plants , 2012, Journal of Lipid Research.

[59]  J. Ohlrogge,et al.  Compartmentation of Triacylglycerol Accumulation in Plants* , 2011, The Journal of Biological Chemistry.

[60]  A. Kelly,et al.  Seed Storage Oil Mobilization Is Important But Not Essential for Germination or Seedling Establishment in Arabidopsis1[W] , 2011, Plant Physiology.

[61]  I. Feussner,et al.  The lipoxygenase-dependent oxygenation of lipid body membranes is promoted by a patatin-type phospholipase in cucumber cotyledons , 2010, Journal of experimental botany.

[62]  F. Inagaki,et al.  Atg8‐family interacting motif crucial for selective autophagy , 2010, FEBS letters.

[63]  I. Graham.  Seed storage oil mobilization. , 2008, Annual review of plant biology.

[64]  Christopher Grefen,et al.  Split‐Ubiquitin System for Identifying Protein‐Protein Interactions in Membrane and Full‐Length Proteins , 2007, Current protocols in neuroscience.

[65]  P. Lipovová,et al.  Structural properties of caleosin: a MS and CD study. , 2007, Archives of biochemistry and biophysics.

[66]  J. Browse,et al.  A role for caleosin in degradation of oil-body storage lipid during seed germination. , 2006, The Plant journal : for cell and molecular biology.

[67]  Rodrigo M. P. Siloto,et al.  The Accumulation of Oleosins Determines the Size of Seed Oilbodies in Arabidopsis[W][OA] , 2006, The Plant Cell Online.

[68]  P. Eastmond.  SUGAR-DEPENDENT1 Encodes a Patatin Domain Triacylglycerol Lipase That Initiates Storage Oil Breakdown in Germinating Arabidopsis Seeds[W] , 2006, The Plant Cell Online.

[69]  C. Pikaard,et al.  Gateway-compatible vectors for plant functional genomics and proteomics. , 2006, The Plant journal : for cell and molecular biology.

[70]  B. André,et al.  K+ channel interactions detected by a genetic system optimized for systematic studies of membrane protein interactions. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[71]  H. Nielsen,et al.  Caleosins: Ca2+-binding proteins associated with lipid bodies , 2000, Plant Molecular Biology.

[72]  J. Tzen,et al.  An in vitro system to examine the effective phospholipids and structural domain for protein targeting to seed oil bodies. , 2001, Plant & cell physiology.

[73]  S. Clough,et al.  Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. , 1998, The Plant journal : for cell and molecular biology.