bims-mitdyn 2022-09-25 papers

bims-mitdyn

Biomed News

on Mitochondrial dynamics: mechanisms

Issue of 2022–09–25
eight papers selected by
Edmond Chan, Queen’s University, School of Medicine

A Mitofusin 2 - Hif1α axis sets a maturation checkpoint in regenerating skeletal muscle.
Flow pattern-dependent mitochondrial dynamics regulates the metabolic profile and inflammatory state of endothelial cells.
Oxygen regulation of breathing is abolished in mitochondrial complex III-deficient arterial chemoreceptors.
ORP5/8 and MIB/MICOS link ER-mitochondria and intra-mitochondrial contacts for non-vesicular transport of phosphatidylserine.
Elucidation of the interaction proteome of mitochondrial chaperone Hsp78 highlights its role in protein aggregation during heat stress.
Primary and metastatic tumors exhibit systems-level differences in dependence on mitochondrial respiratory function.
Oocyte mitophagy is critical for extended reproductive longevity.
Protocol to detect in vitro and in cell ubiquitylation of mitochondrial DNA polymerase gamma by mitochondrial E3 ligase MITOL.

J Clin Invest. 2022 Sep 20. pii: e161638. [Epub ahead of print]

A Mitofusin 2 - Hif1α axis sets a maturation checkpoint in regenerating skeletal muscle.

Xun Wang, Yuemeng Jia, Jiawei Zhao, Nicholas P Lesner, Cameron J Menezes, Spencer D Shelton, Siva Sai Krishna Venigalla, Jian Xu, Chunyu Cai, Prashant Mishra.

  A fundamental issue in regenerative medicine is whether there exist endogenous regulatory mechanisms that limit the speed and efficiency of the repair process. We report the existence of a maturation checkpoint during muscle regeneration which pauses myofibers at a neonatal stage. This checkpoint is regulated by the mitochondrial protein mitofusin 2 (Mfn2), whose expression is activated in response to muscle injury. Mfn2 is required for growth and maturation of regenerating myofibers; in the absence of Mfn2, new myofibers arrested at a neonatal stage, characterized by centrally nucleated myofibers and loss of H3K27me3 repressive marks at the neonatal myosin heavy chain gene. A similar arrest at the neonatal stage was observed in infantile cases of human centronuclear myopathy. Mechanistically, Mfn2 upregulation suppressed expression of Hypoxia-induced Factor 1α (Hif1α), which is induced in the setting of muscle damage. Sustained Hif1α signaling blocked maturation of new myofibers at the neonatal-to-adult fate transition, revealing the existence of a checkpoint that delays muscle regeneration. Correspondingly, inhibition of Hif1α allowed myofibers to bypass the checkpoint, thereby accelerating the repair process. We conclude that skeletal muscle contains a regenerative checkpoint which regulates the speed of myofiber maturation in response to Mitofusin 2 and Hif1α activity.

Keywords:  Epigenetics; Mitochondria; Muscle Biology; Stem cells

DOI:  https://doi.org/10.1172/JCI161638
JCI Insight. 2022 Sep 22. pii: e159286. [Epub ahead of print]7(18):

Flow pattern-dependent mitochondrial dynamics regulates the metabolic profile and inflammatory state of endothelial cells.

Soon-Gook Hong, Junchul Shin, Soo Young Choi, Jeffery C Powers, Benjamin M Meister, Jacqueline Sayoc, Jun Seok Son, Ryan Tierney, Fabio A Recchia, Michael D Brown, Xiaofeng Yang, Joon-Young Park.

  Endothelial mitochondria play a pivotal role in maintaining endothelial cell (EC) homeostasis through constantly altering their size, shape, and intracellular localization. Studies show that the disruption of the basal mitochondrial network in EC, forming excess fragmented mitochondria, implicates cardiovascular disease. However, cellular consequences underlying the morphological changes in the endothelial mitochondria under distinctively different, but physiologically occurring, flow patterns (i.e., unidirectional flow [UF] versus disturbed flow [DF]) are largely unknown. The purpose of this study was to investigate the effect of different flow patterns on mitochondrial morphology and its implications in EC phenotypes. We show that mitochondrial fragmentation is increased at DF-exposed vessel regions, where elongated mitochondria are predominant in the endothelium of UF-exposed regions. DF increased dynamin-related protein 1 (Drp1), mitochondrial reactive oxygen species (mtROS), hypoxia-inducible factor 1, glycolysis, and EC activation. Inhibition of Drp1 significantly attenuated these phenotypes. Carotid artery ligation and microfluidics experiments further validate that the significant induction of mitochondrial fragmentation was associated with EC activation in a Drp1-dependent manner. Contrarily, UF in vitro or voluntary exercise in vivo significantly decreased mitochondrial fragmentation and enhanced fatty acid uptake and OXPHOS. Our data suggest that flow patterns profoundly change mitochondrial fusion/fission events, and this change contributes to the determination of proinflammatory and metabolic states of ECs.

Keywords:  Atherosclerosis; Endothelial cells; Mitochondria; Vascular Biology

DOI:  https://doi.org/10.1172/jci.insight.159286
Proc Natl Acad Sci U S A. 2022 Sep 27. 119(39): e2202178119

Oxygen regulation of breathing is abolished in mitochondrial complex III-deficient arterial chemoreceptors.

Daniel Cabello-Rivera, Patricia Ortega-Sáenz, Lin Gao, Ana M Muñoz-Cabello, Victoria Bonilla-Henao, Paul T Schumacker, José López-Barneo.

  Acute oxygen (O2) sensing is essential for adaptation of organisms to hypoxic environments or medical conditions with restricted exchange of gases in the lung. The main acute O2-sensing organ is the carotid body (CB), which contains neurosecretory chemoreceptor (glomus) cells innervated by sensory fibers whose activation by hypoxia elicits hyperventilation and increased cardiac output. Glomus cells have mitochondria with specialized metabolic and electron transport chain (ETC) properties. Reduced mitochondrial complex (MC) IV activity by hypoxia leads to production of signaling molecules (NADH and reactive O2 species) in MCI and MCIII that modulate membrane ion channel activity. We studied mice with conditional genetic ablation of MCIII that disrupts the ETC in the CB and other catecholaminergic tissues. Glomus cells survived MCIII dysfunction but showed selective abolition of responsiveness to hypoxia (increased [Ca2+] and transmitter release) with normal responses to other stimuli. Mitochondrial hypoxic NADH and reactive O2 species signals were also suppressed. MCIII-deficient mice exhibited strong inhibition of the hypoxic ventilatory response and altered acclimatization to sustained hypoxia. These data indicate that a functional ETC, with coupling between MCI and MCIV, is required for acute O2 sensing. O2 regulation of breathing results from the integrated action of mitochondrial ETC complexes in arterial chemoreceptors.

Keywords:  acute O2 sensing; carotid body glomus cell; hypoxia; mitochondrial O2 sensing and signaling; mitochondrial complex III

DOI:  https://doi.org/10.1073/pnas.2202178119
Cell Rep. 2022 Sep 20. pii: S2211-1247(22)01196-2. [Epub ahead of print]40(12): 111364

ORP5/8 and MIB/MICOS link ER-mitochondria and intra-mitochondrial contacts for non-vesicular transport of phosphatidylserine.

Vera F Monteiro-Cardoso, Leila Rochin, Amita Arora, Audrey Houcine, Eeva Jääskeläinen, Annukka M Kivelä, Cécile Sauvanet, Romain Le Bars, Eyra Marien, Jonas Dehairs, Julie Neveu, Naima El Khallouki, Elena Santonico, Johannes V Swinnen, David Tareste, Vesa M Olkkonen, Francesca Giordano.

  Mitochondria are dynamic organelles essential for cell survival whose structural and functional integrity rely on selective and regulated transport of lipids from/to the endoplasmic reticulum (ER) and across the mitochondrial intermembrane space. As they are not connected by vesicular transport, the exchange of lipids between ER and mitochondria occurs at membrane contact sites. However, the mechanisms and proteins involved in these processes are only beginning to emerge. Here, we show that the main physiological localization of the lipid transfer proteins ORP5 and ORP8 is at mitochondria-associated ER membrane (MAM) subdomains, physically linked to the mitochondrial intermembrane space bridging (MIB)/mitochondrial contact sites and cristae junction organizing system (MICOS) complexes that bridge the two mitochondrial membranes. We also show that ORP5/ORP8 mediate non-vesicular transport of phosphatidylserine (PS) lipids from the ER to mitochondria by cooperating with the MIB/MICOS complexes. Overall our study reveals a physical and functional link between ER-mitochondria contacts involved in lipid transfer and intra-mitochondrial membrane contacts maintained by the MIB/MICOS complexes.

Keywords:  CP: Cell biology; MAM; MICOS; Mic60; ORP; SAM50; cristae junctions; membrane contact sites; mitochondria; phosphatidylserine

DOI:  https://doi.org/10.1016/j.celrep.2022.111364
J Biol Chem. 2022 Sep 14. pii: S0021-9258(22)00937-1. [Epub ahead of print] 102494

Elucidation of the interaction proteome of mitochondrial chaperone Hsp78 highlights its role in protein aggregation during heat stress.

Witold Jaworek, Marc Sylvester, Giovanna Cenini, Wolfgang Voos.

  Chaperones of the Hsp100/Clp family represent major components of protein homeostasis, conferring maintenance of protein activity under stress. The ClpB-type members of the family, present in bacteria, fungi, and plants, are able to resolubilize aggregated proteins. The mitochondrial member of the ClpB family in Saccharomyces cerevisiae is Hsp78. Although Hsp78 has been shown to contribute to proteostasis in elevated temperatures, the biochemical mechanisms underlying this mitochondria-specific thermotolerance are still largely unclear. To identify endogenous chaperone substrate proteins, here we generated an Hsp78-ATPase mutant with stabilized substrate binding behavior. We used two stable isotope labeling (SILAC)-based quantitative mass spectrometry approaches to analyze the role of Hsp78 during heat stress-induced mitochondrial protein aggregation and disaggregation on a proteomic level. We first identified the endogenous substrate spectrum of the Hsp78 chaperone, comprising a wide variety of proteins related to metabolic functions including energy production and protein synthesis, as well as other chaperones, indicating its crucial functions in mitochondrial stress resistance. We then compared these interaction data with aggregation and disaggregation processes in mitochondria under heat stress, which revealed specific aggregation-prone protein populations and demonstrated the direct quantitative impact of Hsp78 on stress-dependent protein solubility under different conditions. We conclude that Hsp78, together with its cofactors, represents a recovery system that protects major mitochondrial metabolic functions during heat stress as well as restores protein biogenesis capacity after the return to normal conditions.

Keywords:  Hsp78; cell biology; chaperone; heat stress; mitochondria; protein aggregation; proteostasis; yeast

DOI:  https://doi.org/10.1016/j.jbc.2022.102494
PLoS Biol. 2022 Sep;20(9): e3001753

Primary and metastatic tumors exhibit systems-level differences in dependence on mitochondrial respiratory function.

Neal K Bennett, Hiroki J Nakaoka, Danny Laurent, Ross A Okimoto, Yoshitaka Sei, Andrew E Horvai, Trever G Bivona, Johanna Ten Hoeve, Thomas G Graeber, Ken Nakamura, Jean L Nakamura.

The Warburg effect, aerobic glycolysis, is a hallmark feature of cancer cells grown in culture. However, the relative roles of glycolysis and respiratory metabolism in supporting in vivo tumor growth and processes such as tumor dissemination and metastatic growth remain poorly understood, particularly on a systems level. Using a CRISPRi mini-library enriched for mitochondrial ribosomal protein and respiratory chain genes in multiple human lung cancer cell lines, we analyzed in vivo metabolic requirements in xenograft tumors grown in distinct anatomic contexts. While knockdown of mitochondrial ribosomal protein and respiratory chain genes (mito-respiratory genes) has little impact on growth in vitro, tumor cells depend heavily on these genes when grown in vivo as either flank or primary orthotopic lung tumor xenografts. In contrast, respiratory function is comparatively dispensable for metastatic tumor growth. RNA-Seq and metabolomics analysis of tumor cells expressing individual sgRNAs against mito-respiratory genes indicate overexpression of glycolytic genes and increased sensitivity of glycolytic inhibition compared to control when grown in vitro, but when grown in vivo as primary tumors these cells down-regulate glycolytic mechanisms. These studies demonstrate that discrete perturbations of mitochondrial respiratory chain function impact in vivo tumor growth in a context-specific manner with differential impacts on primary and metastatic tumors.

DOI: https://doi.org/10.1371/journal.pbio.3001753
PLoS Genet. 2022 Sep 20. 18(9): e1010400

Oocyte mitophagy is critical for extended reproductive longevity.

Vanessa Cota, Salman Sohrabi, Rachel Kaletsky, Coleen T Murphy.

Women's reproductive cessation is the earliest sign of human aging and is caused by decreasing oocyte quality. Similarly, C. elegans' reproduction declines in mid-adulthood and is caused by oocyte quality decline. Aberrant mitochondrial morphology is a hallmark of age-related dysfunction, but the role of mitochondrial morphology and dynamics in reproductive aging is unclear. We examined the requirements for mitochondrial fusion and fission in oocytes of both wild-type worms and the long-lived, long-reproducing insulin-like receptor mutant daf-2. We find that normal reproduction requires both fusion and fission, but that daf-2 mutants utilize a shift towards fission, but not fusion, to extend their reproductive span and oocyte health. daf-2 mutant oocytes' mitochondria are punctate (fissioned) and this morphology is primed for mitophagy, as loss of the mitophagy regulator PINK-1 shortens daf-2's reproductive span. daf-2 mutants maintain oocyte mitochondria quality with age at least in part through a shift toward punctate mitochondrial morphology and subsequent mitophagy. Supporting this model, Urolithin A, a metabolite that promotes mitophagy, extends reproductive span in wild-type mothers-even in mid-reproduction-by maintaining youthful oocytes with age. Our data suggest that promotion of mitophagy may be an effective strategy to maintain oocyte health with age.

DOI: https://doi.org/10.1371/journal.pgen.1010400
STAR Protoc. 2022 Sep 21. pii: S2666-1667(22)00590-1. [Epub ahead of print]3(4): 101710

Protocol to detect in vitro and in cell ubiquitylation of mitochondrial DNA polymerase gamma by mitochondrial E3 ligase MITOL.

Mansoor Hussain, Shabnam Saifi, Aftab Mohammed, Sagar Sengupta.

  Mitochondrial polymerase gamma (PolγA) is the only replicative polymerase in mitochondria. To determine PolγA ubiquitylation in cells, Flag-PolγA and MITOL are overexpressed, and subsequently the immunoprecipitated Flag-PolγA is checked for ubiquitylation. Alternately, in vitro synthesized PolγA and MITOL are used to determine whether PolγA is ubiquitylated. Either anti-ubiquitin or anti-Flag antibody is used to detect the ubiquitylated product. Thus, we provide a detailed, reliable, highly reproducible protocol for detecting ubiquitylation of PolγA by MITOL, both in cells and in vitro. For complete details on the use and execution of this protocol, please refer to Hussain et al. (2021).

Keywords:  Cell biology; Genetics; Molecular biology; Protein biochemistry; Protein expression and purification

DOI:  https://doi.org/10.1016/j.xpro.2022.101710