Inhibition of Very Long Chain Fatty Acids Synthesis Mediates PI3P Homeostasis at Endosomal Compartments

Abstract

A main characteristic of sphingolipids is the presence of a very long chain fatty acid (VLCFA) whose function in cellular processes is not yet fully understood. VLCFAs of sphingolipids are involved in the intracellular traffic to the vacuole and the maturation of early endosomes into late endosomes is one of the major pathways for vacuolar traffic. Additionally, the anionic phospholipid phosphatidylinositol-3-phosphate (PtdIns (3)P or PI3P) is involved in protein sorting and recruitment of small GTPase effectors at late endosomes/multivesicular bodies (MVBs) during vacuolar trafficking. In contrast to animal cells, PI3P mainly localizes to late endosomes in plant cells and to a minor extent to a discrete sub-domain of the plant’s early endosome (EE)/trans-Golgi network (TGN) where the endosomal maturation occurs. However, the mechanisms that control the relative levels of PI3P between TGN and MVBs are unknown. Using metazachlor, an inhibitor of VLCFA synthesis, we found that VLCFAs are involved in the TGN/MVB distribution of PI3P. This effect is independent from either synthesis of PI3P by PI3-kinase or degradation of PI(3,5)P2 into PI3P by the SUPPRESSOR OF ACTIN1 (SAC1) phosphatase. Using high-resolution live cell imaging microscopy, we detected transient associations between TGNs and MVBs but VLCFAs are not involved in those interactions. Nonetheless, our results suggest that PI3P might be transferable from TGN to MVBs and that VLCFAs act in this process.

Crossing paths: recent insights in the interplay between autophagy and intracellular trafficking in plants

“…Lipid identity has been shown to be consequential in many steps of the autophagy pathway [67]. A noteworthy example of the lipid‐associated implications and crosstalk that may occur between the autophagy pathway and vesicle trafficking is the role of the length of very‐long chain fatty acids in mediating PI3P homeostasis, a crucial lipid for both autophagosome biogenesis and LE/MVB identity [68] (Fig. 2).…”

ER-to-Golgi traffickingviaa dynamic intermediatecis-Golgi tubular network in Arabidopsis

“…The dynamic analysis was done using the Detect, Track, and Colocalize (DTC) plugin of ImageJ (https://github.com/fabricecordelieres/IJ-Plugin_DTC) 27 . Images were pre-treated as follow before being analysed with the DTC: individual cells were manually delimited using the polygon selection tool and added to the ROI manager; a Gaussian blur filter (sigma 2) and a subtract background (rolling radius 10, except for p35S::NAG1-EGFP were the rolling radius was 5) were applied to all images.…”

“…Quantitative live tracking of compartment association. The dynamic analysis was done using the Detect, Track, and Colocalize (DTC) plugin of ImageJ (https://github.com/fabricecordelieres/IJ-Plugin_DTC) 27 .…”

“…Thus, our next question was to address how these independent cis -Golgi behave dynamically. To address this question, we setup a workflow composed of lived airyscan microscopy acquisition, tracking of individual objects in both channels using the Detect Track and Co-localize (DTC) plugin of imageJ 27 . This plug-in identifies the interactions occurring between compartments of two channels and calculates their durations.…”

“…A.N provided scientific input and initial materials to this study. Y.I setup the initial airyscan live acquisition and quantification 27 , performed the cloning and solved the initial issues of the RUSH system in Arabidopsis and conceptualized the research. Y.B conceptualized and designed the research, got the financial support for, supervised all aspects of this study and wrote the manuscript.…”