@article {67339, title = {Lipid droplets in the nervous system.}, journal = {Journal of Cell Biology}, volume = {220}, year = {2021}, month = {2021 Jul 05}, abstract = {

Lipid droplets are dynamic intracellular lipid storage organelles that respond to the physiological state of cells. In addition to controlling cell metabolism, they play a protective role for many cellular stressors, including oxidative stress. Despite prior descriptions of lipid droplets appearing in the brain as early as a century ago, only recently has the role of lipid droplets in cells found in the brain begun to be understood. Lipid droplet functions have now been described for cells of the nervous system in the context of development, aging, and an increasing number of neuropathologies. Here, we review the basic mechanisms of lipid droplet formation, turnover, and function and discuss how these mechanisms enable lipid droplets to function in different cell types of the nervous system under healthy and pathological conditions.

}, issn = {1540-8140}, doi = {10.1083/jcb.202102136}, author = {Ralhan, Isha and Chang, Chi-Lun and Lippincott-Schwartz, Jennifer and Ioannou, Maria S} } @article {65658, title = {Microdomains form on the luminal face of neuronal extracellular vesicle membranes.}, journal = {Scientific Reports}, volume = {10}, year = {2020}, month = {2020 Jul 20}, pages = {11953}, abstract = {

Extracellular vesicles (EVs) are important mediators of cell-to-cell communication and have been implicated in several pathologies including those of the central nervous system. They are released by all cell types, including neurons, and are highly heterogenous in size and composition. Yet much remains unknown regarding the biophysical characteristics of different EVs. Here, using cryo-electron microscopy (cryoEM), we analyzed the size distribution and morphology of EVs released from primary cortical neurons. We discovered massive macromolecular clusters on the luminal face of EV membranes. These clusters are predominantly found on medium-sized vesicles, suggesting that they may be specific to microvesicles as opposed to exosomes. We propose that these clusters serve as microdomains for EV signaling and play an important role in EV physiology.

}, issn = {2045-2322}, doi = {10.1038/s41598-020-68436-x}, author = {Matthies, Doreen and Lee, Nathanael Y J and Gatera, Ian and Pasolli, H Amalia and Zhao, Xiaowei and Liu, Hui and Walpita, Deepika and Liu, Zhe and Yu, Zhiheng and Ioannou, Maria S} } @article {65067, title = {Neuron-astrocyte metabolic coupling protects against activity-induced fatty acid toxicity.}, journal = {Cell}, volume = {177}, year = {2019}, month = {2019 May 30}, pages = {1522-1535.e14}, abstract = {

Metabolic coordination between neurons and astrocytes is critical for the health of the brain. However, neuron-astrocyte coupling of lipid metabolism, particularly in response to neural activity, remains largely uncharacterized. Here, we demonstrate that toxic fatty acids (FAs) produced in hyperactive neurons are transferred to astrocytic lipid droplets by ApoE-positive lipid particles. Astrocytes consume the FAs stored in lipid droplets via mitochondrial β-oxidation in response to neuronal activity and turn on a detoxification gene expression program. Our findings reveal that FA metabolism is coupled in neurons and astrocytes to protect neurons from FA toxicity during periods of enhanced activity. This coordinated mechanism for metabolizing FAs could underlie both homeostasis and a variety of disease states of the brain.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2019.04.001}, author = {Ioannou, Maria S and Jackson, Jesse and Sheu, Shu-Hsien and Chang, Chi-Lun and Weigel, Aubrey V and Liu, Hui and Pasolli, H Amalia and Xu, C Shan and Pang, Song and Matthies, Doreen and Hess, Harald F and Lippincott-Schwartz, Jennifer and Liu, Zhe} } @article {65139, title = {A neuron-glia Co-culture system for studying intercellular lipid transport.}, journal = {Curr Protoc Cell Biol}, volume = {84}, year = {2019}, month = {2019 Sep 01}, pages = {e95}, abstract = {

Neurons and glia operate in a highly coordinated fashion in the brain. Although glial cells have long been known to supply lipids to neurons via lipoprotein particles, new evidence reveals that lipid transport between neurons and glia is bidirectional. Here, we describe a co-culture system to study transfer of lipids and lipid-associated proteins from neurons to glia. The assay entails culturing neurons and glia on separate coverslips, pulsing the neurons with fluorescently labeled fatty acids, and then incubating the coverslips together. As astrocytes internalize and store neuron-derived fatty acids in lipid droplets, analyzing the number, size, and fluorescence intensity of lipid droplets containing the fluorescent fatty acids provides an easy and quantifiable measure of fatty acid transport. {\textcopyright} 2019 The Authors.

}, issn = {1934-2616}, doi = {10.1002/cpcb.95}, author = {Ioannou, Maria S and Liu, Zhe and Lippincott-Schwartz, Jennifer} } @article {64795, title = {Spastin tethers lipid droplets to peroxisomes and directs fatty acid trafficking through ESCRT-III.}, journal = {Journal of Cell Biology}, volume = {218}, year = {2019}, month = {2019 Jun 21}, pages = {2583-99}, abstract = {

Lipid droplets (LDs) are neutral lipid storage organelles that transfer lipids to various organelles including peroxisomes. Here, we show that the hereditary spastic paraplegia protein M1 Spastin, a membrane-bound AAA ATPase found on LDs, coordinates fatty acid (FA) trafficking from LDs to peroxisomes through two inter-related mechanisms. First, M1 Spastin forms a tethering complex with peroxisomal ABCD1 to promote LD-peroxisome contact formation. Second, M1 Spastin recruits the membrane-shaping ESCRT-III proteins IST1 and CHMP1B to LDs via its MIT domain to facilitate LD-to-peroxisome FA trafficking, possibly through IST1 and CHMP1B modifying LD membrane morphology. Furthermore, M1 Spastin, IST1 and CHMP1B are all required to relieve LDs of lipid peroxidation. The roles of M1 Spastin in tethering LDs to peroxisomes and in recruiting ESCRT-III components to LD-peroxisome contact sites for FA trafficking may help explain the pathogenesis of diseases associated with defective FA metabolism in LDs and peroxisomes.

}, doi = {10.1101/544023}, author = {Chang, Chi-Lun and Weigel, Aubrey V and Ioannou, Maria S and Pasolli, H Amalia and Xu, C Shan and Peale, David R and Shtengel, Gleb and Freeman, Melanie and Hess, Harald F and Blackstone, Craig and Lippincott-Schwartz, Jennifer} } @article {49382, title = {Macropinosome formation by tent pole ruffling in macrophages.}, journal = {The Journal of Cell Biology}, volume = {217}, year = {2018}, month = {2018 Aug 27}, pages = {3873-85}, abstract = {

Pathogen-mediated activation of macrophages arms innate immune responses that include enhanced surface ruffling and macropinocytosis for environmental sampling and receptor internalization and signaling. Activation of macrophages with bacterial lipopolysaccharide (LPS) generates prominent dorsal ruffles, which are precursors for macropinosomes. Very rapid, high-resolution imaging of live macrophages with lattice light sheet microscopy (LLSM) reveals new features and actions of dorsal ruffles, which redefine the process of macropinosome formation and closure. We offer a new model in which ruffles are erected and supported by F-actin tent poles that cross over and twist to constrict the forming macropinosomes. This process allows for formation of large macropinosomes induced by LPS. We further describe the enrichment of active Rab13 on tent pole ruffles and show that CRISPR deletion of Rab13 results in aberrant tent pole ruffles and blocks the formation of large LPS-induced macropinosomes. Based on the exquisite temporal and spatial resolution of LLSM, we can redefine the ruffling and macropinosome processes that underpin innate immune responses.

}, issn = {1540-8140}, doi = {10.1083/jcb.201804137}, author = {Condon, Nicholas D and Heddleston, John M and Chew, Teng-Leong and Luo, Lin and McPherson, Peter S and Ioannou, Maria S and Hodgson, Louis and Stow, Jennifer L and Wall, Adam A} } @article {49085, title = {Model-free quantification and visualization of colocalization in fluorescence images.}, journal = {Cytometry. Part A : the journal of the International Society for Analytical Cytology}, year = {2018}, month = {2018 Mar 13}, abstract = {

The spatial association between fluorescently tagged biomolecules in situ provides valuable insight into their biological relationship. Within the limits of diffraction, such association can be measured using either Pearson{\textquoteright}s Correlation Coefficient (PCC) or Spearman{\textquoteright}s Rank Coefficient (SRC), which are designed to measure linear and monotonic correlations, respectively. However, the relationship between real biological signals is often more complex than these measures assume, rendering their results difficult to interpret. Here, we have adapted methods from the field of information theory to measure the association between two probes{\textquoteright} concentrations based on their statistical dependence. Our approach is mathematically more general than PCC or SRC, making no assumptions about the type of relationship between the probes. We show that when applied to biological images, our measures provide more intuitive results that are also more robust to outliers and the presence of multiple relationships than PCC or SRC. We also devise a display technique to highlight regions in the input images where the probes{\textquoteright} association is higher versus lower. We expect that our methods will allow biologists to more accurately and robustly quantify and visualize the association between two probes in a pair of fluorescence images. {\textcopyright} 2018 International Society for Advancement of Cytometry.

}, issn = {1552-4930}, doi = {10.1002/cyto.a.23356}, author = {Taylor, Aaron B and Ioannou, Maria S and Aaron, Jesse and Chew, Teng-Leong} } @article {48346, title = {Stochastic protein labeling enables long-term single molecule observation in vivo.}, journal = {bioRxiv}, year = {2017}, month = {2017 Mar 13}, abstract = {Our ability to unambiguously image and track individual molecules in live cells is limited by packing of multiple copies of labeled molecules within the resolution limit. Here we devise a universal genetic strategy to precisely control protein copy number in a cell. This system has a dynamic titration range of more than 10,000 fold, enabling sparse labeling of proteins expressed at widely different levels. Combined with fluorescence signal amplification tags, this system extends the duration of automated single-molecule tracking by 2 orders of magnitude. We demonstrate long-term imaging of synaptic vesicle dynamics in cultured neurons as well as in live zebrafish. We found that axon initial segment utilizes a waterfall mechanism gating synaptic vesicle transport polarity by promoting anterograde transport processivity. Long-time observation also reveals that transcription factor Sox2 samples clustered binding sites in spatially-restricted sub-nuclear regions, suggesting that topological structures in the nucleus shape local gene activities by a sequestering mechanism. This strategy thus greatly expands the spatiotemporal length scales of live-cell single-molecule measurements for a quantitative understanding of complex control of molecular dynamics in vivo.}, doi = {10.1101/116186}, author = {Liu, Hui and Dong, Peng and Ioannou, Maria S and Li, Li and Shea, Jamien and Pasolli, H Amalia and Grimm, Jonathan B and Rivlin, Patricia K and Lavis, Luke D and Koyama, Minoru and Liu, Zhe} } @article {48752, title = {Visualizing long-term single-molecule dynamics in vivo by stochastic protein labeling.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {115}, year = {2017}, month = {2017 Jan 09}, pages = {343-8}, abstract = {

Our ability to unambiguously image and track individual molecules in live cells is limited by packing of multiple copies of labeled molecules within the resolution limit. Here we devise a universal genetic strategy to precisely control copy number of fluorescently labeled molecules in a cell. This system has a dynamic titration range of \>10,000 fold, enabling sparse labeling of proteins expressed at different abundance levels. Combined with photostable labels, this system extends the duration of automated single-molecule tracking by 2 orders of magnitude. We demonstrate long-term imaging of synaptic vesicle dynamics in cultured neurons as well as in intact\ zebrafish. We found axon initial segment utilizes a "waterfall" mechanism gating synaptic vesicle transport polarity by promoting anterograde transport processivity. Long-time observation also reveals that transcription factor hops between clustered binding sites in spatially-restricted sub-nuclear regions, suggesting that topological structures in the nucleus shape local gene activities by a sequestering mechanism. This strategy thus greatly expands the spatiotemporal length scales of live-cell single-molecule measurements, enabling new experiments to quantitatively understand complex control of molecular dynamics\ in vivo.

}, doi = {10.1073/pnas.1713895115}, author = {Liu, Hui and Dong, Peng and Ioannou, Maria S and Li, Li and Shea, Jamien and Pasolli, H Amalia and Grimm, Jonathan B and Rivlin, Patricia K and Lavis, Luke D and Koyama, Minoru and Liu, Zhe} }