bims-nenemi 2023-03-05 papers

bims-nenemi

Biomed News

on Neuroinflammation, neurodegeneration and mitochondria

Issue of 2023–03–05
thirteen papers selected by
Marco Tigano, Thomas Jefferson University

ANKLE1 cleaves mitochondrial DNA and contributes to cancer risk by promoting apoptosis resistance and metabolic dysregulation.
Impaired mitophagy causes mitochondrial DNA leakage and STING activation in ultraviolet B-irradiated human keratinocytes HaCaT.
Mitochondria-localized cGAS suppresses ferroptosis to promote cancer progression.
Analysis of mitochondrial double-stranded RNAs in human cells.
TFAM deficiency in dendritic cells leads to mitochondrial dysfunction and enhanced antitumor immunity through cGAS-STING pathway.
PM2.5 induces mitochondrial dysfunction via AHR-mediated cyp1a1 overexpression during zebrafish heart development.
MERR APEX-seq protocol for profiling the subcellular nascent transcriptome in mammalian cells.
NF-κB activation enhances STING signaling by altering microtubule-mediated STING trafficking.
Protocol for mitochondrial isolation and sub-cellular localization assay for mitochondrial proteins.
Memory of stochastic single-cell apoptotic signaling promotes chemoresistance in neuroblastoma.
Lysosomal control of senescence and inflammation through cholesterol partitioning.
Glia Connect Inflammation and Neurodegeneration in Multiple Sclerosis.
Polynucleotide phosphorylase protects against renal tubular injury via blocking mt-dsRNA-PKR-eIF2α axis.

Commun Biol. 2023 Mar 01. 6(1): 231

ANKLE1 cleaves mitochondrial DNA and contributes to cancer risk by promoting apoptosis resistance and metabolic dysregulation.

Piotr Przanowski, Róża K Przanowska, Michael J Guertin.

Alleles within the chr19p13.1 locus are associated with increased risk of both ovarian and breast cancer and increased expression of the ANKLE1 gene. ANKLE1 is molecularly characterized as an endonuclease that efficiently cuts branched DNA and shuttles between the nucleus and cytoplasm. However, the role of ANKLE1 in mammalian development and homeostasis remains unknown. In normal development ANKLE1 expression is limited to the erythroblast lineage and we found that ANKLE1's role is to cleave the mitochondrial genome during erythropoiesis. We show that ectopic expression of ANKLE1 in breast epithelial-derived cells leads to genome instability and mitochondrial DNA (mtDNA) cleavage. mtDNA degradation then leads to mitophagy and causes a shift from oxidative phosphorylation to glycolysis (Warburg effect). Moreover, mtDNA degradation activates STAT1 and expression of epithelial-mesenchymal transition (EMT) genes. Reduction in mitochondrial content contributes to apoptosis resistance, which may allow precancerous cells to avoid apoptotic checkpoints and proliferate. These findings provide evidence that ANKLE1 is the causal cancer susceptibility gene in the chr19p13.1 locus and describe mechanisms by which higher ANKLE1 expression promotes cancer risk.

DOI: https://doi.org/10.1038/s42003-023-04611-w
Arch Biochem Biophys. 2023 Feb 25. pii: S0003-9861(23)00052-8. [Epub ahead of print]737 109553

Impaired mitophagy causes mitochondrial DNA leakage and STING activation in ultraviolet B-irradiated human keratinocytes HaCaT.

Can Li, Yuying Zhu, Weiwei Liu, Wendie Xiang, Sijun He, Toshihiko Hayashi, Kazunori Mizuno, Shunji Hattori, Hitomi Fujisaki, Takashi Ikejima.

  Ultraviolet B (UVB) irradiation causes skin damages. In this study, we focus on the involvement of mitochondrial disorders in UVB injury. Surprisingly, UVB irradiation increases the amounts of mitochondria in human immortalized keratinocytes HaCaT. However, further analysis shows that ATP levels decreased by UVB treatment in accordance with the collapse of mitochondrial membrane potential (MMP), suggesting an accumulation of dysfunctional mitochondria in UVB-irradiated HaCaT cells. Mitophagy, mainly mediated by PINK1 and parkin, is critical for the elimination of damaged mitochondria. Western blot results show that the levels of both PINK1 and parkin are decreased in UVB-irradiated cells, indicating the impairment of mitophagy. Silencing the expression of PINK1 or parkin by transfection of siRNA shows essentially the same damage to the cells as UVB irradiation does, including increased mitochondrial amount, decreased MMP and ATP production, and enhanced apoptosis, evidencing that repression of PINK1/parkin-mediated mitophagy plays a primary cause of UVB-caused cells damages. We previously found that HaCaT cells exposed to UVB showed activation of the cGAS-STING pathway and apoptosis. Here, silencing PINK1 or parkin also increases the protein levels of cGAS and STING, facilitates nuclear accumulation of NF-κB, and promotes the transcription of IFNβ, suggesting for the activation of STING pathway. Mitophagy impairment either by UVB-irradiation or by PINK1/parkin silencing initiates caspase-3-mediated apoptosis, as shown by the activation of caspase-3 and cleavage of PARP, as well as the increase of Hoechst-positive stained cells and Annexin V-positive cells. Further studies find that Bax-mediated permeabilization of mitochondrial membrane is critical for cell apoptosis, as well as the cytosolic leakage of mtDNA in UVB-treated cells, which results in cGAS-STING activation, and these processes are negatively-regulated by PINK1/parkin-mediated mitophagy. This study reveals the involvement of dysfunctional mitochondria due to impaired mitophagy in the damaging effect of UVB irradiation on HaCaT cells. Restoring the mitophagy has the potential to be developed as a new strategy to protect skin from UVB damages.

Keywords:  Apoptosis; Mitochondrial DNA; PINK1; Parkin; STING; UVB

DOI:  https://doi.org/10.1016/j.abb.2023.109553
Cell Res. 2023 Mar 02.

Mitochondria-localized cGAS suppresses ferroptosis to promote cancer progression.

Shiqiao Qiu, Xiuying Zhong, Xiang Meng, Shiting Li, Xiaoyu Qian, Hui Lu, Jin Cai, Yi Zhang, Mingjie Wang, Zijian Ye, Huafeng Zhang, Ping Gao.

A well-established role of cyclic GMP-AMP synthase (cGAS) is the recognition of cytosolic DNA, which is linked to the activation of host defense programs against pathogens via stimulator of interferon genes (STING)-dependent innate immune response. Recent advance has also revealed that cGAS may be involved in several noninfectious contexts by localizing to subcellular compartments other than the cytosol. However, the subcellular localization and function of cGAS in different biological conditions is unclear; in particular, its role in cancer progression remains poorly understood. Here we show that cGAS is localized to mitochondria and protects hepatocellular carcinoma cells from ferroptosis in vitro and in vivo. cGAS anchors to the outer mitochondrial membrane where it associates with dynamin-related protein 1 (DRP1) to facilitate its oligomerization. In the absence of cGAS or DRP1 oligomerization, mitochondrial ROS accumulation and ferroptosis increase, inhibiting tumor growth. Collectively, this previously unrecognized role for cGAS in orchestrating mitochondrial function and cancer progression suggests that cGAS interactions in mitochondria can serve as potential targets for new cancer interventions.

DOI: https://doi.org/10.1038/s41422-023-00788-1
STAR Protoc. 2023 Jan 18. pii: S2666-1667(22)00887-5. [Epub ahead of print]4(1): 102007

Analysis of mitochondrial double-stranded RNAs in human cells.

Sujin Kim, Jimin Yoon, Keonyong Lee, Yoosik Kim.

  Human mitochondrial genome is transcribed bidirectionally, generating long complementary RNAs that can form double-stranded RNAs (mt-dsRNAs). When released to the cytosol, these mt-dsRNAs can activate antiviral signaling. Here, we present a detailed protocol for the analysis of mt-dsRNA expression. The protocol provides three approaches that can complement one another in examining mt-dsRNAs. While the described protocol is optimized for human cells, this approach can be adapted for use in other animal cell lines and tissue samples. For complete details on the use and execution of this protocol, please refer to Kim et al. (2022).1.

Keywords:  Biotechnology and bioengineering; Cell biology; Immunology; Molecular biology; Protein biochemistry; Protein expression and purification

DOI:  https://doi.org/10.1016/j.xpro.2022.102007
J Immunother Cancer. 2023 Mar;pii: e005430. [Epub ahead of print]11(3):

TFAM deficiency in dendritic cells leads to mitochondrial dysfunction and enhanced antitumor immunity through cGAS-STING pathway.

Tianqi Lu, Ziqi Zhang, Zhenfei Bi, Tianxia Lan, Hao Zeng, Yu Liu, Fei Mo, Jingyun Yang, Siyuan Chen, Xuemei He, Weiqi Hong, Zhe Zhang, Ruyu Pi, Wenyan Ren, Xiaohe Tian, Yuquan Wei, Min Luo, Xiawei Wei.

BACKGROUND: Mitochondrial transcription factor A (TFAM) is a transcription factor that maintains mitochondrial DNA (mtDNA) stabilization and initiates mtDNA replication. However, little is known about the immune regulation function and TFAM expression in immune cells in the tumors.
METHODS: Mouse tumor models were applied to analyze the effect of TFAM deficiency in myeloid cell lineage on tumor progression and tumor microenvironment (TME) modification. In vitro, primary mouse bone marrow-derived dendritic cells (BMDCs) were used in the investigation of the altered function and the activated pathway. OVA was used as the model antigen to validate the activation of immune responses in vivo. STING inhibitors were used to confirm the STING activation provoked by Tfam deficient in DCs.
RESULTS: The deletion of TFAM in DCs led to mitochondrial dysfunction and mtDNA cytosolic leakage resulting in the cGAS-STING pathway activation in DCs, which contributed to the enhanced antigen presentation. The deletion of TFAM in DCs has interestingly reversed the immune suppressive TME and inhibited tumor growth and metastasis in tumor models.
CONCLUSIONS: We have revealed that TFAM knockout in DCs ameliorated immune-suppressive microenvironment in tumors through STING pathway. Our work suggests that specific TFAM knockout in DCs might be a compelling strategy for designing novel immunotherapy methods in the future.

DOI: https://doi.org/10.1136/jitc-2022-005430
Toxicology. 2023 Feb 24. pii: S0300-483X(23)00052-5. [Epub ahead of print]487 153466

PM2.5 induces mitochondrial dysfunction via AHR-mediated cyp1a1 overexpression during zebrafish heart development.

Jin Chen, Mingxuan Zhang, Hongmei Zou, Stanley Aniagu, Yan Jiang, Tao Chen.

  Accumulating evidence suggests an association between maternal PM2.5 exposure and congenital heart diseases, but the underlying mechanisms remain unclear. We previously reported that PM2.5 induces cardiac malformations in zebrafish embryos via the aryl hydrocarbon receptor (AHR) pathway, which mediates the generation of reactive oxygen species (ROS). Since mitochondria are not only the main source of ROS but also sensitive to oxidative damage, we hypothesize that mitochondria may play an important role in the cardiac developmental toxicity of PM2.5. In this study, we demonstrated that extractable organic matter (EOM) from PM2.5 caused mitochondrial dysfunction in the heart of zebrafish embryos, including increased mitochondrial ROS (mtROS) levels, mitochondrial permeability transition pore (mPTP) opening, mitochondrial membrane potential (MMP) collapse, reduced mitochondrial ATP levels, and decreased expression levels of the mRNAs encoding mitochondrial proteins, which were attenuated by either pharmacological or genetic inhibition of AHR. We further demonstrated that improving mitochondrial function by inhibiting mPTP opening with Cyclosporin A suppressed the EOM-induced intracellular ROS and mtROS generation, MMP collapse, intrinsic apoptosis, and heart defects. Moreover, the EOM-induced mPTP opening was counteracted by inhibiting mtROS with mitoquinone mesylate (MitoQ). Supplementation with MitoQ also attenuated the EOM-induced mitochondrial dysfunction, apoptosis and heart defects. Additionally, knockdown of cyp1a1 but not cyp1b1 attenuated the EOM-induced mtROS generation and heart defects. Taken together, this study indicates that PM2.5 triggers mtROS generation via AHR-mediated cyp1a1 overexpression, which then causes mPTP opening and mitochondrial dysfunction, leading to apoptosis and heart defects.

Keywords:  AHR; Heart development; Mitochondrial dysfunction; PM2.5; ROS; Zebrafish

DOI:  https://doi.org/10.1016/j.tox.2023.153466
STAR Protoc. 2023 Jan 24. pii: S2666-1667(23)00015-1. [Epub ahead of print]4(1): 102057

MERR APEX-seq protocol for profiling the subcellular nascent transcriptome in mammalian cells.

Ran Li, Peng Zou.

  Knowledge about the spatial organization of RNAs in eukaryotic cells is crucial for understanding their functions. Here, we present a detailed MERR APEX-seq protocol to achieve spatiotemporally resolved mapping of the subcellular transcriptome in cultured mammalian cells. This protocol provides detailed description of constructing cell lines stably expressing APEX2, immunofluorescence characterization, MERR APEX labeling, enrichment of biotinylated RNA, library construction and high-throughput sequencing, and MERR APEX-seq data analysis. For complete details on the use and execution of this protocol, please refer to Li et al. (2022).1.

Keywords:  Biotechnology and Bioengineering; Chemistry; Molecular Biology; Molecular/Chemical Probes; RNAseq; Sequence Analysis; Sequencing

DOI:  https://doi.org/10.1016/j.xpro.2023.102057
Cell Rep. 2023 Feb 28. pii: S2211-1247(23)00196-1. [Epub ahead of print]42(3): 112185

NF-κB activation enhances STING signaling by altering microtubule-mediated STING trafficking.

Lulu Zhang, Xubiao Wei, Zhimeng Wang, Peiyuan Liu, Yanfei Hou, Yifang Xu, Huili Su, Matthew D Koci, Hang Yin, Conggang Zhang.

  It is widely known that stimulator of interferon genes (STING) can trigger nuclear factor κB (NF-κB) signaling. However, whether and how the NF-κB pathway affects STING signaling remains largely unclear. Here, we report that Toll-like receptor (TLR)-, interleukin-1 receptor (IL-1R)-, tumor necrosis factor receptor (TNFR)-, growth factor receptor (GF-R)-, and protein kinase C (PKC)-mediated NF-κB signaling activation dramatically enhances STING-mediated immune responses. Mechanistically, we find that STING interacts with microtubules, which plays a crucial role in STING intracellular trafficking. We further uncover that activation of the canonical NF-κB pathway induces microtubule depolymerization, which inhibits STING trafficking to lysosomes for degradation. This leads to increased levels of activated STING that persist for a longer period of time. The synergy between NF-κB and STING triggers a cascade-amplified interferon response and robust host antiviral defense. In addition, we observe that several gain-of-function mutations of STING abolish the microtubule-STING interaction and cause abnormal STING trafficking and ligand-independent STING autoactivation. Collectively, our data demonstrate that NF-κB activation enhances STING signaling by regulating microtubule-mediated STING trafficking.

Keywords:  CP: Immunology; NF-κB signaling pathways; SAVI; STING degradation; Toll-like receptors; innate immunity; microtubule depolymerization

DOI:  https://doi.org/10.1016/j.celrep.2023.112185
STAR Protoc. 2023 Feb 03. pii: S2666-1667(23)00046-1. [Epub ahead of print]4(1): 102088

Protocol for mitochondrial isolation and sub-cellular localization assay for mitochondrial proteins.

Danyi Zhou, Sheng Zhong, Xinyu Han, Dandan Liu, Hezhi Fang, Ya Wang.

  Here, we provide a protocol to isolate mitochondria from cultured cells and extract differently located mitochondrial proteins. We detail steps to separate both integral and peripheral membrane proteins from soluble proteins using sonication. We describe the separation of integral membrane proteins from the peripheral membrane and soluble proteins using sodium carbonate extraction. Furthermore, we detail the use of proteinase K and Triton X-100 to distinguish outer membrane proteins from mitochondrial proteins.

Keywords:  Cell Membrane; Cell culture; Cell separation/fractionation; Protein Biochemistry

DOI:  https://doi.org/10.1016/j.xpro.2023.102088
Sci Adv. 2023 Mar 03. 9(9): eabp8314

Memory of stochastic single-cell apoptotic signaling promotes chemoresistance in neuroblastoma.

Jordan F Hastings, Sharissa L Latham, Alvin Kamili, Madeleine S Wheatley, Jeremy Z R Han, Marie Wong-Erasmus, Monica Phimmachanh, Max Nobis, Chiara Pantarelli, Antonia L Cadell, Yolande E I O'Donnell, King Ho Leong, Sophie Lynn, Fan-Suo Geng, Lujing Cui, Sabrina Yan, Joanna Achinger-Kawecka, Clare Stirzaker, Murray D Norris, Michelle Haber, Toby N Trahair, Frank Speleman, Katleen De Preter, Mark J Cowley, Ozren Bogdanovic, Paul Timpson, Thomas R Cox, Walter Kolch, Jamie I Fletcher, Dirk Fey, David R Croucher.

Gene expression noise is known to promote stochastic drug resistance through the elevated expression of individual genes in rare cancer cells. However, we now demonstrate that chemoresistant neuroblastoma cells emerge at a much higher frequency when the influence of noise is integrated across multiple components of an apoptotic signaling network. Using a JNK activity biosensor with longitudinal high-content and in vivo intravital imaging, we identify a population of stochastic, JNK-impaired, chemoresistant cells that exist because of noise within this signaling network. Furthermore, we reveal that the memory of this initially random state is retained following chemotherapy treatment across a series of in vitro, in vivo, and patient models. Using matched PDX models established at diagnosis and relapse from individual patients, we show that HDAC inhibitor priming cannot erase the memory of this resistant state within relapsed neuroblastomas but improves response in the first-line setting by restoring drug-induced JNK activity within the chemoresistant population of treatment-naïve tumors.

DOI: https://doi.org/10.1126/sciadv.abp8314
Nat Metab. 2023 Mar 02.

Lysosomal control of senescence and inflammation through cholesterol partitioning.

Kyeonghwan Roh, Jeonghwan Noh, Yeonju Kim, Yeji Jang, Jaejin Kim, Haebeen Choi, Yeonghyeon Lee, Moongi Ji, Donghyun Kang, Mi-Sung Kim, Man-Jeong Paik, Jongkyeong Chung, Jin-Hong Kim, Chanhee Kang.

Whereas cholesterol is vital for cell growth, proliferation, and remodeling, dysregulation of cholesterol metabolism is associated with multiple age-related pathologies. Here we show that senescent cells accumulate cholesterol in lysosomes to maintain the senescence-associated secretory phenotype (SASP). We find that induction of cellular senescence by diverse triggers enhances cellular cholesterol metabolism. Senescence is associated with the upregulation of the cholesterol exporter ABCA1, which is rerouted to the lysosome, where it moonlights as a cholesterol importer. Lysosomal cholesterol accumulation results in the formation of cholesterol-rich microdomains on the lysosomal limiting membrane enriched with the mammalian target of rapamycin complex 1 (mTORC1) scaffolding complex, thereby sustaining mTORC1 activity to support the SASP. We further show that pharmacological modulation of lysosomal cholesterol partitioning alters senescence-associated inflammation and in vivo senescence during osteoarthritis progression in male mice. Our study reveals a potential unifying theme for the role of cholesterol in the aging process through the regulation of senescence-associated inflammation.

DOI: https://doi.org/10.1038/s42255-023-00747-5
Neurosci Bull. 2023 Feb 28.

Glia Connect Inflammation and Neurodegeneration in Multiple Sclerosis.

Ye Sun, Haojun Yu, Yangtai Guan.

  Multiple sclerosis (MS) is regarded as a chronic inflammatory disease that leads to demyelination and eventually to neurodegeneration. Activation of innate immune cells and other inflammatory cells in the brain and spinal cord of people with MS has been well described. However, with the innovation of technology in glial cell research, we have a deep understanding of the mechanisms of glial cells connecting inflammation and neurodegeneration in MS. In this review, we focus on the role of glial cells, including microglia, astrocytes, and oligodendrocytes, in the pathogenesis of MS. We mainly focus on the connection between glial cells and immune cells in the process of axonal damage and demyelinating neuron loss.

Keywords:  Glial cells; Inflammation; Multiple sclerosis; Neurodegeneration

DOI:  https://doi.org/10.1007/s12264-023-01034-9
Nat Commun. 2023 Mar 03. 14(1): 1223

Polynucleotide phosphorylase protects against renal tubular injury via blocking mt-dsRNA-PKR-eIF2α axis.

Yujie Zhu, Mingchao Zhang, Weiran Wang, Shuang Qu, Minghui Liu, Weiwei Rong, Wenwen Yang, Hongwei Liang, Caihong Zeng, Xiaodong Zhu, Limin Li, Zhihong Liu, Ke Zen.

Renal tubular atrophy is a hallmark of chronic kidney disease. The cause of tubular atrophy, however, remains elusive. Here we report that reduction of renal tubular cell polynucleotide phosphorylase (PNPT1) causes renal tubular translation arrest and atrophy. Analysis of tubular atrophic tissues from renal dysfunction patients and male mice with ischemia-reperfusion injuries (IRI) or unilateral ureteral obstruction (UUO) treatment shows that renal tubular PNPT1 is markedly downregulated under atrophic conditions. PNPT1 reduction leads to leakage of mitochondrial double-stranded RNA (mt-dsRNA) into the cytoplasm where it activates protein kinase R (PKR), followed by phosphorylation of eukaryotic initiation factor 2α (eIF2α) and protein translational termination. Increasing renal PNPT1 expression or inhibiting PKR activity largely rescues IRI- or UUO-induced mouse renal tubular injury. Moreover, tubular-specific PNPT1-knockout mice display Fanconi syndrome-like phenotypes with impaired reabsorption and significant renal tubular injury. Our results reveal that PNPT1 protects renal tubules by blocking the mt-dsRNA-PKR-eIF2α axis.

DOI: https://doi.org/10.1038/s41467-023-36664-0