bims-mecmid Biomed News
on Membrane communication in mitochondrial dynamics
Issue of 2022‒05‒15
eight papers selected by
Mauricio Cardenas Rodriguez
University of Padova
  1. Acute Manipulation of Outer Membrane Phospholipid Composition Directly Alters Mitochondrial Dynamics and Ultrastructure.
  2. The role of IR-induced swelling in OPA1-MICOS interaction in cardiac mitochondria.
  3. Crosstalk between Mitochondrial Protein Import and Lipids.
  4. Mitochondrial micropeptide STMP1 enhances mitochondrial fission to promote tumor metastasis.
  5. The Role of the Optic Atrophy 1 (OPA1) Protein in Drug-Induced Liver Injury.
  6. Mitochondrial Fission is Essential to Maintain Cristae Morphology and Bioenergetics.
  7. Characterisation of a novel OPA1 splice variant resulting in cryptic splice site activation and mitochondrial dysfunction.
  8. Lifespan Extension of Podospora anserina Mic60-Subcomplex Mutants Depends on Cardiolipin Remodeling.
  1. FASEB J. 2022 May;36 Suppl 1
    Acute Manipulation of Outer Membrane Phospholipid Composition Directly Alters Mitochondrial Dynamics and Ultrastructure.
    Joshua Pemberton, Yeun Ju Kim, Elizabeth Ferrer, Vijay Joshi, Tara Fischer, Marek Korzeniowski, Richard Youle, John Heuser, Tamas Balla.
      Mitochondria undergo coordinated rounds of fusion and fission that are critical for maintaining the functional integrity of this essential organelle. While a growing number of proteins have been identified as important regulators of mitochondrial dynamics, the direct role of membrane lipid composition during the fusion and fission processes is poorly understood. To address these shortcomings, we devised a protein-engineering platform that allows for the acute remodeling of structural phospholipids within the outer mitochondrial membrane (OMM) of intact cells. Specifically, we modified a bacterial phospholipase C (Bacillus cereus (Bc)PI-PLC) to initiate the rapid hydrolysis of phosphatidylinositol (PI) and locally generate diacylglycerol (DAG); an important intracellular signaling molecule and metabolic precursor that is used in diverse lipid biosynthetic pathways. Spatial restriction of enzyme activity was achieved using a chemically inducible system consisting of a rapamycin-dependent dimerization module (FKBP-BcPI-PLC) along with an OMM targeting sequence tagged with the FKBP-rapamycin binding domain (OMM-FRB). Using these unique molecular tools, we show that recruitment of FKBP-BcPI-PLC to the OMM not only causes the expected local accumulation of DAG, but also initiates the rapid and uniform fragmentation of the mitochondrial network. Mitochondrial fission induced by FKBP-BcPI-PLC is accompanied by profound swelling of the mitochondrial matrix along with vesiculation of the inner mitochondrial membrane (IMM) and a general loss of cristae, which all occur within minutes of tethering FKBP-BcPI-PLC to the OMM. Expression of dominant-negative constructs targeting essential GTPases known to regulate OMM fission suggest that both dynamin-related protein 1 (Drp1) and dynamin 2 (Dnm2) work together to drive efficient BcPI-PLC-induced mitochondrial division. However, results using a validated Drp1 knockout cell line show that the loss of Drp1 alone is sufficient to prevent the mitochondrial fragmentation initiated by FKBP-BcPI-PLC recruitment, indicating that Drp1 likely functions upstream or independent of Dnm2 in this context. Interestingly, unlike the induced OMM fission, removal of Drp1 from cells does not prevent the matrix swelling or OMM constrictions observed in response to acute generation of DAG within the OMM. Ongoing experiments are now focused on characterizing new methods to sequentially metabolize the DAG generated within the OMM as well as investigate how local lipid composition influences the binding and oligomerization of membrane-shaping proteins that may function in concert with Drp1 to regulate mitochondrial remodeling. Overall, these studies establish a direct relationship between lipid metabolism within the OMM and clinically relevant morphological changes that are known to manifest in mitochondrial-associated diseases.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R3682
  2. FASEB J. 2022 May;36 Suppl 1
    The role of IR-induced swelling in OPA1-MICOS interaction in cardiac mitochondria.
    Keishla M Rodríguez-Graciani, Xavier R Chapa-Dubocq, Ivana Chaves-Negrón, Sabzali Javadov.
      Mitochondria are membrane-bound organelles composed of two membranes, the inner mitochondrial membrane (IMM) and the outer mitochondrial membrane (OMM), with the intermembrane space (IMS) localized between them. The IMM is divided into two subcompartments, the inner boundary membrane (IBM) that lies parallel to the OMM, and the cristae membrane (CM), which are IMM invaginations folded into cristae. The IBM and CM are connected at terminals forming circular-like openings known as cristae junctions (CJs). The structural integrity of the mitochondrial CJs is maintained by the mitochondrial contact site and cristae organizing system (MICOS), a specific protein complex containing several structural and regulatory proteins. The MICOS proteins control the IMM architecture through direct membrane shaping, formation of contact sites, and biogenesis of proteins and lipids. In addition, the optic atrophy 1 (OPA1), a mitochondrial fusion protein, localized also in the CM, has been shown to participate in IMM remodeling which mediates the fusion of the IMM. The structural organization of the IMM is regulated by changes in the matrix volume of mitochondria; excessive matrix swelling in response to energetic and oxidative stress induced by pathological stimuli such as cardiac ischemia-reperfusion (IR) impairs the integrity of CJs and thereby, alters mitochondrial function. The main goal of this study is to investigate the effects of cardiac IR-induced swelling on OPA1 and MICOS proteins. Hearts were isolated from male Sprague Dawley rats and perfused with Krebs-Henseleit solution (KHS) using the Langendorff-mode technique at a constant flow rate (10-12 ml/min). The animals were randomly assigned to the following groups: i) perfusion (no ischemia) for 55 min (C-55 group), ii) ischemia (I group) for 25 min, iii) ischemia in the presence of sanglifehrin A (SfA), an inhibitor of the permeability transition pore (IS), iv) perfusion (no ischemia) for 95 min (C-95), v) ischemia for 25-min followed by 40-min reperfusion (IR group), and vi) IR in the presence of SfA (IRS). SfA (0.5 µM) was present 10 min before ischemia (IS group) and throughout the entire period of reperfusion (IRS group). LDH activity was determined in the coronary effluent as a marker of cell death. At the end of reperfusion, mitochondria were isolated from the hearts by differential centrifugation for analysis of oxygen consumption rates, mitochondrial swelling, and protein levels of long and short forms of OPA1, and MICOS proteins. Our results show that ischemia diminished post-ischemic recovery of the hearts as evidenced by impaired cardiac contractility, increased LDH activity in the coronary effluent, and reduced mitochondrial respiration. Ischemia alone and IR differently affected the expression of both, OPA1 and mitofilin (Mic-60, a core MICOS protein) as well as other MICOS components. In conclusion, our data suggest that OPA1 and MICOS proteins are affected differently by ischemia alone and IR.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.L8099
  3. Int J Mol Sci. 2022 May 09. pii: 5274. [Epub ahead of print]23(9):
    Crosstalk between Mitochondrial Protein Import and Lipids.
    Juliane J Hoffmann, Thomas Becker.
      Mitochondria import about 1000 precursor proteins from the cytosol. The translocase of the outer membrane (TOM complex) forms the major entry site for precursor proteins. Subsequently, membrane-bound protein translocases sort the precursor proteins into the outer and inner membrane, the intermembrane space, and the matrix. The phospholipid composition of mitochondrial membranes is critical for protein import. Structural and biochemical data revealed that phospholipids affect the stability and activity of mitochondrial protein translocases. Integration of proteins into the target membrane involves rearrangement of phospholipids and distortion of the lipid bilayer. Phospholipids are present in the interface between subunits of protein translocases and affect the dynamic coupling of partner proteins. Phospholipids are required for full activity of the respiratory chain to generate membrane potential, which in turn drives protein import across and into the inner membrane. Finally, outer membrane protein translocases are closely linked to organellar contact sites that mediate lipid trafficking. Altogether, intensive crosstalk between mitochondrial protein import and lipid biogenesis controls mitochondrial biogenesis.
    Keywords:  SAM complex; TOM complex; mitochondria; phospholipids; protein import
    DOI:  https://doi.org/10.3390/ijms23095274
  4. Cancer Res. 2022 May 11. pii: canres.3910.2021. [Epub ahead of print]
    Mitochondrial micropeptide STMP1 enhances mitochondrial fission to promote tumor metastasis.
    Chen Xie, Feng-Yi Wang, Ye Sang, Bin Chen, Jia-Hui Huang, Feng-Jun He, Hui Li, Ying Zhu, Xingguo Liu, Shi-Mei Zhuang, Jian-Hong Fang.
      Micropeptides are a recently discovered class of molecules that play vital roles in various cellular processes, including differentiation, proliferation, and apoptosis. Here, we sought to identify cancer-associated micropeptides and to uncover their mechanistic functions. A micropeptide named short trans-membrane protein 1 (STMP1) that localizes at the inner mitochondrial membrane was identified to be upregulated in various cancer types and associated with metastasis and recurrence of hepatocellular carcinoma. Both gain- and loss-of-function studies revealed that STMP1 increased dynamin-related protein 1 (DRP1) activation to promote mitochondrial fission and enhanced migration of tumor cells. STMP1 silencing inhibited in vivo tumor metastasis in xenograft mouse models. Overexpression of STMP1 led to redistribution of mitochondria to the leading edge of cells and enhanced lamellipodia formation. Treatment with a DRP1 inhibitor abrogated the promotive effect of STMP1 on mitochondrial fission, lamellipodia formation, and tumor cell migration in vitro and metastasis in vivo. Furthermore, STMP1 interacted with myosin heavy chain 9 (MYH9), the subunit of non-muscle myosin II, and silencing MYH9 abrogated STMP1-induced DRP1 activation, mitochondrial fission, and cell migration. Collectively, this study identifies STMP1 as a critical regulator of metastasis and a novel unit of the mitochondrial fission protein machinery, providing a potential therapeutic target for treating metastases.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-21-3910
  5. FASEB J. 2022 May;36 Suppl 1
    The Role of the Optic Atrophy 1 (OPA1) Protein in Drug-Induced Liver Injury.
    Hakjoo Lee, Hiromi Sesaki, Yisang Yoon.
      The liver is the metabolic hub, and is responsible for the myriad of processes including the nutrient homeostasis and detoxification. Mitochondria of liver are critical for these functions. The detoxification process in liver, when severe, often results in liver damage through causing oxidative stress. Mitochondria are the main source of ROS and are also vulnerable to oxidant damage. Therefore, mitochondrial dysfunction is one of the prominent causes for drug-induced liver injury. Mitochondrial fission and fusion, the main processes of mitochondrial dynamics, determine mitochondrial shape, and are important for functional maintenance of mitochondria. However, the role of mitochondrial dynamics in drug-induced liver injury is poorly understood. In the current study, we examined the role of the optic atrophy 1 (OPA1) protein in drug-induced liver injury. OPA1 is associated with mitochondrial inner membrane (IM) and mediates IM fusion. OPA1 also regulates cristate structure, and is required for proper electron transport and ATP production. To investigate the OPA1's role, we used liver-specific OPA1-knockout (OPA1-LKO) mice with acetaminophen (APAP) administration as a model for drug-induced liver injury. We generated OPA1-LKO mice by crossing OPA1 flox mice with mice carrying the Cre recombinase under the albumin promoter. Whereas whole body KO of OPA1 causes embryonic lethality, OPA1-LKO mice appeared healthy and showed normal growth and behavior. Although the OPA1 gene in the liver was disrupted, OPA1-LKO mice showed approximately 30% of OPA1 remaining in the liver, presumably due to less efficient albumin promoter-mediated Cre expression and from other cell types of the liver. Liver histology revealed that OPA1-KO livers have disorganized hepatic cords with enlarged hepatocytes. Despite a reduced OPA1 level, mitochondria in OPA1-KO liver show near intact cristae structure and respiration, suggesting that a low level of OPA1 would support mitochondrial function in liver. We then tested the effect of OPA1 LKO on liver function under APAP stress. In APAP overdose, excess APAP metabolite depletes GSH in hepatocytes, which causes mitochondrial oxidative stress, leading to mitochondrial permeability transition, mitochondrial dysfunction, ATP depletion, and ultimately necrotic cell death. Upon administration of excess APAP, we found that OPA1-LKO mice were more sensitive to APAP-induced liver injury compared with the control mice. Histological analyses showed significantly more expanded focal centrilobular necrosis with vacuolization, cell swelling, and nuclear disintegration in OPA1-KO livers. Alanine aminotransferase levels, as a clinical chemistry parameter, were higher in OPA1-LKO mice than in control mice with APAP overdose. Furthermore, phospho-JNK levels were higher in OPA1-KO livers, indicating increased initial oxidative stress. However, depletion of hepatic glutathione (GSH) contents and reduction of GSH/GSSG ratio upon APAP treatment were similar between the LKO and control liver. Together, our experimental results indicate that although liver is tolerant to a reduced level of OPA1, OPA1 depletion lowers the stress threshold and makes hepatocytes more sensitive to APAP-induced liver injury.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R3945
  6. FASEB J. 2022 May;36 Suppl 1
    Mitochondrial Fission is Essential to Maintain Cristae Morphology and Bioenergetics.
    Gabriella Robertson, Mira Patel, Stellan Riffle, Andrea Marshall, Heather Beasley, Edgar Garza-Lopez, Antentor O Hinton, Maria Stoll, Jason Mears, Vivian Gama.
      Mitochondria and peroxisomes are both dynamic signaling organelles that constantly undergo fission. While mitochondrial fission and fusion are known to coordinate cellular metabolism, proliferation, and apoptosis, the physiological relevance of peroxisome dynamics and the implications for cell fate are not fully understood. DRP1 (dynamin-related protein 1) is an essential GTPase that executes both mitochondrial and peroxisomal fission. Patients with de novo heterozygous missense mutations in the gene that encodes DRP1, DNM1L, present with encephalopathy due to mitochondrial and peroxisomal elongation (EMPF). EMPF is a devastating neurodevelopmental disease with no effective treatment. To interrogate the molecular mechanisms by which DRP1 mutations cause developmental defects, we are using patient-derived fibroblasts and iPSC-derived models from patients with mutations in different domains of DRP1 who present with clinically disparate conditions. Using super resolution imaging, we find that patient cells, in addition to displaying elongated mitochondrial and peroxisomal morphology, present with aberrant cristae structure. Given the direct link between cristae morphology and oxidative phosphorylation efficiency, we explored the impact of these mutations on cellular energy production. Patient cells display a lower coupling efficiency of the electron transport chain, increased proton leak, and Complex III deficiency. In addition to these metabolic abnormalities, mitochondrial hyperfusion results in hyperpolarized mitochondrial membrane potential. Intriguingly, human fibroblasts are capable of cellular reprogramming into iPSCs and appear to display peroxisome-mediated mitochondrial adaptations that could help sustain these cell fate transitions. Understanding the mechanism by which DRP1 mutations cause cellular dysfunction will give insight into the role of mitochondrial and peroxisome dynamics in neurodevelopment.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R3665
  7. Eur J Hum Genet. 2022 May 09.
    Characterisation of a novel OPA1 splice variant resulting in cryptic splice site activation and mitochondrial dysfunction.
    Joshua Paul Harvey, Patrick Yu-Wai-Man, Michael Edward Cheetham.
      Autosomal dominant optic atrophy (DOA) is an inherited optic neuropathy that results in progressive, bilateral visual acuity loss and field defects. OPA1 is the causative gene in around 60% of cases of DOA. The majority of patients have a pure ocular phenotype, but 20% have extra-ocular features (DOA +). We report on a patient with DOA + manifesting as bilateral optic atrophy, spastic paraparesis, urinary incontinence and white matter changes in the central nervous system associated with a novel heterozygous splice variant NM_015560.2(OPA1):c.2356-1 G > T. Further characterisation, which was performed using fibroblasts obtained from a skin biopsy, demonstrated that this variant altered mRNA splicing of the OPA1 transcript, specifically a 21 base pair deletion at the start of exon 24, NM_015560.2(OPA1):p.Cys786_Lys792del. The majority of variant transcripts were shown to escape nonsense-mediated decay and modelling of the predicted protein structure suggests that the in-frame 7 amino acid deletion may affect OPA1 oligomerisation. Fibroblasts carrying the c.2356-1 G > T variant demonstrated impaired mitochondrial bioenergetics, membrane potential, increased cell death, and disrupted and fragmented mitochondrial networks in comparison to WT cells. This study suggests that the c.2356-1 G > T OPA1 splice site variant leads to a cryptic splice site activation and may manifest in a dominant-negative manner, which could account for the patient's severe syndromic phenotype.
    DOI:  https://doi.org/10.1038/s41431-022-01102-0
  8. Int J Mol Sci. 2022 Apr 25. pii: 4741. [Epub ahead of print]23(9):
    Lifespan Extension of Podospora anserina Mic60-Subcomplex Mutants Depends on Cardiolipin Remodeling.
    Lisa-Marie Marschall, Verena Warnsmann, Anja C Meeßen, Timo Löser, Heinz D Osiewacz.
      Function of mitochondria largely depends on a characteristic ultrastructure with typical invaginations, namely the cristae of the inner mitochondrial membrane. The mitochondrial signature phospholipid cardiolipin (CL), the F1Fo-ATP-synthase, and the 'mitochondrial contact site and cristae organizing system' (MICOS) complex are involved in this process. Previous studies with Podospora anserina demonstrated that manipulation of MICOS leads to altered cristae structure and prolongs lifespan. While longevity of Mic10-subcomplex mutants is induced by mitohormesis, the underlying mechanism in the Mic60-subcomplex deletion mutants was unclear. Since several studies indicated a connection between MICOS and phospholipid composition, we now analyzed the impact of MICOS on mitochondrial phospholipid metabolism. Data from lipidomic analysis identified alterations in phospholipid profile and acyl composition of CL in Mic60-subcomplex mutants. These changes appear to have beneficial effects on membrane properties and promote longevity. Impairments of CL remodeling in a PaMIC60 ablated mutant lead to a complete abrogation of longevity. This effect is reversed by supplementation of the growth medium with linoleic acid, a fatty acid which allows the formation of tetra-octadecanoyl CL. In the PaMic60 deletion mutant, this CL species appears to lead to longevity. Overall, our data demonstrate a tight connection between MICOS, the regulation of mitochondrial phospholipid homeostasis, and aging of P. anserina.
    Keywords:  MICOS; Podospora anserina; aging; cardiolipin; cristae; mitochondria; tafazzin
    DOI:  https://doi.org/10.3390/ijms23094741
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