bims-nenemi Biomed News
on Neuroinflammation, neurodegeneration and mitochondria
Issue of 2024–04–28
eleven papers selected by
Marco Tigano, Thomas Jefferson University
  1. OMA1-Mediated Mitochondrial Dynamics Balance Organellar Homeostasis Upstream of Cellular Stress Responses.
  2. Single-cell analysis reveals context-dependent, cell-level selection of mtDNA.
  3. Zebrafish polg2 knock-out recapitulates human POLG-disorders; implications for drug treatment.
  4. Mitofusin-mediated contacts between mitochondria and peroxisomes regulate mitochondrial fusion.
  5. IRF3 regulates neuroinflammatory responses and the expression of genes associated with Alzheimer's disease.
  6. Classification and functional characterization of regulators of intracellular STING trafficking identified by genome-wide optical pooled screening.
  7. SUCLG1 restricts POLRMT succinylation to enhance mitochondrial biogenesis and leukemia progression.
  8. Replicative senescence and high glucose induce the accrual of self-derived cytosolic nucleic acids in human endothelial cells.
  9. Mitochondria inside acute myeloid leukemia cells hydrolyze ATP to resist chemotherapy.
  10. Nucleoside supplements as treatments for mitochondrial DNA depletion syndrome.
  11. Mitochondrial and Nuclear DNA Variants in Amyotrophic Lateral Sclerosis: Enrichment in the Mitochondrial Control Region and Sirtuin Pathway Genes in Spinal Cord Tissue.
  1. Int J Mol Sci. 2024 Apr 22. pii: 4566. [Epub ahead of print]25(8):
    OMA1-Mediated Mitochondrial Dynamics Balance Organellar Homeostasis Upstream of Cellular Stress Responses.
    Robert Gilkerson, Harpreet Kaur, Omar Carrillo, Isaiah Ramos.
      In response to cellular metabolic and signaling cues, the mitochondrial network employs distinct sets of membrane-shaping factors to dynamically modulate organellar structures through a balance of fission and fusion. While these organellar dynamics mediate mitochondrial structure/function homeostasis, they also directly impact critical cell-wide signaling pathways such as apoptosis, autophagy, and the integrated stress response (ISR). Mitochondrial fission is driven by the recruitment of the cytosolic dynamin-related protein-1 (DRP1), while fusion is carried out by mitofusins 1 and 2 (in the outer membrane) and optic atrophy-1 (OPA1) in the inner membrane. This dynamic balance is highly sensitive to cellular stress; when the transmembrane potential across the inner membrane (Δψm) is lost, fusion-active OPA1 is cleaved by the overlapping activity with m-AAA protease-1 (OMA1 metalloprotease, disrupting mitochondrial fusion and leaving dynamin-related protein-1 (DRP1)-mediated fission unopposed, thus causing the collapse of the mitochondrial network to a fragmented state. OMA1 is a unique regulator of stress-sensitive homeostatic mitochondrial balance, acting as a key upstream sensor capable of priming the cell for apoptosis, autophagy, or ISR signaling cascades. Recent evidence indicates that higher-order macromolecular associations within the mitochondrial inner membrane allow these specialized domains to mediate crucial organellar functionalities.
    Keywords:  DRP1; OMA1; OPA1; apoptosis; autophagy; bioenergetics; cristae; fission; fusion; integrated stress response; mitochondria; transmembrane potential
    DOI:  https://doi.org/10.3390/ijms25084566
  2. Nature. 2024 Apr 24.
    Single-cell analysis reveals context-dependent, cell-level selection of mtDNA.
    Anna V Kotrys, Timothy J Durham, Xiaoyan A Guo, Venkata R Vantaku, Sareh Parangi, Vamsi K Mootha.
      Heteroplasmy occurs when wild-type and mutant mitochondrial DNA (mtDNA) molecules co-exist in single cells1. Heteroplasmy levels change dynamically in development, disease and ageing2,3, but it is unclear whether these shifts are caused by selection or drift, and whether they occur at the level of cells or intracellularly. Here we investigate heteroplasmy dynamics in dividing cells by combining precise mtDNA base editing (DdCBE)4 with a new method, SCI-LITE (single-cell combinatorial indexing leveraged to interrogate targeted expression), which tracks single-cell heteroplasmy with ultra-high throughput. We engineered cells to have synonymous or nonsynonymous complex I mtDNA mutations and found that cell populations in standard culture conditions purge nonsynonymous mtDNA variants, whereas synonymous variants are maintained. This suggests that selection dominates over simple drift in shaping population heteroplasmy. We simultaneously tracked single-cell mtDNA heteroplasmy and ancestry, and found that, although the population heteroplasmy shifts, the heteroplasmy of individual cell lineages remains stable, arguing that selection acts at the level of cell fitness in dividing cells. Using these insights, we show that we can force cells to accumulate high levels of truncating complex I mtDNA heteroplasmy by placing them in environments where loss of biochemical complex I activity has been reported to benefit cell fitness. We conclude that in dividing cells, a given nonsynonymous mtDNA heteroplasmy can be harmful, neutral or even beneficial to cell fitness, but that the 'sign' of the effect is wholly dependent on the environment.
    DOI:  https://doi.org/10.1038/s41586-024-07332-0
  3. Cell Death Dis. 2024 Apr 20. 15(4): 281
    Zebrafish polg2 knock-out recapitulates human POLG-disorders; implications for drug treatment.
    Raquel Brañas Casas, Alessandro Zuppardo, Giovanni Risato, Alberto Dinarello, Rudy Celeghin, Camilla Fontana, Eleonora Grelloni, Alexandru Ionut Gilea, Carlo Viscomi, Andrea Rasola, Luisa Dalla Valle, Tiziana Lodi, Enrico Baruffini, Nicola Facchinello, Francesco Argenton, Natascia Tiso.
      The human mitochondrial DNA polymerase gamma is a holoenzyme, involved in mitochondrial DNA (mtDNA) replication and maintenance, composed of a catalytic subunit (POLG) and a dimeric accessory subunit (POLG2) conferring processivity. Mutations in POLG or POLG2 cause POLG-related diseases in humans, leading to a subset of Mendelian-inherited mitochondrial disorders characterized by mtDNA depletion (MDD) or accumulation of multiple deletions, presenting multi-organ defects and often leading to premature death at a young age. Considering the paucity of POLG2 models, we have generated a stable zebrafish polg2 mutant line (polg2ia304) by CRISPR/Cas9 technology, carrying a 10-nucleotide deletion with frameshift mutation and premature stop codon. Zebrafish polg2 homozygous mutants present slower development and decreased viability compared to wild type siblings, dying before the juvenile stage. Mutants display a set of POLG-related phenotypes comparable to the symptoms of human patients affected by POLG-related diseases, including remarkable MDD, altered mitochondrial network and dynamics, and reduced mitochondrial respiration. Histological analyses detected morphological alterations in high-energy demanding tissues, along with a significant disorganization of skeletal muscle fibres. Consistent with the last finding, locomotor assays highlighted a decreased larval motility. Of note, treatment with the Clofilium tosylate drug, previously shown to be effective in POLG models, could partially rescue MDD in Polg2 mutant animals. Altogether, our results point at zebrafish as an effective model to study the etiopathology of human POLG-related disorders linked to POLG2, and a suitable platform to screen the efficacy of POLG-directed drugs in POLG2-associated forms.
    DOI:  https://doi.org/10.1038/s41419-024-06622-9
  4. PLoS Biol. 2024 Apr 26. 22(4): e3002602
    Mitofusin-mediated contacts between mitochondria and peroxisomes regulate mitochondrial fusion.
    Cynthia Alsayyah, Manish K Singh, Maria Angeles Morcillo-Parra, Laetitia Cavellini, Nadav Shai, Christine Schmitt, Maya Schuldiner, Einat Zalckvar, Adeline Mallet, Naïma Belgareh-Touzé, Christophe Zimmer, Mickaël M Cohen.
      Mitofusins are large GTPases that trigger fusion of mitochondrial outer membranes. Similarly to the human mitofusin Mfn2, which also tethers mitochondria to the endoplasmic reticulum (ER), the yeast mitofusin Fzo1 stimulates contacts between Peroxisomes and Mitochondria when overexpressed. Yet, the physiological significance and function of these "PerMit" contacts remain unknown. Here, we demonstrate that Fzo1 naturally localizes to peroxisomes and promotes PerMit contacts in physiological conditions. These contacts are regulated through co-modulation of Fzo1 levels by the ubiquitin-proteasome system (UPS) and by the desaturation status of fatty acids (FAs). Contacts decrease under low FA desaturation but reach a maximum during high FA desaturation. High-throughput genetic screening combined with high-resolution cellular imaging reveal that Fzo1-mediated PerMit contacts favor the transit of peroxisomal citrate into mitochondria. In turn, citrate enters the TCA cycle to stimulate the mitochondrial membrane potential and maintain efficient mitochondrial fusion upon high FA desaturation. These findings thus unravel a mechanism by which inter-organelle contacts safeguard mitochondrial fusion.
    DOI:  https://doi.org/10.1371/journal.pbio.3002602
  5. bioRxiv. 2024 Mar 12. pii: 2024.03.08.582968. [Epub ahead of print]
    IRF3 regulates neuroinflammatory responses and the expression of genes associated with Alzheimer's disease.
    Radhika Joshi, Veronika Brezani, Gabrielle M Mey, Sergi Guixé-Muntet, Marti Ortega-Ribera, Yuan Zhuang, Adam Zivny, Sebastian Werneburg, Jordi Gracia-Sancho, Gyongyi Szabo.
      The pathological role of interferon signaling is emerging in neuroinflammatory disorders, yet, the specific role of Interferon Regulatory Factor 3 (IRF3) in neuroinflammation remains poorly understood. Here, we show that global IRF3 deficiency delays TLR4-mediated signaling in microglia and attenuates the hallmark features of LPS-induced inflammation such as cytokine release, microglial reactivity, astrocyte activation, myeloid cell infiltration, and inflammasome activation. Moreover, expression of a constitutively active IRF3 (S388D/S390D:IRF3-2D) in microglia induces a transcriptional program reminiscent of the Activated Response Microglia and the expression of genes associated with Alzheimer's Disease, notably apolipoprotein-e . Lastly, using bulk-RNAseq of IRF3-2D brain myeloid cells, we identified Z-DNA binding protein-1 as a target of IRF3 that is relevant across various neuroinflammatory disorders. Together, our results identify IRF3 as an important regulator of LPS-mediated neuroinflammatory responses and highlight IRF3 as a central regulator of disease-specific gene activation in different neuroinflammatory diseases.
    DOI:  https://doi.org/10.1101/2024.03.08.582968
  6. bioRxiv. 2024 Apr 09. pii: 2024.04.07.588166. [Epub ahead of print]
    Classification and functional characterization of regulators of intracellular STING trafficking identified by genome-wide optical pooled screening.
    Matteo Gentili, Rebecca J Carlson, Bingxu Liu, Quentin Hellier, Jocelyn Andrews, Yue Qin, Paul C Blainey, Nir Hacohen.
      STING is an innate immune sensor that traffics across many cellular compartments to carry out its function of detecting cyclic di-nucleotides and triggering defense processes. Mutations in factors that regulate this process are often linked to STING-dependent human inflammatory disorders. To systematically identify factors involved in STING trafficking, we performed a genome-wide optical pooled screen and examined the impact of genetic perturbations on intracellular STING localization. Based on subcellular imaging of STING protein and trafficking markers in 45 million cells perturbed with sgRNAs, we defined 464 clusters of gene perturbations with similar cellular phenotypes. A higher-dimensional focused optical pooled screen on 262 perturbed genes which assayed 11 imaging channels identified 73 finer phenotypic clusters. In a cluster containing USE1, a protein that mediates Golgi to ER transport, we found a gene of unknown function, C19orf25. Consistent with the known role of USE1, loss of C19orf25 enhanced STING signaling. Other clusters contained subunits of the HOPS, GARP and RIC1-RGP1 complexes. We show that HOPS deficiency delayed STING degradation and consequently increased signaling. Similarly, GARP/RIC1-RGP1 loss increased STING signaling by delaying STING exit from the Golgi. Our findings demonstrate that genome-wide genotype-phenotype maps based on high-content cell imaging outperform other screening approaches, and provide a community resource for mining for factors that impact STING trafficking as well as other cellular processes observable in our dataset.
    DOI:  https://doi.org/10.1101/2024.04.07.588166
  7. EMBO J. 2024 Apr 22.
    SUCLG1 restricts POLRMT succinylation to enhance mitochondrial biogenesis and leukemia progression.
    Weiwei Yan, Chengmei Xie, Sijun Sun, Quan Zheng, Jingyi Wang, Zihao Wang, Cheuk-Him Man, Haiyan Wang, Yunfan Yang, Tianshi Wang, Leilei Shi, Shengjie Zhang, Chen Huang, Shuangnian Xu, Yi-Ping Wang.
      Mitochondria are cellular powerhouses that generate energy through the electron transport chain (ETC). The mitochondrial genome (mtDNA) encodes essential ETC proteins in a compartmentalized manner, however, the mechanism underlying metabolic regulation of mtDNA function remains unknown. Here, we report that expression of tricarboxylic acid cycle enzyme succinate-CoA ligase SUCLG1 strongly correlates with ETC genes across various TCGA cancer transcriptomes. Mechanistically, SUCLG1 restricts succinyl-CoA levels to suppress the succinylation of mitochondrial RNA polymerase (POLRMT). Lysine 622 succinylation disrupts the interaction of POLRMT with mtDNA and mitochondrial transcription factors. SUCLG1-mediated POLRMT hyposuccinylation maintains mtDNA transcription, mitochondrial biogenesis, and leukemia cell proliferation. Specifically, leukemia-promoting FMS-like tyrosine kinase 3 (FLT3) mutations modulate nuclear transcription and upregulate SUCLG1 expression to reduce succinyl-CoA and POLRMT succinylation, resulting in enhanced mitobiogenesis. In line, genetic depletion of POLRMT or SUCLG1 significantly delays disease progression in mouse and humanized leukemia models. Importantly, succinyl-CoA level and POLRMT succinylation are downregulated in FLT3-mutated clinical leukemia samples, linking enhanced mitobiogenesis to cancer progression. Together, SUCLG1 connects succinyl-CoA with POLRMT succinylation to modulate mitochondrial function and cancer development.
    Keywords:  FMS-like Tyrosine Kinase 3; Lysine Succinylation; Mitochondrial Biogenesis; Mitochondrial RNA Polymerase; Succinate-CoA Ligase
    DOI:  https://doi.org/10.1038/s44318-024-00101-9
  8. Cell Death Discov. 2024 Apr 20. 10(1): 184
    Replicative senescence and high glucose induce the accrual of self-derived cytosolic nucleic acids in human endothelial cells.
    Deborah Ramini, Angelica Giuliani, Katarzyna Malgorzata Kwiatkowska, Michele Guescini, Gianluca Storci, Emanuela Mensà, Rina Recchioni, Luciano Xumerle, Elisa Zago, Jacopo Sabbatinelli, Spartaco Santi, Paolo Garagnani, Massimiliano Bonafè, Fabiola Olivieri.
      Recent literature shows that loss of replicative ability and acquisition of a proinflammatory secretory phenotype in senescent cells is coupled with the build-in of nucleic acids in the cytoplasm. Its implication in human age-related diseases is under scrutiny. In human endothelial cells (ECs), we assessed the accumulation of intracellular nucleic acids during in vitro replicative senescence and after exposure to high glucose concentrations, which mimic an in vivo condition of hyperglycemia. We showed that exposure to high glucose induces senescent-like features in ECs, including telomere shortening and proinflammatory cytokine release, coupled with the accrual in the cytoplasm of telomeres, double-stranded DNA and RNA (dsDNA, dsRNA), as well as RNA:DNA hybrid molecules. Senescent ECs showed an activation of the dsRNA sensors RIG-I and MDA5 and of the DNA sensor TLR9, which was not paralleled by the involvement of the canonical (cGAS) and non-canonical (IFI16) activation of the STING pathway. Under high glucose conditions, only a sustained activation of TLR9 was observed. Notably, senescent cells exhibit increased proinflammatory cytokine (IL-1β, IL-6, IL-8) production without a detectable secretion of type I interferon (IFN), a phenomenon that can be explained, at least in part, by the accumulation of methyl-adenosine containing RNAs. At variance, exposure to exogenous nucleic acids enhances both IL-6 and IFN-β1 expression in senescent cells. This study highlights the accrual of cytoplasmic nucleic acids as a marker of senescence-related endothelial dysfunction, that may play a role in dysmetabolic age-related diseases.
    DOI:  https://doi.org/10.1038/s41420-024-01954-z
  9. bioRxiv. 2024 Apr 15. pii: 2024.04.12.589110. [Epub ahead of print]
    Mitochondria inside acute myeloid leukemia cells hydrolyze ATP to resist chemotherapy.
    James T Hagen, Mclane M Montgomery, Raphael T Aruleba, Brett R Chrest, Thomas D Green, Miki Kassai, Tonya N Zeczycki, Cameron A Schmidt, Debajit Bhowmick, Su-Fern Tan, David J Feith, Charles E Chalfant, Thomas P Loughran, Darla Liles, Mark D Minden, Aaron D Schimmer, Myles C Cabot, Joseph M Mclung, Kelsey H Fisher-Wellman.
      Despite early optimism, therapeutics targeting oxidative phosphorylation (OxPhos) have faced clinical setbacks, stemming from their inability to distinguish healthy from cancerous mitochondria. Herein, we describe an actionable bioenergetic mechanism unique to cancerous mitochondria inside acute myeloid leukemia (AML) cells. Unlike healthy cells which couple respiration to the synthesis of ATP, AML mitochondria were discovered to support inner membrane polarization by consuming ATP. Because matrix ATP consumption allows cells to survive bioenergetic stress, we hypothesized that AML cells may resist cell death induced by OxPhos damaging chemotherapy by reversing the ATP synthase reaction. In support of this, targeted inhibition of BCL-2 with venetoclax abolished OxPhos flux without impacting mitochondrial membrane potential. In surviving AML cells, sustained polarization of the mitochondrial inner membrane was dependent on matrix ATP consumption. Mitochondrial ATP consumption was further enhanced in AML cells made refractory to venetoclax, consequential to downregulations in both the proton-pumping respiratory complexes, as well as the endogenous F 1 -ATPase inhibitor ATP5IF1 . In treatment-naive AML, ATP5IF1 knockdown was sufficient to drive venetoclax resistance, while ATP5IF1 overexpression impaired F 1 -ATPase activity and heightened sensitivity to venetoclax. Collectively, our data identify matrix ATP consumption as a cancer-cell intrinsic bioenergetic vulnerability actionable in the context of mitochondrial damaging chemotherapy.
    DOI:  https://doi.org/10.1101/2024.04.12.589110
  10. Front Cell Dev Biol. 2024 ;12 1260496
    Nucleoside supplements as treatments for mitochondrial DNA depletion syndrome.
    Eszter Dombi, Tony Marinaki, Paolo Spingardi, Val Millar, Nastasia Hadjichristou, Janet Carver, Iain G Johnston, Carl Fratter, Joanna Poulton.
      Introduction: In mitochondrial DNA (mtDNA) depletion syndrome (MDS), patients cannot maintain sufficient mtDNA for their energy needs. MDS presentations range from infantile encephalopathy with hepatopathy (Alpers syndrome) to adult chronic progressive external ophthalmoplegia. Most are caused by nucleotide imbalance or by defects in the mtDNA replisome. There is currently no curative treatment available. Nucleoside therapy is a promising experimental treatment for TK2 deficiency, where patients are supplemented with exogenous deoxypyrimidines. We aimed to explore the benefits of nucleoside supplementation in POLG and TWNK deficient fibroblasts. Methods: We used high-content fluorescence microscopy with software-based image analysis to assay mtDNA content and membrane potential quantitatively, using vital dyes PicoGreen and MitoTracker Red CMXRos respectively. We tested the effect of 15 combinations (A, T, G, C, AT, AC, AG, CT, CG, GT, ATC, ATG, AGC, TGC, ATGC) of deoxynucleoside supplements on mtDNA content of fibroblasts derived from four patients with MDS (POLG1, POLG2, DGUOK, TWNK) in both a replicating (10% dialysed FCS) and quiescent (0.1% dialysed FCS) state. We used qPCR to measure mtDNA content of supplemented and non-supplemented fibroblasts following mtDNA depletion using 20 µM ddC and after 14- and 21-day recovery in a quiescent state. Results: Nucleoside treatments at 200 µM that significantly increased mtDNA content also significantly reduced the number of cells remaining in culture after 7 days of treatment, as well as mitochondrial membrane potential. These toxic effects were abolished by reducing the concentration of nucleosides to 50 µM. In POLG1 and TWNK cells the combination of ATGC treatment increased mtDNA content the most after 7 days in non-replicating cells. ATGC nucleoside combination significantly increased the rate of mtDNA recovery in quiescent POLG1 cells following mtDNA depletion by ddC. Conclusion: High-content imaging enabled us to link mtDNA copy number with key read-outs linked to patient wellbeing. Elevated G increased mtDNA copy number but severely impaired fibroblast growth, potentially by inhibiting purine synthesis and/or causing replication stress. Combinations of nucleosides ATGC, T, or TC, benefited growth of cells harbouring POLG mutations. These combinations, one of which reflects a commercially available preparation, could be explored further for treatment of POLG patients.
    Keywords:  POLG; TWNK; alpers syndrome; heavy isotope labelling mass spectroscopy; high-content imaging; mitochondrial DNA; mitochondrial DNA depletion syndrome; nucleoside bypass therapy
    DOI:  https://doi.org/10.3389/fcell.2024.1260496
  11. Biomolecules. 2024 Mar 28. pii: 411. [Epub ahead of print]14(4):
    Mitochondrial and Nuclear DNA Variants in Amyotrophic Lateral Sclerosis: Enrichment in the Mitochondrial Control Region and Sirtuin Pathway Genes in Spinal Cord Tissue.
    Sharon Natasha Cox, Claudio Lo Giudice, Anna Lavecchia, Maria Luana Poeta, Matteo Chiara, Ernesto Picardi, Graziano Pesole.
      Amyotrophic Lateral Sclerosis (ALS) is a progressive disease with prevalent mitochondrial dysfunctions affecting both upper and lower motor neurons in the motor cortex, brainstem, and spinal cord. Despite mitochondria having their own genome (mtDNA), in humans, most mitochondrial genes are encoded by the nuclear genome (nDNA). Our study aimed to simultaneously screen for nDNA and mtDNA genomes to assess for specific variant enrichment in ALS compared to control tissues. Here, we analysed whole exome (WES) and whole genome (WGS) sequencing data from spinal cord tissues, respectively, of 6 and 12 human donors. A total of 31,257 and 301,241 variants in nuclear-encoded mitochondrial genes were identified from WES and WGS, respectively, while mtDNA reads accounted for 73 and 332 variants. Despite technical differences, both datasets consistently revealed a specific enrichment of variants in the mitochondrial Control Region (CR) and in several of these genes directly associated with mitochondrial dynamics or with Sirtuin pathway genes within ALS tissues. Overall, our data support the hypothesis of a variant burden in specific genes, highlighting potential actionable targets for therapeutic interventions in ALS.
    Keywords:  Amyotrophic Lateral Sclerosis; WES; WGS; bioinformatic pipeline; heteroplasmy; mtDNA; nDNA; variants
    DOI:  https://doi.org/10.3390/biom14040411
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