Membrane Dynamics and Intracellular Trafficking

CATHY JACKSON & JEAN-MARC VERBAVATZ

Eukaryotic cells are characterized by their internal membrane compartments (organelles), which allow different cellular processes to take place in a well-adapted environment. However, in order for the cell to function as a whole, the various membrane-bound organelles, as well as the plasma membrane, must communicate through vesicular and non-vesicular trafficking. Our research focuses on intracellular lipid trafficking pathways. Their disruption leads to various human pathologies such as cancer, diabetes and neurodegenerative diseases, and these pathways are also subverted for the propagation of numerous viruses, including Hepatitis C and SARS-CoV-2.

Keywords : Organelle, membrane trafficking, membrane contact site, polarity, lipid, endoplasmic reticulum, Golgi, plasma membrane

+33 (0)157278004 /  +33 (0)157278004     cathy.jackson(at)ijm.fr / jean-marc.verbavatz(at)ijm.fr

Eukaryotic cells are characterized by their internal membrane compartments (organelles), which allow different cellular processes to take place in a well-adapted environment. However, in order for the cell to function as a whole, the various membrane-bound organelles, including the endoplasmic reticulum (ER), Golgi apparatus, endosomes, lipid droplets, and mitochondria, as well as the plasma membrane, communicate through vesicular and non-vesicular trafficking (Jackson, Walch and Verbavatz, 2016; Kaczmarek, Verbavatz and Jackson, 2017; Jackson 2019). In vesicular trafficking, vesicles are transported from one compartment to their target, whereas non vesicular trafficking occurs at sites of close contact between organelles, named membrane contact sites (MCS), where the membranes of both organelles come together at a distance of 10-30 nm without fusing. Lipids can be transported by both vesicular and non-vesicular routes, but maintenance of the lipid composition of membranes requires lipid transfer at MCS (Jackson, Walch and Verbavatz, 2016). Disruption of lipid trafficking pathways leads to various human pathologies such as cancer, diabetes and neurodegenerative diseases, and these pathways are also subverted for the propagation of numerous viruses, including Hepatitis C and SARS-CoV-2.

Figure 1. Diagram illustrating the major organelles and membrane contact sites in a typical eukaryotic cell. ER, endoplasmic reticulum; PM, plasma membrane; LD, lipid droplet (Adapted from Jackson, Walch and Verbavatz, 2016).

 

We are using a multidisciplinary approach consisting of advanced imaging (light and electron microscopy, live cell imaging), biochemical approaches and proteomics analyses, to explore the following research questions:

 

  1. Arf family GTPase regulation of organelle dynamics.

Arf family proteins are crucial regulators of membrane dynamics in all eukaryotic cells, having essential functions at multiple organelles, in vesicular trafficking and at MCS. For a number of years, our group has studied the small G protein (GTPase) ADP ribosylation factor 1 (Arf1) and its activators (Donaldson and Jackson 2011), uncovering new functions for these regulators.

            Organelle dynamics in cells depends on microtubule motors that bind membrane compartments through adaptor proteins and pull them towards one or the other end of a microtubule. We discovered that the small G protein Arf1 and its activator GBF1 regulate the transport of mitochondria along microtubules and their spatial organization (Walch et al. 2018).

Lipid droplets (LDs) are the major energy storage depots of eukaryotic cells, which interface with membrane trafficking pathways. Our group uncovered a crucial role of Arf1 and its regulators in recruitment of LD-associated proteins to the LD surface (Donaldson and Jackson 2011, Bouvet et al. 2013, Jackson 2019). We have also described the mechanisms by which amphipathic helix-containing proteins such as perilipins are targeted to the LD surface (Donaldson and Jackson 2011, Copic et al. 2018).

            In an exciting recent development, Arf-related proteins have been identified in newly discovered archaea lineages (the Asgard archaea), suggesting that these proteins were present in the common ancestor of archaea and eukaryotes. Thus Asgard archaea appear to be the long-sought lineage from which eukaryotes emerged, i.e. the host that entered into endosymbiosis with the mitochondrial progenitor. We are collaborating with experts in bioinformatics, biophysics and structural biology, to determine if these predicted Asgard Arf-related proteins possess the hallmark features of eukaryotic Arf family GTPases, and to explore their potential roles in eukaryogenesis.

 

 

  1. Physiological functions of endoplasmic reticulum (ER) – organelle membrane contact sites (MCS)

Our studies have demonstrated functions of MCS in lipid transport from the ER to the plasma membrane, and their potential role in membrane growth (Moser von Filseck et al. 2015, Petkovic et al. 2014). The transport of lipids, including phosphoinositides at MCS, is an important determinant of the specific lipid composition of cell membranes (Jackson, Walch and Verbavatz, 2016), and it plays important roles in cellular rearrangements that occur during fundamental cell processes. Our group is currently studying the regulation and functions of MCS in processes such as cell division, cell migration, cell-cell contacts and apico-basal polarity and their implication in human disease.

 

2.1 MCS in cell division

Dramatic reorganizations of cellular organelles occur during cell division, essential for its successful completion. Improper cell division can result in numerous abnormalities, including aneuploidy and cancer. During division, membrane organelles undergo multiple steps of disassembly (nuclear membrane, Golgi apparatus…), fragmentation (mitochondria…) and spatial reorganization (mitochondria, ER…), followed by their reassembly at the end of mitosis. This is accompanied by coordinated changes in the distribution of phosphoinositides at the plasma membrane. We are studying the dynamic rearrangements of MCS in this process, the regulation of their function in lipid transport and their role in the choreography of membrane organelles during cell division.

 

2.2 MCS in epithelial cell polarity

The establishment and maintenance of cell polarity are crucial for the functions of epithelia, and are misregulated in human diseases such as cancer. In polarized epithelial cells, phosphoinositides are important determinants of the apical and basolateral plasma membrane identities. For example, the phosphoinositide PI(3,4,5)P3 is localized to the basolateral plasma membrane, but excluded from the apical membrane. Our goal is to decipher the role of MCS in determining the lipid composition of apical and basolateral membranes of epithelial cells and in the establishment and maintenance of cell polarity. Using cell biology approaches, we are investigating the membrane distribution of protein complexes at MCS, including lipid transfer proteins, and their function in the establishment of cell polarity.

Figure 2.  Junction between two epithelial MDCK cells showing the presence of multiple ER-PM contact sites (MCS, yellow arrows in the inset, right panel). ER, endoplasmic reticulum; PM, plasma membrane. Photos, L. Daunas and V. Proux-Gillardeaux.

 

2.3 MCS in cell motility

Cell motility highly relies on the distribution of phosphoinositides that act as lipidic messengers mediating cytoskeleton reshaping and adhesion dynamics. We investigate the role of VAPA, an ER-resident MCS tether, during cancer cell motility processes including collective and single cell migration, cell adhesion and mecano-sensing. Using high-resolution microscopy, quantitative imaging and optogenetic tools, we intend to define in space and time how lipid exchange taking place at VAPA-mediated MCS controls cell motility.

Figure 3. A. Colon adenocarcinoma Caco-2 cells migrating during a wound healing experiment. Here, a leader cell has been tracked for several hours (blue line). B. ER-PM contact sites identified by electron microscopy at the leading edge of Caco-2 migrating cells (yellow boxes). C. We use fluorescent probes to study the dynamics of MCS during cell motility. The picture has been taken at the tip of the leading edge of a migrating cell (green : ER marker ; magenta : ER-PM contact sites). Photos, M. Heuzé and H. Siegfried

 

Group Leaders:

Cathy JACKSON
Téléphone : +33 (0)157278004 / cathy.jackson (at) ijm.fr

Jean-Marc VERBAVATZ
Téléphone : +33 (0)157278004 / jean-marc.verbavatz (at) ijm.fr

 

Members :

DAUNAS Lucien Engineer in biology
HEUZÉ Mélina Researcher
JACKSON Cathy Researcher
PROUX-GILLARDEAUX Véronique Researcher
SIEGFRIED Hugo PhD Student
VERBAVATZ Jean-Marc Researcher
VERRAES Agathe Assistant engineer in biology
VERTUEUX Anais PhD Student

Lipid droplet biogenesis.

Jackson CL. Curr Opin Cell Biol. 2019 Aug;59:88-96. doi: 10.1016/j.ceb.2019.03.018. Epub 2019 May 7. PMID: 31075519 Review.

Correlative single-molecule localization microscopy and electron tomography reveals endosome nanoscale domains.

Franke C, Repnik U, Segeletz S, Brouilly N, Kalaidzidis Y, Verbavatz JM, Zerial M. Traffic. 2019 Aug;20(8):601-617. doi: 10.1111/tra.12671. PMID: 31206952

GBF1 and Arf1 interact with Miro and regulate mitochondrial positioning within cells.

Walch L, Pellier E, Leng W, Lakisic G, Gautreau A, Contremoulins V, Verbavatz JM#, Jackson CL#. Sci Rep. 2018 Nov 20;8(1):17121. doi: 10.1038/s41598-018-35190-0. PMID: 30459446 #co-corresponding authors

A giant amphipathic helix from a perilipin that is adapted for coating lipid droplets.

Čopič A, Antoine-Bally S, Giménez-Andrés M, La Torre Garay C, Antonny B, Manni MM, Pagnotta S, Guihot J, Jackson CL.Nat Commun. 2018 Apr 6;9(1):1332. doi: 10.1038/s41467-018-03717-8.PMID: 29626194

GBF1 and Arf1 function in vesicular trafficking, lipid homoeostasis and organelle dynamics.

Kaczmarek B, Verbavatz JM, Jackson CL. Biol Cell. 2017 Dec;109(12):391-399. doi: 10.1111/boc.201700042. Epub 2017 Nov 6. PMID: 28985001

Lipids and Their Trafficking: An Integral Part of Cellular Organization.

Jackson CL, Walch L, Verbavatz JM. Dev Cell. 2016 Oct 24;39(2):139-153. doi: 10.1016/j.devcel.2016.09.030. PMID: 27780039 Review.

The SNARE Sec22b has a non-fusogenic function in plasma membrane expansion.

Petkovic M, Jemaiel A, Daste F, Specht CG, Izeddin I, Vorkel D, Verbavatz JM, Darzacq X, Triller A, Pfenninger KH, Tareste D, Jackson CL#, Galli T#. Nat Cell Biol. 2014 May;16(5):434-44. doi: 10.1038/ncb2937. Epub 2014 Apr 6. PMID: 24705552  #co-corresponding authors

Targeting of the Arf-GEF GBF1 to lipid droplets and Golgi membranes.

Bouvet S, Golinelli-Cohen MP, Contremoulins V, Jackson CL.J Cell Sci. 2013 Oct 15;126(Pt 20):4794-805. doi: 10.1242/jcs.134254. Epub 2013 Aug 13.PMID: 23943872

Automated tracing of microtubules in electron tomograms of plastic embedded samples of Caenorhabditis elegans embryos.

Weber B, Greenan G, Prohaska S, Baum D, Hege HC, Müller-Reichert T, Hyman AA, Verbavatz JM.J Struct Biol. 2012 May;178(2):129-38. doi: 10.1016/j.jsb.2011.12.004. Epub 2011 Dec 13.PMID: 22182731

α-Synuclein and ALPS motifs are membrane curvature sensors whose contrasting chemistry mediates selective vesicle binding.

Pranke IM, Morello V, Bigay J, Gibson K, Verbavatz JM, Antonny B, Jackson CL. J Cell Biol. 2011 Jul 11;194(1):89-103. doi: 10.1083/jcb.201011118. PMID: 21746853

ARF family G proteins and their regulators: roles in membrane transport, development and disease.

Donaldson JG, Jackson CL.Nat Rev Mol Cell Biol. 2011 Jun;12(6):362-75. doi: 10.1038/nrm3117. Epub 2011 May 18.PMID: 21587297

2019

Jackson CL. Lipid droplet biogenesis. (2019) Curr Opin Cell Biol. 59:88-96. Review. doi: 10.1016/j.ceb.2019.03.018.

Franke C, Repnik U, Segeletz S, Brouilly N, Kalaidzidis Y, Verbavatz JM, Zerial M. (2019) Correlative single-molecule localization microscopy and electron tomography reveals endosome nanoscale domains. Traffic. 20:601-617. doi: 10.1111/tra.12671.

Lebsir N, Goueslain L, Farhat R, Callens N, Dubuisson J, Jackson CL#, Rouillé Y#. (2019 ) Functional and Physical Interaction between the Arf Activator GBF1 and Hepatitis C Virus NS3 Protein. J Virol. 93(6). pii: e01459-18. doi: 10.1128/JVI.01459-18. #co-corresponding authors

2018

Walch L, Pellier E, Leng W, Lakisic G, Gautreau A, Contremoulins V, Verbavatz JM#, Jackson CL#. (2018) GBF1 and Arf1 interact with Miro and regulate mitochondrial positioning within cells. Scientific Reports 8(1):17121. doi: 10.1038/s41598-018-35190-0.  #co-corresponding authors

Jackson CL. (2018) Activators and Effectors of the Small G Protein Arf1 in Regulation of Golgi Dynamics During the Cell Division Cycle. Front Cell Dev Biol. 6:29.  doi: 10.3389/fcell.2018.00029.

Magliozzi R, Carrero ZI, Low TY, Yuniati L, Valdes-Quezada C, Kruiswijk F, van Wijk K, Heck AJR, Jackson CL, Guardavaccaro D. (2018) Inheritance of the Golgi Apparatus and Cytokinesis Are Controlled by Degradation of GBF1. Cell Rep. 23(11):3381-3391.e4.  doi: 10.1016/j.celrep.2018.05.031.

Jackson CL. (2018) Membrane Trafficking: A Little Flexibility Helps Vesicles Get into Shape. Curr Biol. 28(12):R706-R709 Review. doi: 10.1016/j.cub.2018.04.068.

Copic A, Antoine-Bally S, Giménez-Andrés M, La Torre Garay C, Antonny B, Manni MM, Pagnotta S, Guihot J, Jackson CL. (2018) A giant amphipathic helix from a perilipin that is adapted for coating lipid droplets. Nat Commun. 9:1332. doi: 10.1038/s41467-018-03717-8.

Farhat R, Ankavay M, Lebsir N, Gouttenoire J, Jackson CL, Wychowski C, Moradpour D, Dubuisson J, Rouillé Y, Cocquerel L. (2018) Identification of GBF1 as a cellular factor required for hepatitis E virus RNA replication. Cell Microbiol. 20(1):1-11. doi: 10.1111/cmi.12572.

2017

Kaczmarek B, Verbavatz JM, Jackson CL. (2017) GBF1 and Arf1 function in vesicular trafficking, lipid homoeostasis and organelle dynamics. Biol Cell. 109(12):391-399 Review. doi: 10.1111/boc.201700042.

Bacle A, Gautier R, Jackson CL, Fuchs PF, Vanni S. (2017) Interdigitation between Triglyceridesand Lipids Modulates Surface Properties of Lipid Droplets. Biophys J. 112(7):1417-1430. doi: 10.1016/j.bpj.2017.02.032.

Laband K, Le Borgne R, Edwards F, Stefanutti M, Canman JC, Verbavatz JM, Dumont J. (2017) Chromosome segregation occurs by microtubule pushing in oocytes. Nat Commun. 8:1499. doi: 10.1038/s41467-017-01539-8.

Müller A, Neukam M, Ivanova A, Sönmez A, Münster C, Kretschmar S, Kalaidzidis  Y, Kurth T, Verbavatz JM, Solimena M. (2017) A Global Approach for Quantitative Super Resolution and Electron Microscopy on Cryo and Epoxy Sections Using Self-labeling Protein Tags. Sci Rep. 7:23. doi: 10.1038/s41598-017-00033-x.

2016

Jackson CL, Walch L, Verbavatz JM. (2016) Lipids and Their Trafficking: An Integral Part of Cellular Organization. Developmental Cell. 39(2):139-153 Review. doi: 10.1016/j.devcel.2016.09.030.

2015

Moser von Filseck J*, Čopič A*, Delfosse V*, Vanni S, Jackson CL, Bourguet W, Drin G. (2015) Phosphatidylserine transport by ORP/Osh proteins is driven by phosphatidylinositol 4-phosphate. Science 349(6246):432-6. *co-first authors  doi: 10.1126/science.aab1346

Malinovska L, Palm S, Gibson K, Verbavatz JM, Alberti S. 2015.
Dictyostelium discoideum has a highly Q/N-rich proteome and shows an unusual resilience to protein aggregation. Proc Natl Acad Sci U S A. 2015 112(20):E2620-9. doi: 10.1073/pnas.1504459112

Walch L, Čopič A, Jackson CL. (2015) Fatty acid metabolism meets organelle dynamics. Developmental Cell 32(6):657-8. doi: 10.1016/j.devcel.2015.03.008.

 

Kuster A, Nola S, Dingli F, Vacca B, Gauchy C, Beaujouan JC, Nunez M, Moncion T, Loew D, Formstecher E, Galli T, Proux-Gillardeaux V. (2015) The Q-soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor (Q-SNARE) SNAP-47 Regulates Trafficking of Selected Vesicle-associated Membrane Proteins (VAMPs). J Biol Chem. Nov 20;290(47):28056-28069. doi: 10.1074/jbc.M115.666362.

2020, Manuel Giménez Andrés, “Interaction of perilipin amphipathic helices with lipid droplets”, Université Paris-Saclay

2013, Samuel Bouvet, “Lipids and Trafficking: Roles of GBF1, exchange factor for the small G protein Arf1”, Université Paris Sud

2013, Aymen Jemaiel, “Study of vesicular and non-vesicular membrane trafficking in yeast “, Université Paris Sud

2011, Iwona M. Pranke “Specificity of membrane targeting by ALPS motifs and α-synuclein”, Université Paris Sud