bims-nenemi 2021-12-26 papers

bims-nenemi

Biomed News

on Neuroinflammation, neurodegeneration and mitochondria

Issue of 2021–12–26
thirteen papers selected by
Marco Tigano, Thomas Jefferson University

BCL-2-family protein tBID can act as a BAX-like effector of apoptosis.
Mitochondrial DNA damage as driver of cellular outcomes.
ATAD3A has a scaffolding role regulating mitochondria inner membrane structure and protein assembly.
Cytosolic Self-DNA-A Potential Source of Chronic Inflammation in Aging.
Implications of Membrane Binding by the Fe-S Cluster-Containing N-Terminal Domain in the Drosophila Mitochondrial Replicative DNA Helicase.
A minimal motif for sequence recognition by mitochondrial transcription factor A (TFAM).
Inactivity of Peptidase ClpP Causes Primary Accumulation of Mitochondrial Disaggregase ClpX with Its Interacting Nucleoid Proteins, and of mtDNA.
DdCBE mediates efficient and inheritable modifications in mouse mitochondrial genome.
Transcriptomic profiling and pathway analysis of cultured human lung microvascular endothelial cells following ionizing radiation exposure.
Inhibition of the Anti-Apoptotic Bcl-2 Family by BH3 Mimetics Sensitize the Mitochondrial Permeability Transition Pore Through Bax and Bak.
Activation mechanism of PINK1.
Mitochondrial C5aR1 activity in macrophages controls IL-1β production underlying sterile inflammation.
Peroxiredoxin 6 protects irradiated cells from oxidative stress and shapes their senescence-associated cytokine landscape.

EMBO J. 2021 Dec 21. e108690

BCL-2-family protein tBID can act as a BAX-like effector of apoptosis.

Hector Flores-Romero, Lisa Hohorst, Malina John, Marie-Christine Albert, Louise E King, Laura Beckmann, Tamas Szabo, Vanessa Hertlein, Xu Luo, Andreas Villunger, Lukas P Frenzel, Hamid Kashkar, Ana J Garcia-Saez.

  During apoptosis, the BCL-2-family protein tBID promotes mitochondrial permeabilization by activating BAX and BAK and by blocking anti-apoptotic BCL-2 members. Here, we report that tBID can also mediate mitochondrial permeabilization by itself, resulting in release of cytochrome c and mitochondrial DNA, caspase activation and apoptosis even in absence of BAX and BAK. This previously unrecognized activity of tBID depends on helix 6, homologous to the pore-forming regions of BAX and BAK, and can be blocked by pro-survival BCL-2 proteins. Importantly, tBID-mediated mitochondrial permeabilization independent of BAX and BAK is physiologically relevant for SMAC release in the immune response against Shigella infection. Furthermore, it can be exploited to kill leukaemia cells with acquired venetoclax resistance due to lack of active BAX and BAK. Our findings define tBID as an effector of mitochondrial permeabilization in apoptosis and provide a new paradigm for BCL-2 proteins, with implications for anti-bacterial immunity and cancer therapy.

Keywords:  BCL-2 proteins; apoptosis; mitochondrial permeabilization; pore formation

DOI:  https://doi.org/10.15252/embj.2021108690
Am J Physiol Cell Physiol. 2021 Dec 22.

Mitochondrial DNA damage as driver of cellular outcomes.

Cristina A Nadalutti, Sylvette Ayala-Peña, Janine H Santos.

  Mitochondria are primarily involved in energy production through the process of oxidative phosphorylation (OXPHOS). Increasing evidence has shown that mitochondrial function impacts a plethora of different cellular activities, including metabolism, epigenetics and innate immunity. Like the nucleus, mitochondria own their genetic material, which is maternally inherited. The mitochondrial DNA (mtDNA) encodes 37 genes that are solely involved in OXPHOS. Maintenance of mtDNA, through replication and repair, requires the import of nuclear DNA encoded proteins. Thus, mitochondria completely rely on the nucleus to prevent mitochondrial genetic alterations. As every cell contains hundreds to thousands of mitochondria, it follows that the shear number of organelles allow for the buffering of dysfunction - at least to some extent - before tissue homeostasis becomes impaired. Only red blood cells lack mitochondria entirely. Impaired mitochondrial function is a hallmark of aging and is involved in a number of different disorders, including neurodegenerative diseases, diabetes, cancer, and autoimmunity. While alterations in mitochondrial processes unrelated to OXPHOS, such as fusion and fission, contribute to aging and disease, maintenance of mtDNA integrity is critical for proper organellar function. Here, we focus on how mtDNA damage contributes to cellular dysfunction and health outcomes.

Keywords:  DNA repair; cellular outcomes; mitochondrial dysfunction; mtDNA damage

DOI:  https://doi.org/10.1152/ajpcell.00389.2021
Cell Rep. 2021 Dec 21. pii: S2211-1247(21)01635-1. [Epub ahead of print]37(12): 110139

ATAD3A has a scaffolding role regulating mitochondria inner membrane structure and protein assembly.

Tania Arguello, Susana Peralta, Hana Antonicka, Gabriel Gaidosh, Francisca Diaz, Ya-Ting Tu, Sofia Garcia, Ramin Shiekhattar, Antonio Barrientos, Carlos T Moraes.

  The ATPase Family AAA Domain Containing 3A (ATAD3A), is a mitochondrial inner membrane protein conserved in metazoans. ATAD3A has been associated with several mitochondrial functions, including nucleoid organization, cholesterol metabolism, and mitochondrial translation. To address its primary role, we generated a neuronal-specific conditional knockout (Atad3 nKO) mouse model, which developed a severe encephalopathy by 5 months of age. Pre-symptomatic mice showed aberrant mitochondrial cristae morphogenesis in the cortex as early as 2 months. Using a multi-omics approach in the CNS of 2-to-3-month-old mice, we found early alterations in the organelle membrane structure. We also show that human ATAD3A associates with different components of the inner membrane, including OXPHOS complex I, Letm1, and prohibitin complexes. Stochastic Optical Reconstruction Microscopy (STORM) shows that ATAD3A is regularly distributed along the inner mitochondrial membrane, suggesting a critical structural role in inner mitochondrial membrane and its organization, most likely in an ATPase-dependent manner.

Keywords:  ATAD3; cardiolipin; cristae; inner membrane; mitochondria

DOI:  https://doi.org/10.1016/j.celrep.2021.110139
Cells. 2021 Dec 15. pii: 3544. [Epub ahead of print]10(12):

Cytosolic Self-DNA-A Potential Source of Chronic Inflammation in Aging.

Mansour Akbari, Daryl P Shanley, Vilhelm A Bohr, Lene Juel Rasmussen.

  Aging is the consequence of a lifelong accumulation of stochastic damage to tissues and cellular components. Advancing age closely associates with elevated markers of innate immunity and low-grade chronic inflammation, probably reflecting steady increasing incidents of cellular and tissue damage over the life course. The DNA sensing cGAS-STING signaling pathway is activated by misplaced cytosolic self-DNA, which then initiates the innate immune responses. Here, we hypothesize that the stochastic release of various forms of DNA from the nucleus and mitochondria, e.g., because of DNA damage, altered nucleus integrity, and mitochondrial damage, can result in chronic activation of inflammatory responses that characterize the aging process. This cytosolic self-DNA-innate immunity axis may perturb tissue homeostasis and function that characterizes human aging and age-associated pathology. Proper techniques and experimental models are available to investigate this axis to develop therapeutic interventions.

Keywords:  DNA repair; aging; cGAS-STING; cytosolic self-DNA; inflammation; mitochondria

DOI:  https://doi.org/10.3390/cells10123544
Front Genet. 2021 ;12 790521

Implications of Membrane Binding by the Fe-S Cluster-Containing N-Terminal Domain in the Drosophila Mitochondrial Replicative DNA Helicase.

Minyoung So, Johnny Stiban, Grzegorz L Ciesielski, Stacy L Hovde, Laurie S Kaguni.

  Recent evidence suggests that iron-sulfur clusters (ISCs) in DNA replicative proteins sense DNA-mediated charge transfer to modulate nuclear DNA replication. In the mitochondrial DNA replisome, only the replicative DNA helicase (mtDNA helicase) from Drosophila melanogaster (Dm) has been shown to contain an ISC in its N-terminal, primase-like domain (NTD). In this report, we confirm the presence of the ISC and demonstrate the importance of a metal cofactor in the structural stability of the Dm mtDNA helicase. Further, we show that the NTD also serves a role in membrane binding. We demonstrate that the NTD binds to asolectin liposomes, which mimic phospholipid membranes, through electrostatic interactions. Notably, membrane binding is more specific with increasing cardiolipin content, which is characteristically high in the mitochondrial inner membrane (MIM). We suggest that the N-terminal domain of the mtDNA helicase interacts with the MIM to recruit mtDNA and initiate mtDNA replication. Furthermore, Dm NUBPL, the known ISC donor for respiratory complex I and a putative donor for Dm mtDNA helicase, was identified as a peripheral membrane protein that is likely to execute membrane-mediated ISC delivery to its target proteins.

Keywords:  NUBPL/Ind1; genome stability; iron-sulfur clusters; liposomes; membrane binding; mitochondria; replicative helicase

DOI:  https://doi.org/10.3389/fgene.2021.790521
Nucleic Acids Res. 2021 Dec 20. pii: gkab1230. [Epub ahead of print]

A minimal motif for sequence recognition by mitochondrial transcription factor A (TFAM).

Woo Suk Choi, Miguel Garcia-Diaz.

Mitochondrial transcription factor A (TFAM) plays a critical role in mitochondrial transcription initiation and mitochondrial DNA (mtDNA) packaging. Both functions require DNA binding, but in one case TFAM must recognize a specific promoter sequence, while packaging requires coating of mtDNA by association with non sequence-specific regions. The mechanisms by which TFAM achieves both sequence-specific and non sequence-specific recognition have not yet been determined. Existing crystal structures of TFAM bound to DNA allowed us to identify two guanine-specific interactions that are established between TFAM and the bound DNA. These interactions are observed when TFAM is bound to both specific promoter sequences and non-sequence specific DNA. These interactions are established with two guanine bases separated by 10 random nucleotides (GN10G). Our biochemical results demonstrate that the GN10G consensus is essential for transcriptional initiation and contributes to facilitating TFAM binding to DNA substrates. Furthermore, we report a crystal structure of TFAM in complex with a non sequence-specific sequence containing a GN10G consensus. The structure reveals a unique arrangement in which TFAM bridges two DNA substrates while maintaining the GN10G interactions. We propose that the GN10G consensus is key to facilitate the interaction of TFAM with DNA.

DOI: https://doi.org/10.1093/nar/gkab1230
Cells. 2021 Nov 29. pii: 3354. [Epub ahead of print]10(12):

Inactivity of Peptidase ClpP Causes Primary Accumulation of Mitochondrial Disaggregase ClpX with Its Interacting Nucleoid Proteins, and of mtDNA.

Jana Key, Sylvia Torres-Odio, Nina C Bach, Suzana Gispert, Gabriele Koepf, Marina Reichlmeir, A Phillip West, Holger Prokisch, Peter Freisinger, William G Newman, Stavit Shalev, Stephan A Sieber, Ilka Wittig, Georg Auburger.

  Biallelic pathogenic variants in CLPP, encoding mitochondrial matrix peptidase ClpP, cause a rare autosomal recessive condition, Perrault syndrome type 3 (PRLTS3). It is characterized by primary ovarian insufficiency and early sensorineural hearing loss, often associated with progressive neurological deficits. Mouse models showed that accumulations of (i) its main protein interactor, the substrate-selecting AAA+ ATPase ClpX, (ii) mitoribosomes, and (iii) mtDNA nucleoids are the main cellular consequences of ClpP absence. However, the sequence of these events and their validity in human remain unclear. Here, we studied global proteome profiles to define ClpP substrates among mitochondrial ClpX interactors, which accumulated consistently in ClpP-null mouse embryonal fibroblasts and brains. Validation work included novel ClpP-mutant patient fibroblast proteomics. ClpX co-accumulated in mitochondria with the nucleoid component POLDIP2, the mitochondrial poly(A) mRNA granule element LRPPRC, and tRNA processing factor GFM1 (in mouse, also GRSF1). Only in mouse did accumulated ClpX, GFM1, and GRSF1 appear in nuclear fractions. Mitoribosomal accumulation was minor. Consistent accumulations in murine and human fibroblasts also affected multimerizing factors not known as ClpX interactors, namely, OAT, ASS1, ACADVL, STOM, PRDX3, PC, MUT, ALDH2, PMPCB, UQCRC2, and ACADSB, but the impact on downstream metabolites was marginal. Our data demonstrate the primary impact of ClpXP on the assembly of proteins with nucleic acids and show nucleoid enlargement in human as a key consequence.

Keywords:  ClpB; ERAL1; HARS2; LARS2; Parkinson’s disease; TWNK; ataxia; leukodystrophy

DOI:  https://doi.org/10.3390/cells10123354
Mol Ther Nucleic Acids. 2022 Mar 08. 27 73-80

DdCBE mediates efficient and inheritable modifications in mouse mitochondrial genome.

Jiayin Guo, Xiaoxu Chen, Zhiwei Liu, Haifeng Sun, Yu Zhou, Yichen Dai, Yu'e Ma, Lei He, Xuezhen Qian, Jianying Wang, Jie Zhang, Yichen Zhu, Jun Zhang, Bin Shen, Fei Zhou.

  Critical mutations of mitochondrial DNA (mtDNA) generally lead to maternally inheritable diseases that affect multiple organs and systems; however, it was difficult to alter mtDNA in mammalian cells to intervene in or cure mitochondrial disorders. Recently, the discovery of DddA-derived cytosine base editor (DdCBE) enabled the precise manipulation of mtDNA. To test its feasibility for in vivo use, we selected several sites in mouse mtDNA as DdCBE targets to resemble the human pathogenic mtDNA G-to-A mutations. The efficiency of DdCBE-mediated mtDNA editing was first screened in mouse Neuro-2A cells and DdCBE pairs with the best performance were chosen for in vivo targeting. Microinjection of the mRNAs of DdCBE halves in the mouse zygotes or 2-cell embryo successfully generated edited founder mice with a base conversion rate ranging from 2.48% to 28.51%. When backcrossed with wild-type male mice, female founders were able to transmit the mutations to their offspring with different mutation loads. Off-target analyses demonstrated a high fidelity for DdCBE-mediated base editing in mouse mtDNA both in vitro and in vivo. Our study demonstrated that the DdCBE is feasible for generation of mtDNA mutation models to facilitate disease study and for potential treatment of mitochondrial disorders.

Keywords:  DdCBE; base editing; mitochondrial disorder; mouse model; mtDNA

DOI:  https://doi.org/10.1016/j.omtn.2021.11.016
Sci Rep. 2021 Dec 20. 11(1): 24214

Transcriptomic profiling and pathway analysis of cultured human lung microvascular endothelial cells following ionizing radiation exposure.

Roxane M Bouten, Clifton L Dalgard, Anthony R Soltis, John E Slaven, Regina M Day.

The vascular system is sensitive to radiation injury, and vascular damage is believed to play a key role in delayed tissue injury such as pulmonary fibrosis. However, the response of endothelial cells to radiation is not completely understood. We examined the response of primary human lung microvascular endothelial cells (HLMVEC) to 10 Gy (1.15 Gy/min) X-irradiation. HLMVEC underwent senescence (80-85%) with no significant necrosis or apoptosis. Targeted RT-qPCR showed increased expression of genes CDKN1A and MDM2 (10-120 min). Western blotting showed upregulation of p2/waf1, MDM2, ATM, and Akt phosphorylation (15 min-72 h). Low levels of apoptosis at 24-72 h were identified using nuclear morphology. To identify novel pathway regulation, RNA-seq was performed on mRNA using time points from 2 to 24 h post-irradiation. Gene ontology and pathway analysis revealed increased cell cycle inhibition, DNA damage response, pro- and anti- apoptosis, and pro-senescence gene expression. Based on published literature on inflammation and endothelial-to-mesenchymal transition (EndMT) pathway genes, we identified increased expression of pro-inflammatory genes and EndMT-associated genes by 24 h. Together our data reveal a time course of integrated gene expression and protein activation leading from early DNA damage response and cell cycle arrest to senescence, pro-inflammatory gene expression, and endothelial-to-mesenchymal transition.

DOI: https://doi.org/10.1038/s41598-021-03636-7
Front Cell Dev Biol. 2021 ;9 765973

Inhibition of the Anti-Apoptotic Bcl-2 Family by BH3 Mimetics Sensitize the Mitochondrial Permeability Transition Pore Through Bax and Bak.

Pooja Patel, Arielys Mendoza, Dexter J Robichaux, Meng C Wang, Xander H T Wehrens, Jason Karch.

  Mitochondrial permeability transition pore (MPTP)-dependent necrosis contributes to numerous pathologies in the heart, brain, and skeletal muscle. The MPTP is a non-selective pore in the inner mitochondrial membrane that is triggered by high levels of matrix Ca2+, and sustained opening leads to mitochondrial dysfunction. Although the MPTP is defined by an increase in inner mitochondrial membrane permeability, the expression of pro-apoptotic Bcl-2 family members, Bax and Bak localization to the outer mitochondrial membrane is required for MPTP-dependent mitochondrial dysfunction and subsequent necrotic cell death. Contrary to the role of Bax and Bak in apoptosis, which is dependent on their oligomerization, MPTP-dependent necrosis does not require oligomerization as monomeric/inactive forms of Bax and Bak can facilitate mitochondrial dysfunction. However, the relationship between Bax and Bak activation/oligomerization and MPTP sensitization remains to be explored. Here, we use a combination of in vitro and ex vivo approaches to determine the role of the anti-apoptotic Bcl-2 family members, which regulate Bax/Bak activity, in necrotic cell death and MPTP sensitivity. To study the role of each predominantly expressed anti-apoptotic Bcl-2 family member (i.e., Mcl-1, Bcl-2, and Bcl-xL) in MPTP regulation, we utilize various BH3 mimetics that specifically bind to and inhibit each. We determined that the inhibition of each anti-apoptotic Bcl-2 family member lowers mitochondrial calcium retention capacity and sensitizes MPTP opening. Furthermore, the inhibition of each Bcl-2 family member exacerbates both apoptotic and necrotic cell death in vitro in a Bax/Bak-dependent manner. Our findings suggests that mitochondrial Ca2+ retention capacity and MPTP sensitivity is influenced by Bax/Bak activation/oligomerization on the outer mitochondrial membrane, providing further evidence of the crosstalk between the apoptotic and necrotic cell death pathways.

Keywords:  BCL-2 family; BH3 mimetics; calcium; mitochondria; mitochondrial dysfunction; necrosis; permeability transition

DOI:  https://doi.org/10.3389/fcell.2021.765973
Nature. 2021 Dec 21.

Activation mechanism of PINK1.

Zhong Yan Gan, Sylvie Callegari, Simon A Cobbold, Thomas R Cotton, Michael J Mlodzianoski, Alexander F Schubert, Niall D Geoghegan, Kelly L Rogers, Andrew Leis, Grant Dewson, Alisa Glukhova, David Komander.

Mutations in the protein kinase PINK1 lead to defects in mitophagy and cause autosomal recessive early onset Parkinson's Disease (EOPD)1,2. PINK1 has many unique features that enable it to phosphorylate ubiquitin and the ubiquitin-like domain of Parkin3-9. Structural analysis of PINK1 from diverse insect species10-12 with and without ubiquitin provided snapshots of distinct structural states yet did not explain how PINK1 is activated. We here elucidate the activation mechanism of PINK1 by crystallography and cryo-EM. A crystal structure of unphosphorylated Pediculus humanus corporis (Ph) PINK1 resolves a previously omitted N-terminal helix revealing how unphosphorylated yet active PINK1 is oriented on mitochondria. We further reveal a 2.35 Å cryo-EM structure of a symmetric PhPINK1 dimer trapped during the process of trans-autophosphorylation, and a 3.1 Å cryo-EM structure of phosphorylated PhPINK1 in the process of undergoing a conformational change to become an active ubiquitin kinase. Structures and phosphorylation studies further identify a role for regulatory PINK1 oxidation. Together, our work delineates the complete activation mechanism of PINK1, illuminates how PINK1 interacts with the mitochondrial outer membrane, and reveals how PINK1 activity may be modulated by mitochondrial reactive oxygen species.

DOI: https://doi.org/10.1038/s41586-021-04340-2
Sci Immunol. 2021 Dec 24. 6(66): eabf2489

Mitochondrial C5aR1 activity in macrophages controls IL-1β production underlying sterile inflammation.

Nathalie Niyonzima, Jubayer Rahman, Natalia Kunz, Erin E West, Tilo Freiwald, Jigar V Desai, Nicolas S Merle, Alexandre Gidon, Bjørnar Sporsheim, Michail S Lionakis, Kristin Evensen, Beate Lindberg, Karolina Skagen, Mona Skjelland, Parul Singh, Markus Haug, Marieta M Ruseva, Martin Kolev, Jack Bibby, Olivia Marshall, Brett O'Brien, Nigel Deeks, Behdad Afzali, Richard J Clark, Trent M Woodruff, Milton Pryor, Zhi-Hong Yang, Alan T Remaley, Tom E Mollnes, Stephen M Hewitt, Bingyu Yan, Majid Kazemian, Máté G Kiss, Christoph J Binder, Bente Halvorsen, Terje Espevik, Claudia Kemper.

[Figure: see text].

DOI: https://doi.org/10.1126/sciimmunol.abf2489
Redox Biol. 2021 Dec 11. pii: S2213-2317(21)00372-4. [Epub ahead of print]49 102212

Peroxiredoxin 6 protects irradiated cells from oxidative stress and shapes their senescence-associated cytokine landscape.

Barbora Salovska, Alexandra Kondelova, Kristyna Pimkova, Zuzana Liblova, Miroslav Pribyl, Ivo Fabrik, Jiri Bartek, Marie Vajrychova, Zdenek Hodny.

  Cellular senescence is a complex stress response defined as an essentially irreversible cell cycle arrest mediated by the inhibition of cell cycle-specific cyclin dependent kinases. The imbalance in redox homeostasis and oxidative stress have been repeatedly observed as one of the hallmarks of the senescent phenotype. However, a large-scale study investigating protein oxidation and redox signaling in senescent cells in vitro has been lacking. Here we applied a proteome-wide analysis using SILAC-iodoTMT workflow to quantitatively estimate the level of protein sulfhydryl oxidation and proteome level changes in ionizing radiation-induced senescence (IRIS) in hTERT-RPE-1 cells. We observed that senescent cells mobilized the antioxidant system to buffer the increased oxidation stress. Among the antioxidant proteins with increased relative abundance in IRIS, a unique 1-Cys peroxiredoxin family member, peroxiredoxin 6 (PRDX6), was identified as an important contributor to protection against oxidative stress. PRDX6 silencing increased ROS production in senescent cells, decreased their resistance to oxidative stress-induced cell death, and impaired their viability. Subsequent SILAC-iodoTMT and secretome analysis after PRDX6 silencing showed the downregulation of PRDX6 in IRIS affected protein secretory pathways, decreased expression of extracellular matrix proteins, and led to unexpected attenuation of senescence-associated secretory phenotype (SASP). The latter was exemplified by decreased secretion of pro-inflammatory cytokine IL-6 which was also confirmed after treatment with an inhibitor of PRDX6 iPLA2 activity, MJ33. In conclusion, by combining different methodological approaches we discovered a novel role of PRDX6 in senescent cell viability and SASP development. Our results suggest PRDX6 could have a potential as a drug target for senolytic or senomodulatory therapy.

Keywords:  Cellular senescence; Interleukin 6; Peroxiredoxin 6; Redox proteomics; SILAC-iodoTMT; Senescence-associated secretory phenotype

DOI:  https://doi.org/10.1016/j.redox.2021.102212