scholarly journals Miro clusters regulate ER-mitochondria contact sites and link cristae organization to the mitochondrial transport machinery

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Souvik Modi ◽  
Guillermo López-Doménech ◽  
Elise F. Halff ◽  
Christian Covill-Cooke ◽  
Davor Ivankovic ◽  
...  
Keyword(s):  
Mitochondrial Membrane ◽  
Super Resolution ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Localization ◽  
Mitochondrial Membranes ◽  
Mouse Embryonic Fibroblasts ◽  
Contact Site ◽  
Embryonic Fibroblasts ◽  
Contact Sites ◽  
Regulated Trafficking

Abstract Mitochondrial Rho (Miro) GTPases localize to the outer mitochondrial membrane and are essential machinery for the regulated trafficking of mitochondria to defined subcellular locations. However, their sub-mitochondrial localization and relationship with other critical mitochondrial complexes remains poorly understood. Here, using super-resolution fluorescence microscopy, we report that Miro proteins form nanometer-sized clusters along the mitochondrial outer membrane in association with the Mitochondrial Contact Site and Cristae Organizing System (MICOS). Using knockout mouse embryonic fibroblasts we show that Miro1 and Miro2 are required for normal mitochondrial cristae architecture and Endoplasmic Reticulum-Mitochondria Contacts Sites (ERMCS). Further, we show that Miro couples MICOS to TRAK motor protein adaptors to ensure the concerted transport of the two mitochondrial membranes and the correct distribution of cristae on the mitochondrial membrane. The Miro nanoscale organization, association with MICOS complex and regulation of ERMCS reveal new levels of control of the Miro GTPases on mitochondrial functionality.

scholarly journals Dual role of Miro protein clusters in mitochondrial cristae organisation and ER-Mitochondria Contact Sites

2019 ◽  
Author(s):  
Souvik Modi ◽  
Guillermo López-Doménech ◽  
Elise F. Halff ◽  
Christian Covill-Cooke ◽  
Davor Ivankovic ◽  
...  
Keyword(s):  
Mitochondrial Membrane ◽  
Super Resolution ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Localization ◽  
Mitochondrial Membranes ◽  
Mouse Embryonic Fibroblasts ◽  
Contact Site ◽  
Embryonic Fibroblasts ◽  
Regulated Trafficking

AbstractMitochondrial Rho (Miro) GTPases localize to the outer mitochondrial membrane and are essential machinery for the regulated trafficking of mitochondria to defined subcellular locations. However, their sub-mitochondrial localization and relationship with other critical mitochondrial complexes remains poorly understood. Here, using super-resolution fluorescence microscopy, we report that Miro proteins form nanometer-sized clusters along the mitochondrial outer membrane in association with the Mitochondrial Contact Site and Cristae Organizing System (MICOS). Using knockout mouse embryonic fibroblasts (MEF) we show that Miro1 and Miro2 are required for normal mitochondrial cristae architecture and endoplasmic reticulum-mitochondria contacts sites (ERMCS). Further, we show that Miro couples MICOS to TRAK motor protein adaptors to ensure the concerted transport of the two mitochondrial membranes and the correct distribution of cristae on the mitochondrial membrane. The Miro nanoscale organization, association with MICOS complex and regulation of ERMCS reveal new levels of control of the Miro GTPases on mitochondrial functionality.

scholarly journals Protease OMA1 Modulates Mitochondrial Bioenergetics and Ultrastructure through Dynamic Association with MICOS Complex

2020 ◽  
Author(s):  
Martonio Ponte Viana ◽  
Roman M. Levytskyy ◽  
Ruchika Anand ◽  
Andreas S. Reichert ◽  
Oleh Khalimonchuk
Keyword(s):  
Mitochondrial Membrane ◽  
Outer Mitochondrial Membrane ◽  
Mitochondrial Ultrastructure ◽  
Mitochondrial Membranes ◽  
Complex Dynamic ◽  
Contact Site ◽  
Outer Membranes ◽  
Mitochondrial Functions ◽  
Membrane Contacts ◽  
Dynamic Association

ABSTRACTRemodeling of mitochondrial ultrastructure is a complex dynamic process that is critical for a variety of mitochondrial functions and apoptosis. Although the key regulators of this process - mitochondrial contact site and cristae junction organizing system (MICOS) and GTPase Optic Atrophy 1 (OPA1) have been characterized, the mechanisms behind this regulation remain incompletely defined. Here, we found that in addition to its role in mitochondrial division, metallopeptidase OMA1 is required for maintenance of contacts between the inner and outer membranes through a dynamic association with MICOS. This association is independent of OPA1, appears to be mediated via the MICOS subunit MIC60, and is important for stability of MICOS machinery and the inner-outer mitochondrial membrane contacts. We find that OMA1-MICOS relay is required for stability of respiratory supercomplexes, optimal bioenergetic output in response to cellular insults, and apoptosis. Loss of OMA1 affects these activities; remarkably it can be partially compensated for by an artificial MICOS-emulating tether protein that bridges the inner and outer mitochondrial membranes. Our data show that OMA1-mediated support of mitochondrial ultrastructure is required for maintenance of mitochondrial architecture and bioenergetics under both basal and homeostasis-challenging conditions, and suggest a previously unrecognized role for this protease in mitochondrial physiology.

scholarly journals The distribution of anionic sites on the surfaces of mitochondrial membranes. Visual probing with polycationic ferritin.

1975 ◽  
Vol 65 (3) ◽  
pp. 615-630 ◽  
Author(s):  
C R Hackenbrock ◽  
K J Miller
Keyword(s):  
Outer Membrane ◽  
Mitochondrial Membrane ◽  
High Density ◽  
Intermembrane Space ◽  
Mitochondrial Membranes ◽  
Inner Mitochondrial Membrane ◽  
Anionic Sites ◽  
Contact Sites ◽  
Intact Mitochondrion ◽  
Boundary Membrane

Polycationic ferritin, a multivalent ligand, was used as a visual probe to determine the distribution and density of anionic sites on the surfaces of rat liver mitochondrial membranes. Both the distribution of bound polycationic ferritin and the topography of the outer surface of the inner mitochondrial membrane were studied in depth by utilizing thin sections and critical-point dried, whole mount preparations for transmission electron microscopy and by scanning electron microscopy. Based on its relative affinity for polycationic ferritin, the surface of the inner membrane contains discrete regions of high density and low density anionic sites. Whereas the surface of the cristal membrane contains a low density of anionic sites, the surface of the inner boundary membrane contains patches of high density anionic sites. The high density anionic sites on the inner boundary membrane were found to persist as stable patches and did not dissociate or randomize freely when the membrane was converted osmotically to a spherical configuration. The observations suggest that the inner mitochondrial membrane is composed of two major regions of anionic macromolecular distinction. It is well-known that an intermembrane space exists between the two membranes of the intact mitochondrion; however, a number of contact sites occur between the two membranes. We determined that the outer membrane, partially disrupted by treatment with digitonin, remains attached to the inner membrane at these contact sites as inverted vesicles. Such attached vesicles show that the inner surface of the outer membrane contains anionic sites, but of decreased density, surrounding the contact sites. Thus, the intermembrane space in the intact mitochondrion may be maintained by electronegative surfaces of the two mitochondrial membranes. The distribution of anionic sites on the outer surface of the outer membrane is random. The nature and function of fixed anionic surface charges and membrane contact sites are discussed with regard to recent reports relating to calcium transport, protein assembly into mitochondrial membranes, and membrane fluidity.

scholarly journals Mdv1p Is a Wd Repeat Protein That Interacts with the Dynamin-Related Gtpase, Dnm1p, to Trigger Mitochondrial Division

2000 ◽  
Vol 151 (2) ◽  
pp. 353-366 ◽  
Author(s):  
Quinton Tieu ◽  
Jodi Nunnari
Keyword(s):  
Outer Membrane ◽  
Mitochondrial Membrane ◽  
Mitochondrial Fission ◽  
Coiled Coil ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Targeting ◽  
Mitochondrial Membranes ◽  
Mitochondrial Division ◽  
Targeting Signal ◽  
Two Hybrid

Mitochondrial fission is mediated by the dynamin-related GTPase, Dnm1p, which assembles on the mitochondrial outer membrane into punctate structures associated with sites of membrane constriction and fission. We have identified additional nuclear genes required for mitochondrial fission, termed MDV (for mitochondrial division). MDV1 encodes a predicted soluble protein, containing a coiled-coil motif and seven COOH-terminal WD repeats. Genetic and two-hybrid analyses indicate that Mdv1p interacts with Dnm1p to mediate mitochondrial fission. In addition, Mdv1p colocalizes with Dnm1p in fission-mediating punctate structures on the mitochondrial outer membrane. Whereas localization of Mdv1p to these structures requires Dnm1p, localization of Mdv1p to mitochondrial membranes does not. This indicates that Mdv1p possesses a Dnm1p-independent mitochondrial targeting signal. Dnm1p-independent targeting of Mdv1p to mitochondria requires MDV2. Our data indicate that MDV2 also functions separately to regulate the assembly of Dnm1p into punctate structures. In contrast, Mdv1p is not required for the assembly of Dnm1p, but Dnm1p-containing punctate structures lacking Mdv1p are not able to complete division. Our studies suggest that mitochondrial fission is a multi-step process in which Mdv2p regulates the assembly of Dnm1p into punctate structures and together with Mdv1p functions later during fission to facilitate Dnm1p-dependent mitochondrial membrane constriction and/or division.

scholarly journals Loss of SHMT2 and Folate Deficiency Impair Energy Metabolism in Mouse Embryonic Fibroblasts Cells

2021 ◽  
Vol 5 (Supplement_2) ◽  
pp. 1311-1311
Author(s):  
Joanna Fiddler ◽  
Jamie Blum ◽  
Anna Thalacker-Mercer ◽  
Martha Field
Keyword(s):  
Enzyme Activity ◽  
Membrane Potential ◽  
Energy Metabolism ◽  
Pyruvate Kinase ◽  
Mitochondrial Membrane ◽  
Folate Deficiency ◽  
Oxidative Capacity ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Protein Levels

Abstract Objectives Serine hydroxymethyltransferase 2 (SHMT2) catalyzes the reversible conversion of tetrahydrofolate (THF) and serine producing THF-conjugated one-carbon units and glycine in the mitochondria. Recently biallelic SHMT2 variants were identified in humans and suggested to alter the protein's active site, potentially disrupting enzymatic function. Indeed, patient fibroblasts exhibited impaired oxidative capacity, however, with no changes in oxidative phosphorylation protein levels. Furthermore, immortalized cells models of total SHMT2 loss or folate deficiency exhibit decreased oxidative capacity and impaired mitochondrial complex I assembly and protein levels suggesting folate-mediated one-carbon metabolism (FOCM) and the oxidative phosphorylation system are functionally coordinated. We have developed a mouse model of reduced Shmt2 expression that provides an appropriate reference model to further study the deleterious human SHMT2 variants and their connection to folate metabolism. Methods Primary mouse embryonic fibroblasts (MEF) were isolated from a C57Bl/6 dam crossed with a heterozygous Shmt2+/− (HET) male. MEF embryos were genotyped, and Shmt2+/+ (wild type) or Shmt2+/− MEF cells were cultured for 4 doubling in 2 nM (deficient) or 25 nM folate (sufficient) medium. Cells were examined for proliferation, total folate levels, mtDNA content, protein levels of pyruvate kinase and PGC1α, pyruvate kinase enzyme activity, and mitochondrial membrane potential and function. Results Loss of Shmt2 and folate deficiency significantly reduced cell proliferation and oxygen consumption rate. Total cellular folate levels and pyruvate kinase enzyme activity and protein levels were reduced with folate deficiency. While there were no changes in mitochondrial DNA content or complex 1 proteins NDUFA9 or ND3, mitochondrial membrane potential was impaired and PGC1α protein levels were elevated in Shmt2+/− MEF cells. Conclusions Our results demonstrate that loss of SHMT2 impairs cellular proliferation and that SHMT2 expression and folate status impact cellular energy metabolism and mitochondrial health. Funding Sources This study was financially supported by a President's Council of Cornell Women Award and Cornell University Division of Nutritional Sciences funds.

Structure of contact sites between the outer and inner mitochondrial membranes investigated by HVEM tomography

1996 ◽  
Vol 54 ◽  
pp. 966-967
Author(s):  
C.A. Mannella ◽  
K.F. Buttle ◽  
K.A. O‘Farrell ◽  
A. Leith ◽  
M. Marko
Keyword(s):  
Liver Mitochondrion ◽  
Adenine Nucleotide ◽  
Protein Complexes ◽  
Mitochondrial Membranes ◽  
Electron Microscopic ◽  
Contact Sites ◽  
Transmission Electron ◽  
Precursor Proteins ◽  
Membrane Contacts ◽  
And Function

Early transmission electron microscopy of plastic-embedded, thin-sectioned mitochondria indicated that there are numerous junctions between the outer and inner membranes of this organelle. More recent studies have suggested that the mitochondrial membrane contacts may be the site of protein complexes engaged in specialized functions, e.g., import of mitochondrial precursor proteins, adenine nucleotide channeling, and even intermembrane signalling. It has been suggested that the intermembrane contacts may be sites of membrane fusion involving non-bilayer lipid domains in the two membranes. However, despite growing interest in the nature and function of intramitochondrial contact sites, little is known about their structure.We are using electron microscopic tomography with the Albany HVEM to determine the internal organization of mitochondria. We have reconstructed a 0.6-μm section through an isolated, plasticembedded rat-liver mitochondrion by combining 123 projections collected by tilting (+/- 70°) around two perpendicular tilt axes. The resulting 3-D image has confirmed the basic inner-membrane organization inferred from lower-resolution reconstructions obtained from single-axis tomography.

scholarly journals Regulation of c-Fos Gene Expression by NF-κB: A p65 Homodimer Binding Site in Mouse Embryonic Fibroblasts but Not Human HEK293 Cells

PLoS ONE ◽  
2013 ◽  
Vol 8 (12) ◽  
pp. e84062 ◽  
Author(s):  
Yu-Cheng Tu ◽  
Duen-Yi Huang ◽  
Shine-Gwo Shiah ◽  
Jang-Shiun Wang ◽  
Wan-Wan Lin
Keyword(s):  
Gene Expression ◽  
Binding Site ◽  
Hek293 Cells ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts

scholarly journals Introduction of Mouse Embryonic Fibroblasts into Early Embryos: From Confusion to Constructive Discussion. Comment on Savatier, P. Introduction of Mouse Embryonic Fibroblasts into Early Embryos Causes Reprogramming and (Con)fusion. Cells 2021, 10, 772

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1534
Author(s):  
Krystyna Żyżyńska-Galeńska ◽  
Jolanta Karasiewicz ◽  
Agnieszka Bernat
Keyword(s):  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Early Embryos ◽  
Constructive Discussion

We would like to address the issues raised by Pierre Savatier in “Introduction of Mouse Embryonic Fibroblasts into Early Embryos Causes Reprogramming and (Con)Fusion” [...]

Sign in / Sign up

Export Citation Format

Share Document