bims-mecosi Biomed News
on Membrane contact sites
Issue of 2021–09–19
eleven papers selected by
Verena Kohler, Stockholm University



  1. J Neurosci Res. 2021 Sep 12.
      Mitochondria-associated ER membranes (MAMs) are formed by close and specific components in the contact sites between the endoplasmic reticulum (ER) and mitochondria, which participate in several cell functions, including lipid metabolism, autophagy, and Ca2+ signaling. Particularly, the presence of α-synuclein (α-syn) in MAMs was previously demonstrated, indicating a physical interaction among some proteins in this region and a potential involvement in cell dysfunctions. MAMs alterations are associated with neurodegenerative diseases such as Parkinson's disease (PD) and contribute to the pathogenesis features. Here, we investigated the effects of α-syn on MAMs and Ca2+ transfer from the ER to mitochondria in WT- and A30P α-syn-overexpressing SH-SY5Y or HEK293 cells. We observed that α-syn potentiates the mitochondrial membrane potential (Δψm ) loss induced by rotenone, increases mitophagy and mitochondrial Ca2+ overload. Additionally, in α-syn-overexpressing cells, we found a reduction in ER-mitochondria contact sites through the impairment of the GRP75-IP3R interaction, however, with no alteration in VDAC1-GRP75 interaction. Consequently, after Ca2+ release from the ER, α-syn-overexpressing cells demonstrated a reduction in Ca2+ buffering by mitochondria, suggesting a deregulation in MAM activity. Taken together, our data highlight the importance of the α-syn/MAMs/Ca2+ axis that potentially affects cell functions in PD.
    Keywords:  MAMs; Parkinson's disease; RRID:AB_10200697; RRID:AB_1078991; RRID:AB_11179069; RRID:AB_1118910; RRID:AB_1279301; RRID:AB_141607; RRID:AB_141637; RRID:AB_143165; RRID:AB_2085424; RRID:AB_2120468; RRID:AB_216026; RRID:AB_228341; RRID:AB_2536527; RRID:AB_2637028; RRID:AB_2714190; RRID:AB_2750920; RRID:AB_398108; RRID:AB_477593; RRID:AB_570711; RRID:AB_915950; RRID:Addgene_73209; RRID:CVCL_0019; RRID:CVCL_0045; RRID:SCR_002798; RRID:SCR_013672; calcium; mitochondria; α-synuclein
    DOI:  https://doi.org/10.1002/jnr.24952
  2. J Cell Sci. 2021 Sep 15. pii: jcs.255299. [Epub ahead of print]
      The Voltage Dependent Anion channel (VDAC) is a ubiquitous channel in the outer membrane of the mitochondrion with multiple roles in protein, metabolite and small molecule transport. In mammalian cells, VDAC, as part of a larger complex including the inositol triphosphate receptor, has been shown to have a role in mediating contacts between the mitochondria and endoplasmic reticulum (ER). We identify VDAC of the pathogenic apicomplexan Toxoplasma gondii and demonstrate its importance for parasite growth. We show that VDAC is involved in protein import and metabolite transfer to mitochondria. Further, depletion of VDAC resulted in significant morphological changes of the mitochondrion and ER, suggesting a role in mediating contacts between these organelles in T. gondii.
    Keywords:  Calcium; ER; Membrane contact sites; Mitochondria; Mitochondrion; Motility; Porin; Toxoplasma; VDAC
    DOI:  https://doi.org/10.1242/jcs.255299
  3. Plant Cell. 2021 Sep 17. pii: koab229. [Epub ahead of print]
      COPII, a coat protein complex that forms vesicles on the endoplasmic reticulum (ER), mediates trafficking to the Golgi. While metazoans have few genes encoding each COPII component, plants have expanded these gene families, leading to the hypothesis that plant COPII has functionally diversified. In the moss Physcomitrium (Physcomitrella) patens, the Sec23/24 gene families are each composed of seven genes. Silencing Sec23/24 revealed isoform-specific contributions to polarized growth, with the closely related Sec23D/E and Sec24C/D essential for protonemal development. Focusing on Sec23, we discovered that Sec23D/E mediate ER-to Golgi transport and are essential for tip growth, with Sec23D localizing to presumptive ER exit sites. By contrast, Sec23A,B,C,F, and G are dispensable and do not quantitatively affect ER-to-Golgi trafficking. However, Δsec23abcfg plants exhibited reduced secretion of plasma membrane cargo. Of the four highly expressed protonemal Sec23 genes, Sec23F/G are members of a divergent Sec23 clade specifically retained in land plants. Notably, Sec23G accumulates on ER-associated foci that are significantly larger, do not overlap with, and are independent of Sec23D. While Sec23D/E form ER exit sites and function as bona fide COPII components essential for tip-growing protonemata, Sec23G and the closely related Sec23F have likely functionally diversified, forming separate and independent ER exit sites and participating in Golgi-independent trafficking pathways.
    DOI:  https://doi.org/10.1093/plcell/koab229
  4. Dev Cell. 2021 Sep 07. pii: S1534-5807(21)00674-2. [Epub ahead of print]
      Membrane contact between intracellular organelles is important in mediating organelle communication. However, the assembly of molecular machinery at membrane contact site and its internal organization correlating with its functional activity remain unclear. Here, we demonstrate that a gel-like condensation of Cidec, a crucial protein for obesity development by facilitating lipid droplet (LD) fusion, occurs at the LD-LD contact site (LDCS) through phase separation. The homomeric interaction between the multivalent N terminus of Cidec is sufficient to promote its phase separation both in vivo and in vitro. Interestingly, Cidec condensation at LDCSs generates highly plastic and lipid-permeable fusion plates that are geometrically constrained by donor LDs. In addition, Cidec condensates are distributed unevenly in the fusion plate generating stochastic sub-compartments that may represent unique lipid passageways during LD fusion. We have thus uncovered the organization and functional significance of geometry-constrained Cidec phase separation in mediating LD fusion and lipid homeostasis.
    Keywords:  gel-like phase separation; lipid droplet fusion and growth; lipid droplet-lipid droplet contact site; lipid-permeable fusion plate
    DOI:  https://doi.org/10.1016/j.devcel.2021.08.015
  5. Cell Rep. 2021 Sep 14. pii: S2211-1247(21)01154-2. [Epub ahead of print]36(11): 109707
      Secretory cargos are collected at endoplasmic reticulum (ER) exit sites (ERES) before transport to the Golgi apparatus. Decades of research have provided many details of the molecular events underlying ER-Golgi exchanges. Essential questions, however, remain about the organization of the ER-Golgi interface in cells and the type of membrane structures mediating traffic from ERES. To investigate these, we use transgenic tagging in Drosophila flies, 3D-structured illumination microscopy (SIM), and focused ion beam scanning electron microscopy (FIB-SEM) to characterize ERES-Golgi units in collagen-producing fat body, imaginal discs, and imaginal discs overexpressing ERES determinant Tango1. Facing ERES, we find a pre-cis-Golgi region, equivalent to the vertebrate ER-Golgi intermediate compartment (ERGIC), involved in both anterograde and retrograde transport. This pre-cis-Golgi is continuous with the rest of the Golgi, not a separate compartment or collection of large carriers, for which we find no evidence. We observe, however, many vesicles, as well as pearled tubules connecting ERES and Golgi.
    Keywords:  ERGIC; Golgi; Tango1; intermediate compartment; secretion; traffic
    DOI:  https://doi.org/10.1016/j.celrep.2021.109707
  6. Theranostics. 2021 ;11(18): 8855-8873
      Mitochondrial dysfunction and oxidative stress are frequently observed in the early stages of Alzheimer's disease (AD). Studies have shown that presenilin-1 (PS1), the catalytic subunit of γ-secretase whose mutation is linked to familial AD (FAD), localizes to the mitochondrial membrane and regulates its homeostasis. Thus, we investigated how five PS1 mutations (A431E, E280A, H163R, M146V, and Δexon9) observed in FAD affect mitochondrial functions. Methods: We used H4 glioblastoma cell lines genetically engineered to inducibly express either the wild-type PS1 or one of the five PS1 mutants in order to examine mitochondrial morphology, dynamics, membrane potential, ATP production, mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs), oxidative stress, and bioenergetics. Furthermore, we used brains of PS1M146V knock-in mice, 3xTg-AD mice, and human AD patients in order to investigate the role of PS1 in regulating MAMs formation. Results: Each PS1 mutant exhibited slightly different mitochondrial dysfunction. Δexon9 mutant induced mitochondrial fragmentation while A431E, E280A, H163R, and M146V mutants increased MAMs formation. A431E, E280A, M146V, and Δexon9 mutants also induced mitochondrial ROS production. A431E mutant impaired both complex I and peroxidase activity while M146V mutant only impaired peroxidase activity. All PS1 mutants compromised mitochondrial membrane potential and cellular ATP levels were reduced by A431E, M146V, and Δexon9 mutants. Through comparative profiling of hippocampal gene expression in PS1M146V knock-in mice, we found that PS1M146V upregulates Atlastin 2 (ATL2) expression level, which increases ER-mitochondria contacts. Down-regulation of ATL2 after PS1 mutant induction rescued abnormally elevated ER-mitochondria interactions back to the normal level. Moreover, ATL2 expression levels were significantly elevated in the brains of 3xTg-AD mice and AD patients. Conclusions: Overall, our findings suggest that each of the five FAD-linked PS1 mutations has a deleterious effect on mitochondrial functions in a variety of ways. The adverse effects of PS1 mutations on mitochondria may contribute to MAMs formation and oxidative stress resulting in an accelerated age of disease onset in people harboring mutant PS1.
    Keywords:  ATL2; Alzheimer's disease; MAMs; Mitochondria; Presenilin-1
    DOI:  https://doi.org/10.7150/thno.59776
  7. Protein Sci. 2021 Sep 13.
      Phosphatidylserine (PS) synthase 1 (PSS1) of mammalian cells is a multiple membrane-spanning protein of the endoplasmic reticulum (ER) and regulated by inhibition with the product PS. Alanine-scanning mutagenesis of PSS1 has revealed eight amino acid residues as those crucial for its activity and six as those important for its regulation. Furthermore, three missense mutations in the human PSS1 gene, which lead to regulatory dysfunctions of PSS1 and are causative of Lenz-Majewski syndrome, have been identified. In this study, we investigated the membrane topology of PSS1 by means of epitope insertion and immunofluorescence. According to a 10-transmembrane segment model supported by topology analysis of PSS1, all the eight amino acid residues crucial for the enzyme activity were localized to the luminal side of the lipid bilayer or the lumen of the ER, whereas all the nine amino acid residues involved in the enzyme regulation were localized to the cytosol or the cytoplasmic side of the lipid bilayer of the ER. This localization of the functional amino acid residues suggests that PSS1 is regulated by inhibition with PS in the cytoplasmic leaflet of the ER membrane and synthesizes PS at the luminal leaflet. This article is protected by copyright. All rights reserved.
    Keywords:  Biosynthesis; Endoplasmic reticulum; Membrane topology; Metabolic regulation; Phosphatidylserine; Phosphatidylserine synthase; Phospholipids
    DOI:  https://doi.org/10.1002/pro.4182
  8. Cell Calcium. 2021 Sep 05. pii: S0143-4160(21)00122-6. [Epub ahead of print]99 102468
      The gamma secretase catalytic subunit presenilin 1 (PS1) is expressed in the endoplasmic reticulum (ER) of neurons, where it regulates Ca2+ signaling. PS1 is also expressed in heart, but its role in regulation of cardiac Ca2+ transport remains unknown. Since the type 2 sarco/endoplasmic reticulum Ca2+ ATPase (SERCA2a) plays a central role in cardiac Ca2+ homeostasis, we studied whether PS1 regulates the cardiac SERCA2a function. The experiments were conducted in an inducible human SERCA2a stable T-Rex-293 cell line transfected with fluorescently labeled PS1 and the ER Ca2+ sensor R-CEPIA1er. Confocal imaging showed that that PS1 is localized predominantly in the ER membrane. Fluorescent resonance energy transfer (FRET) experiments in HEK293 cells transfected with fluorescently labeled SERCA2a and PS1 revealed that the two proteins directly interact with a 1:1 stoichiometry. The functional significance of this interaction was investigated in a heterologous cellular environment using a novel approach to directly measure ER Ca2+ dynamics. Measurements of SERCA2a-mediated Ca2+ transport showed that PS1 enhanced Ca2+ uptake at low ER Ca2+ loads (<0.15 mM) and reduced uptake at high loads (>0.35 mM). The results of this study revealed that PS1 could act as an important regulator of the cardiac Ca2+ pump function with a complex stimulatory/inhibitory profile.
    Keywords:  Ca(2+) pump; Ca(2+) transport; Endoplasmic reticulum; FRET; HEK293 cells; Presenelin 1
    DOI:  https://doi.org/10.1016/j.ceca.2021.102468
  9. Redox Biol. 2021 Sep 10. pii: S2213-2317(21)00284-6. [Epub ahead of print]46 102125
      Heme is an essential cofactor required for a plethora of cellular processes in eukaryotes. In metazoans the heme biosynthetic pathway is typically partitioned between the cytosol and mitochondria, with the first and final steps taking place in the mitochondrion. The pathway has been extensively studied and its biosynthetic enzymes structurally characterized to varying extents. Nevertheless, understanding of the regulation of heme synthesis and factors that influence this process in metazoans remains incomplete. Therefore, we investigated the molecular organization as well as the physical and genetic interactions of the terminal pathway enzyme, ferrochelatase (Hem15), in the yeast Saccharomyces cerevisiae. Biochemical and genetic analyses revealed dynamic association of Hem15 with Mic60, a core component of the mitochondrial contact site and cristae organizing system (MICOS). Loss of MICOS negatively impacts Hem15 activity, affects the size of the Hem15 high-mass complex, and results in accumulation of reactive and potentially toxic tetrapyrrole precursors that may cause oxidative damage. Restoring intermembrane connectivity in MICOS-deficient cells mitigates these cytotoxic effects. These data provide new insights into how heme biosynthetic machinery is organized and regulated, linking mitochondrial architecture-organizing factors to heme homeostasis.
    Keywords:  Ferrochelatase; Heme; MICOS; Mitochondria; Yeast
    DOI:  https://doi.org/10.1016/j.redox.2021.102125
  10. Front Immunol. 2021 ;12 751865
      
    Keywords:  MSC; extracellular vesicles (EV); glycolytic activity; immunomodulation; macrophages; metabolism; miR (microRNA); mitochondria transfer
    DOI:  https://doi.org/10.3389/fimmu.2021.751865
  11. Nat Commun. 2021 Sep 14. 12(1): 5434
      Vesicle tethers are thought to underpin the efficiency of intracellular fusion by bridging vesicles to their target membranes. However, the interplay between tethering and fusion has remained enigmatic. Here, through optogenetic control of either a natural tether-the exocyst complex-or an artificial tether, we report that tethering regulates the mode of fusion. We find that vesicles mainly undergo kiss-and-run instead of full fusion in the absence of functional exocyst. Full fusion is rescued by optogenetically restoring exocyst function, in a manner likely dependent on the stoichiometry of tether engagement with the plasma membrane. In contrast, a passive artificial tether produces mostly kissing events, suggesting that kiss-and-run is the default mode of vesicle fusion. Optogenetic control of tethering further shows that fusion mode has physiological relevance since only full fusion could trigger lamellipodial expansion. These findings demonstrate that active coupling between tethering and fusion is critical for robust membrane merger.
    DOI:  https://doi.org/10.1038/s41467-021-25465-y