bims-midmar 2021-09-12 papers

bims-midmar

Biomed News

on Mitochondrial DNA maintenance and replication

Issue of 2021‒09‒12
twenty-five papers selected by
Flavia Söllner
Ludwig-Maximilians University

Mitochondrial DNA: Unraveling the "other" genome.
Trapped topoisomerase-DNA covalent complexes in the mitochondria and their role in human diseases.
RNA Granules in the Mitochondria and Their Organization under Mitochondrial Stresses.
Mitochondrial RNA, a new trigger of the innate immune system.
A rapid method to visualize human mitochondrial DNA replication through rotary shadowing and transmission electron microscopy.
A linear five-ring pyrrole-imidazole polyamide-triphenylphosphonium conjugate targeting a mitochondrial DNA mutation efficiently induces apoptosis of HeLa cybrid cells carrying the mutation.
ER-SURF: Riding the Endoplasmic Reticulum Surface to Mitochondria.
Internal validation and improvement of mitochondrial genome sequencing using the Precision ID mtDNA Whole Genome Panel.
A novel integrated approach to estimate the mitochondrial content of neuronal cells and brain tissues.
CRISPR/Cas9-Based Split Fluorescent Protein Tagging.
Mitochondrial DNA Copy Number Variations in Gastrointestinal Tract Cancers: Potential Players.
The Role of Mitochondrial DNA Variation in Drug Response: A Systematic Review.
Oligodendrocytes enhance axonal energy metabolism by deacetylation of mitochondrial proteins through transcellular delivery of SIRT2.
Visualizing and Quantifying Endonuclease-Based Site-Specific DNA Damage.
Mitochondria-Targeted Nanomedicine for Enhanced Efficacy of Cancer Therapy.
The Role of Mitochondrial Genes in Neurodegenerative Disorders.
Modelling SARS-CoV-2 CNS infection.
Structural basis of RNA processing by human mitochondrial RNase P.
Seminal fluid and sperm diluent affect sperm metabolism in an insect: Evidence from NAD(P)H and flavin adenine dinucleotide autofluorescence lifetime imaging.
Clinical Utility of Rapid Exome Sequencing Combined With Mitochondrial DNA Sequencing in Critically Ill Pediatric Patients With Suspected Genetic Disorders.
Low peripheral mitochondrial DNA copy number during manic episodes of bipolar disorders is associated with disease severity and inflammation.
Digging into the lesser-known aspects of CRISPR biology.
Mitokyne: A Ratiometric Raman Probe for Mitochondrial pH.
Genomic landscape of mitochondrial DNA insertions in 23 bat genomes: characteristics, loci, phylogeny, and polymorphism.
Two-photon fluorescent turn-on probes for highly efficient detection and profiling of thiols in live cells and tissues.

J Am Assoc Nurse Pract. 2021 Sep 01. 33(9): 673-675

Mitochondrial DNA: Unraveling the "other" genome.

Beth Heuer.

ABSTRACT: The mitochondrial genome, which contains all of the hereditary information within human mitochondria, consists of 16,569 base pairs of double-stranded DNA that encode 37 genes. Pathogenic mutations of mitochondrial DNA (mtDNA) cause dysfunction of the respiratory chain and the process of oxidative phosphorylation (OXPHOS), leading to impaired adenosine triphosphate synthesis. Nuclear DNA (nDNA) mutations can affect structural subunits or assembly factors of one of the five OXPHOS complexes. Mitochondrial diseases are a heterogeneous group of disorders, ranging from mtDNA single-point mutations and large-scale deletions to mitochondrial depletion syndromes, resulting from nDNA pathogenic mutations. Manifestations of mitochondrial disease are multisystemic, and organs with substantial energy requirements are most typically affected. Mitochondrial disorders are progressive in nature, and prognosis is dependent on the organs involved and the rate and severity of disease progression. A multidisciplinary team approach is needed to monitor and manage disease sequelae.

DOI: https://doi.org/10.1097/JXX.0000000000000646
Mitochondrion. 2021 Sep 06. pii: S1567-7249(21)00118-5. [Epub ahead of print]

Trapped topoisomerase-DNA covalent complexes in the mitochondria and their role in human diseases.

Benu Brata Das, Arijit Ghosh, Sangheeta Bhattacharjee, Arpan Bhattacharyya.

  Topoisomerases regulate DNA topology, organization of the intracellular DNA, the transmission of genetic materials, and gene expressions. Other than the nuclear genome, mitochondria also harbor the small, circular DNA (mtDNA) that encodes a critical subset of proteins for the production of cellular ATP; however, mitochondria are solely dependent on the nucleus for all the mitochondrial proteins necessary for mtDNA replication, repair, and maintenance. Mitochondrial genome compiles topological stress from bidirectional transcription and replication, therefore imports four nuclear encoded topoisomerases (Top1mt, Top2α, Top2β, and Top3α) in the mitochondria to relax mtDNA supercoiling generated during these processes. Trapping of topoisomerase on DNA results in the formation of protein-linked DNA adducts (PDAs), which are widely exploited by topoisomerase-targeting anticancer drugs. Intriguingly mtDNA is potentially exposed to DNA damage that has been attributed to a variety of human diseases, including neurodegeneration, cancer, and premature aging. In this review, we focus on the role of different topoisomerases in the mitochondria and our current understanding of the mitochondrial DNA damage through trapped protein-DNA complexes, and the progress in the molecular mechanisms of the repair for trapped topoisomerase covalent complexes (Topcc). Finally, we have discussed how the pathological DNA lesions that cause mtDNA damage,trigger mitochondrial fission and mitophagy, which serve as quality control events for clearing damaged mtDNA.

Keywords:  DNA repair; Mitochondria; TDP1; TDP2; TFAM; Topoisomerase 1; Topoisomerase II; mitochondrial DNA; neurological diseases

DOI:  https://doi.org/10.1016/j.mito.2021.08.017
Int J Mol Sci. 2021 Sep 01. pii: 9502. [Epub ahead of print]22(17):

RNA Granules in the Mitochondria and Their Organization under Mitochondrial Stresses.

Vanessa Joanne Xavier, Jean-Claude Martinou.

  The human mitochondrial genome (mtDNA) regulates its transcription products in specialised and distinct ways as compared to nuclear transcription. Thanks to its mtDNA mitochondria possess their own set of tRNAs, rRNAs and mRNAs that encode a subset of the protein subunits of the electron transport chain complexes. The RNA regulation within mitochondria is organised within specialised, membraneless, compartments of RNA-protein complexes, called the Mitochondrial RNA Granules (MRGs). MRGs were first identified to contain nascent mRNA, complexed with many proteins involved in RNA processing and maturation and ribosome assembly. Most recently, double-stranded RNA (dsRNA) species, a hybrid of the two complementary mRNA strands, were found to form granules in the matrix of mitochondria. These RNA granules are therefore components of the mitochondrial post-transcriptional pathway and as such play an essential role in mitochondrial gene expression. Mitochondrial dysfunctions in the form of, for example, RNA processing or RNA quality control defects, or inhibition of mitochondrial fission, can cause the loss or the aberrant accumulation of these RNA granules. These findings underline the important link between mitochondrial maintenance and the efficient expression of its genome.

Keywords:  RNA degradation; RNA processing; degradosome; dsRNA; liquid–liquid phase separation (LLPS); mitochondrial RNA granules (MRGs); mitochondrial gene expression; nucleoids

DOI:  https://doi.org/10.3390/ijms22179502
Wiley Interdiscip Rev RNA. 2021 Sep 08. e1690

Mitochondrial RNA, a new trigger of the innate immune system.

Joanna Grochowska, Jolanta Czerwinska, Lukasz S Borowski, Roman J Szczesny.

  Mitochondria play a pivotal role in numerous cellular processes. One of them is regulation of the innate immune pathway. In this instance, mitochondria function in two different aspects of regulatory mechanisms. First, mitochondria are part of the antiviral signaling cascade that is triggered in the cytoplasm and transmitted to effector proteins through mitochondria-localized proteins. Second, mitochondria can become an endogenous source of innate immune stimuli. Under some pathophysiological conditions, mitochondria release to the cytoplasm immunogenic factors, such as mitochondrial nucleic acids. Here, we focus on immunogenic mitochondrial double-stranded RNA (mt-dsRNA) and its origin and metabolism. We discuss factors that are responsible for regulating mt-dsRNA and its escape from mitochondria, emphasizing the contribution of polynucleotide phosphorylase (PNPase, PNPT1). Finally, we review current knowledge of the role of PNPase in human health and disease. This article is categorized under: RNA in Disease and Development > RNA in Disease.

Keywords:  innate immunity; mitochondrial RNA decay and surveillance; mitochondrial dsRNA

DOI:  https://doi.org/10.1002/wrna.1690
Nucleic Acids Res. 2021 Sep 09. pii: gkab770. [Epub ahead of print]

A rapid method to visualize human mitochondrial DNA replication through rotary shadowing and transmission electron microscopy.

Martin Kosar, Daniele Piccini, Marco Foiani, Michele Giannattasio.

We report a rapid experimental procedure based on high-density in vivo psoralen inter-strand DNA cross-linking coupled to spreading of naked purified DNA, positive staining, low-angle rotary shadowing, and transmission electron microscopy (TEM) that allows quick visualization of the dynamic of heavy strand (HS) and light strand (LS) human mitochondrial DNA replication. Replication maps built on linearized mitochondrial genomes and optimized rotary shadowing conditions enable clear visualization of the progression of the mitochondrial DNA synthesis and visualization of replication intermediates carrying long single-strand DNA stretches. One variant of this technique, called denaturing spreading, allowed the inspection of the fine chromatin structure of the mitochondrial genome and was applied to visualize the in vivo three-strand DNA structure of the human mitochondrial D-loop intermediate with unprecedented clarity.

DOI: https://doi.org/10.1093/nar/gkab770
Biochem Biophys Res Commun. 2021 Aug 31. pii: S0006-291X(21)01270-5. [Epub ahead of print]576 93-99

A linear five-ring pyrrole-imidazole polyamide-triphenylphosphonium conjugate targeting a mitochondrial DNA mutation efficiently induces apoptosis of HeLa cybrid cells carrying the mutation.

Nobuko Koshikawa, Yuki Kida, Nanami Yasui, Yoshinao Shinozaki, Kohei Tsuji, Takayoshi Watanabe, Jason Lin, Seigi Yamamoto, Keizo Takenaga, Hiroki Nagase.

  Somatic mutations in mitochondrial DNA may provide a new avenue for cancer therapy due to their associations to a number of cancers and a tendency of homoplasmicity. In consideration of mitochondrial features and its relatively small genome size, a nucleotide-based targeting approach is a considerably more promising option. To explore the efficacy of short linear N-methylpyrrole-N-methylimidazole polyamide (PI polyamide), we synthesized a five-ring short PI polyamide that provided sequence-specific homing for the A3243G mitochondrial mutation upon conjugation with triphenylphosphonium cation (TPP). This PI polyamide-TPP was able to induce cytotoxicity in HeLamtA3243G cybrid cells, while preserving preferential binding for oligonucleotides containing the A3243G motif from melting temperature assays. The PI polyamide-TPP also localized in the mitochondria in HeLamtA3243G cells and induced mitochondrial reactive oxygen species production, mitophagy and apoptosis in a mutation-specific fashion compared to the wild-type HeLamtHeLa cybrids; normal human dermal fibroblasts were also relatively unaffected to suggest discriminating selectivity for the mutant mitochondria, offering a novel outlook for cancer therapy via mitochondrial homing of short linear PIP-TPPs.

Keywords:  Apoptosis; Mitochondrial DNA mutation; Mitophagy; Pyrrole-imidazole polyamide; Triphenylphosphonium; mtROS

DOI:  https://doi.org/10.1016/j.bbrc.2021.08.088
Int J Mol Sci. 2021 Sep 06. pii: 9655. [Epub ahead of print]22(17):

ER-SURF: Riding the Endoplasmic Reticulum Surface to Mitochondria.

Christian Koch, Maya Schuldiner, Johannes M Herrmann.

  Most mitochondrial proteins are synthesized in the cytosol and targeted to the mitochondrial surface in a post-translational manner. The surface of the endoplasmic reticulum (ER) plays an active role in this targeting reaction. ER-associated chaperones interact with certain mitochondrial membrane protein precursors and transfer them onto receptor proteins of the mitochondrial surface in a process termed ER-SURF. ATP-driven proteins in the membranes of mitochondria (Msp1, ATAD1) and the ER (Spf1, P5A-ATPase) serve as extractors for the removal of mislocalized proteins. If the re-routing to mitochondria fails, precursors can be degraded by ER or mitochondria-associated degradation (ERAD or MAD respectively) in a proteasome-mediated reaction. This review summarizes the current knowledge about the cooperation of the ER and mitochondria in the targeting and quality control of mitochondrial precursor proteins.

Keywords:  ER-SURF; chaperones; contact sites; endoplasmic reticulum; membrane extraction; mitochondria; protein targeting

DOI:  https://doi.org/10.3390/ijms22179655
Int J Legal Med. 2021 Sep 07.

Internal validation and improvement of mitochondrial genome sequencing using the Precision ID mtDNA Whole Genome Panel.

Christian Faccinetto, Daniele Sabbatini, Patrizia Serventi, Martina Rigato, Cecilia Salvoro, Gianluca Casamassima, Gianluca Margiotta, Sara De Fanti, Stefania Sarno, Nicola Staiti, Donata Luiselli, Alberto Marino, Giovanni Vazza.

  With the recent advances in next-generation sequencing (NGS), mitochondrial whole-genome sequencing has begun to be applied to the field of the forensic biology as an alternative to the traditional Sanger-type sequencing (STS). However, experimental workflows, commercial solutions, and output data analysis must be strictly validated before being implemented into the forensic laboratory. In this study, we performed an internal validation for an NGS-based typing of the entire mitochondrial genome using the Precision ID mtDNA Whole Genome Panel (Thermo Fisher Scientific) on the Ion S5 sequencer (Thermo Fisher Scientific). Concordance, repeatability, reproducibility, sensitivity, and heteroplasmy detection analyses were assessed using the 2800 M and 9947A standard control DNA as well as typical casework specimens, and results were compared with conventional Sanger sequencing and another NGS sequencer in a different laboratory. We discuss the strengths and limitations of this approach, highlighting some issues regarding noise thresholds and heteroplasmy detection, and suggesting solutions to mitigate these effects and improve overall data interpretation. Results confirmed that the Precision ID Whole mtDNA Genome Panel is highly reproducible and sensitive, yielding useful full mitochondrial DNA sequences also from challenging DNA specimens, thus providing further support for its use in forensic practice.

Keywords:  Forensic genetics; Internal validation; Ion Torrent; Massive parallel sequencing; NGS; Whole mitochondrial genome

DOI:  https://doi.org/10.1007/s00414-021-02686-w
J Neurosci Methods. 2021 Sep 02. pii: S0165-0270(21)00286-7. [Epub ahead of print] 109351

A novel integrated approach to estimate the mitochondrial content of neuronal cells and brain tissues.

Sara Spadini, Gabriella Racchetti, Alice Adiletta, Jacopo Lamanna, Andrea Stefano Moro, Mattia Ferro, Vincenzo Zimarino, Antonio Malgaroli.

  BACKGROUND: Mitochondria and their dynamics fuel most cellular processes both in physiological and pathological conditions. In the central nervous system, mitochondria sustain synaptic transmission and plasticity via multiple mechanisms which include their redistribution and/or expansion to higher energy demanding sites, sustaining activity changes and promoting morphological circuit adaptations.NEW METHOD: To be able to evaluate changes in mitochondrial number and protein phenotype, we propose a novel methodological approach where the simultaneous analysis of both mitochondrial DNA and protein content is performed on each individual microsample, avoiding non-homogeneous loss of material.
RESULTS: We validated this method on neuronal-like cells and tissue samples and obtained estimates for the mitochondrial/genomic DNA ratio as well as for the abundance of protein counterparts. When the mitochondrial content per cell was evaluated in different brain areas, our results matched the known regional variation in aerobic-anaerobic metabolism. When long-term potentiation (LTP) was induced on hippocampal neurons, we detected increases in the abundance of mitochondria that correlated with the degree of synaptic enhancement.
CONCLUSIONS: Our approach can be effectively used to study the mitochondrial content andits changes in different brain cells and tissues.

Keywords:  Energy metabolism; LTP; Mitochondria; Neuronal metabolism; Synaptic plasticity; Synaptic transmission

DOI:  https://doi.org/10.1016/j.jneumeth.2021.109351
Zebrafish. 2021 Sep 07.

CRISPR/Cas9-Based Split Fluorescent Protein Tagging.

Gokul Kesavan, Anja Machate, Michael Brand.

  Genetically encoded fluorescent tags such as green fluorescent protein fused to protein have revolutionized cell biology as they permit high-resolution protein imaging in live systems. Split fluorescent proteins, with a small fragment of 16 amino acids, can be inserted in the coding sequence to label proteins. We demonstrate successful integration of two bright and fast maturing split fluorescent proteins, mNeon green and sfCherry2, in zebrafish, and show that they are suitable for live imaging, including time-lapse series, and that they have a high signal-to-noise ratio. Furthermore, we show that CRISPR/Cas9 can be used to generate fluorescently tagged proteins in vivo.

Keywords:  CRISPR/Cas9; genetically encoded fluorescent protein tags; knock-in; split fluorescent protein

DOI:  https://doi.org/10.1089/zeb.2021.0031
J Gastrointest Cancer. 2021 Sep 06.

Mitochondrial DNA Copy Number Variations in Gastrointestinal Tract Cancers: Potential Players.

Mehdi Alikhani, Eliette Touati, Morteza Karimipoor, Massoud Vosough, Marjan Mohammadi.

  Alterations of mitochondria have been linked to several cancers. Also, the mitochondrial DNA copy number (mtDNA-CN) is altered in various cancers, including gastrointestinal tract (GIT) cancers, and several research groups have investigated its potential as a cancer biomarker. However, the exact causes of mtDNA-CN variations are not yet revealed. This review discussed the conceivable players in this scheme, including reactive oxygen species (ROS), mtDNA genetic variations, DNA methylation, telomere length, autophagy, immune system activation, aging, and infections, and discussed their possible impact in the initiation and progression of cancer. By further exploring such mechanisms, mtDNA-CN variations may be effectively utilized as cancer biomarkers and provide grounds for developing novel cancer therapeutic agents.

Keywords:  Biomarker; Cancer; Causes; Gastrointestinal tract cancers; mtDNA copy number

DOI:  https://doi.org/10.1007/s12029-021-00707-w
Front Genet. 2021 ;12 698825

The Role of Mitochondrial DNA Variation in Drug Response: A Systematic Review.

Samantha W Jones, Amy L Ball, Amy E Chadwick, Ana Alfirevic.

  Background: The triad of drug efficacy, toxicity and resistance underpins the risk-benefit balance of all therapeutics. The application of pharmacogenomics has the potential to improve the risk-benefit balance of a given therapeutic via the stratification of patient populations based on DNA variants. A growth in the understanding of the particulars of the mitochondrial genome, alongside the availability of techniques for its interrogation has resulted in a growing body of literature examining the impact of mitochondrial DNA (mtDNA) variation upon drug response. Objective: To critically evaluate and summarize the available literature, across a defined period, in a systematic fashion in order to map out the current landscape of the subject area and identify how the field may continue to advance. Methods: A systematic review of the literature published between January 2009 and December 2020 was conducted using the PubMed database with the following key inclusion criteria: reference to specific mtDNA polymorphisms or haplogroups, a core objective to examine associations between mtDNA variants and drug response, and research performed using human subjects or human in vitro models. Results: Review of the literature identified 24 articles reporting an investigation of the association between mtDNA variant(s) and drug efficacy, toxicity or resistance that met the key inclusion criteria. This included 10 articles examining mtDNA variations associated with antiretroviral therapy response, 4 articles examining mtDNA variants associated with anticancer agent response and 4 articles examining mtDNA variants associated with antimicrobial agent response. The remaining articles covered a wide breadth of medications and were therefore grouped together and referred to as "other." Conclusions: Investigation of the impact of mtDNA variation upon drug response has been sporadic to-date. Collective assessment of the associations identified in the articles was inconclusive due to heterogeneous methods and outcomes, limited racial/ethnic groups, lack of replication and inadequate statistical power. There remains a high degree of idiosyncrasy in drug response and this area has the potential to explain variation in drug response in a clinical setting, therefore further research is likely to be of clinical benefit.

Keywords:  drug; efficacy; haplogroup; mitochondrial DNA; resistance; response; toxicity

DOI:  https://doi.org/10.3389/fgene.2021.698825
Neuron. 2021 Sep 01. pii: S0896-6273(21)00608-5. [Epub ahead of print]

Oligodendrocytes enhance axonal energy metabolism by deacetylation of mitochondrial proteins through transcellular delivery of SIRT2.

Kelly A Chamberlain, Ning Huang, Yuxiang Xie, Francesca LiCausi, Sunan Li, Yan Li, Zu-Hang Sheng.

  Neurons require mechanisms to maintain ATP homeostasis in axons, which are highly vulnerable to bioenergetic failure. Here, we elucidate a transcellular signaling mechanism by which oligodendrocytes support axonal energy metabolism via transcellular delivery of NAD-dependent deacetylase SIRT2. SIRT2 is undetectable in neurons but enriched in oligodendrocytes and released within exosomes. By deleting sirt2, knocking down SIRT2, or blocking exosome release, we demonstrate that transcellular delivery of SIRT2 is critical for axonal energy enhancement. Mass spectrometry and acetylation analyses indicate that neurons treated with oligodendrocyte-conditioned media from WT, but not sirt2-knockout, mice exhibit strong deacetylation of mitochondrial adenine nucleotide translocases 1 and 2 (ANT1/2). In vivo delivery of SIRT2-filled exosomes into myelinated axons rescues mitochondrial integrity in sirt2-knockout mouse spinal cords. Thus, our study reveals an oligodendrocyte-to-axon delivery of SIRT2, which enhances ATP production by deacetylating mitochondrial proteins, providing a target for boosting axonal bioenergetic metabolism in neurological disorders.

Keywords:  acetylation; adenine nucleotide translocases 1 and 2; axonal ATP; axonal energetics; axonal mitochondria; energy metabolism; exosome; myelin; oligodendrocyte; sirtuin 2

DOI:  https://doi.org/10.1016/j.neuron.2021.08.011
J Vis Exp. 2021 Aug 21.

Visualizing and Quantifying Endonuclease-Based Site-Specific DNA Damage.

Vasiliki Pantazi, Ivett Berzsenyi, Barbara N Borsos, Tibor Pankotai.

Cells are continuously exposed to various DNA damaging agents, inducing different cellular responses. Applying biochemical and genetic approaches is essential in revealing cellular events associated with the recruitment and assembly of DNA repair complexes at the site of DNA damage. In the last few years, several powerful tools have been developed to induce site-specific DNA damage. Moreover, novel seminal techniques allow us to study these processes at the single-cell resolution level using both fixed and living cells. Although these techniques have been used to study various biological processes, herein we present the most widely used protocols in the field of DNA repair, Fluorescence Immunostaining (IF) and Chromatin Immunoprecipitation (ChIP), which in combination with endonuclease-based site-specific DNA damage make it possible to visualize and quantify the genomic occupancy of DNA repair factors in a directed and regulated fashion, respectively. These techniques provide powerful tools for the researchers to identify novel proteins bound to the damaged genomic locus as well as their post-translational modifications necessary for their fine-tune regulation during DNA repair.

DOI: https://doi.org/10.3791/62175
Front Bioeng Biotechnol. 2021 ;9 720508

Mitochondria-Targeted Nanomedicine for Enhanced Efficacy of Cancer Therapy.

Yan Gao, Haibei Tong, Jialiang Li, Jiachen Li, Di Huang, Jisen Shi, Bing Xia.

  Nanomedicines have been designed and developed to deliver anticancer drugs or exert anticancer therapy more selectively to tumor sites. Recent investigations have gone beyond delivering drugs to tumor tissues or cells, but to intracellular compartments for amplifying therapy efficacy. Mitochondria are attractive targets for cancer treatment due to their important functions for cells and close relationships to tumor occurrence and metastasis. Accordingly, multifunctional nanoplatforms have been constructed for cancer therapy with the modification of a variety of mitochondriotropic ligands, to trigger the mitochondria-mediated apoptosis of tumor cells. On this basis, various cancer therapeutic modalities based on mitochondria-targeted nanomedicines are developed by strategies of damaging mitochondria DNA (mtDNA), increasing reactive oxygen species (ROS), disturbing respiratory chain and redox balance. Herein, in this review, we highlight mitochondria-targeted cancer therapies enabled by nanoplatforms including chemotherapy, photothermal therapy (PTT), photodynamic therapy (PDT), chemodynamic therapy (CDT), sonodynamic therapy (SDT), radiodynamic therapy (RDT) and combined immunotherapy, and discussed the ongoing challenges.

Keywords:  chemoteraphy; enhanced efficacy; immunotherapy; mitochondria targeting; nanomedicine; photothermal therapy (PTT)

DOI:  https://doi.org/10.3389/fbioe.2021.720508
Curr Neuropharmacol. 2021 Sep 08.

The Role of Mitochondrial Genes in Neurodegenerative Disorders.

Rajesh Kumar, Seetha Harila, Della Grace Thomas Parambi, S K Kanthlal, Md Atiar Rahman, Athanasios Alexiou, Gaber El-Saber Batiha, Bijo Mathew.

  Mitochondrial disorders are clinically heterogeneous, resulting from nuclear gene and mitochondrial mutations that disturb the mitochondrial functions and dynamics. There is a lack of evidence linking mtDNA mutations to neurodegenerative disorders, mainly due to the absence of noticeable neuropathological lesions in postmortem samples. This review describes various gene mutations in Alzheimer's disease, Parkinson's disease, Amyotrophic Lateral Sclerosis, Multiple Sclerosis, and Stroke. These abnormalities, including PINK1, Parkin, and SOD1 mutations, seem to reveal mitochondrial dysfunctions due to either mtDNA mutation or deletion, the mechanism of which remains unclear in depth.

Keywords:  Alzheimer's disease; Amyotrophic Lateral Sclerosis; Multiple Sclerosis; Neurodegeneration; Parkinson's disease; Stroke; mtDNA

DOI:  https://doi.org/10.2174/1570159X19666210908163839
Nat Cell Biol. 2021 Sep 08.

Modelling SARS-CoV-2 CNS infection.

Christine Weber.

DOI: https://doi.org/10.1038/s41556-021-00756-0
Nat Struct Mol Biol. 2021 Sep 06.

Structural basis of RNA processing by human mitochondrial RNase P.

Arjun Bhatta, Christian Dienemann, Patrick Cramer, Hauke S Hillen.

Human mitochondrial transcripts contain messenger and ribosomal RNAs flanked by transfer RNAs (tRNAs), which are excised by mitochondrial RNase (mtRNase) P and Z to liberate all RNA species. In contrast to nuclear or bacterial RNase P, mtRNase P is not a ribozyme but comprises three protein subunits that carry out RNA cleavage and methylation by unknown mechanisms. Here, we present the cryo-EM structure of human mtRNase P bound to precursor tRNA, which reveals a unique mechanism of substrate recognition and processing. Subunits TRMT10C and SDR5C1 form a subcomplex that binds conserved mitochondrial tRNA elements, including the anticodon loop, and positions the tRNA for methylation. The endonuclease PRORP is recruited and activated through interactions with its PPR and nuclease domains to ensure precise pre-tRNA cleavage. The structure provides the molecular basis for the first step of RNA processing in human mitochondria.

DOI: https://doi.org/10.1038/s41594-021-00637-y
Microsc Res Tech. 2021 Sep 05.

Seminal fluid and sperm diluent affect sperm metabolism in an insect: Evidence from NAD(P)H and flavin adenine dinucleotide autofluorescence lifetime imaging.

Christian Massino, Cornelia Wetzker, Ondřej Balvin, Tomáš Bartonicka, Jana Kremenova, Markéta Sasinkova, Oliver Otti, Klaus Reinhardt.

  Sperm metabolism is fundamental to sperm motility and male fertility. Its measurement is still in its infancy, and recommendations do not exist as to whether or how to standardize laboratory procedures. Here, using the sperm of an insect, the common bedbug, Cimex lectularius, we demonstrate that standardization of sperm metabolism is required with respect to the artificial sperm storage medium and a natural medium, the seminal fluid. We used fluorescence lifetime imaging microscopy (FLIM) in combination with time-correlated single-photon counting (TCSPC) to quantify sperm metabolism based on the fluorescent properties of autofluorescent coenzymes, NAD(P)H and flavin adenine dinucleotide. Autofluorescence lifetimes (decay times) differ for the free and protein-bound state of the co-enzymes, and their relative contributions to the lifetime signal serve to characterize the metabolic state of cells. We found that artificial storage medium and seminal fluid separately, and additively, affected sperm metabolism. In a medium containing sugars and amino acids (Grace's Insect medium), sperm showed increased glycolysis compared with a commonly used storage medium, phosphate-buffered saline (PBS). Adding seminal fluid to the sperm additionally increased oxidative phosphorylation, likely reflecting increased energy production of sperm during activation. Our study provides a protocol to measure sperm metabolism independently from motility, stresses that protocol standardizations for sperm measurements should be implemented and, for the first time, demonstrates that seminal fluid alters sperm metabolism. Equivalent protocol standardizations should be imposed on metabolic investigations of human sperm samples.

Keywords:  Cimex lectularius; FLIM; FLIRR; bedbug; metabolic mapping; multiphoton microscopy; spermatozoa

DOI:  https://doi.org/10.1002/jemt.23914
Front Genet. 2021 ;12 725259

Clinical Utility of Rapid Exome Sequencing Combined With Mitochondrial DNA Sequencing in Critically Ill Pediatric Patients With Suspected Genetic Disorders.

Xuejun Ouyang, Yu Zhang, Lijuan Zhang, Jixuan Luo, Ting Zhang, Hui Hu, Lin Liu, Lieqiang Zhong, Shaoying Zeng, Pingyi Xu, Zhenjiang Bai, Lee-Jun Wong, Jing Wang, Chunli Wang, Bin Wang, Victor Wei Zhang.

  Genetic disorders are a frequent cause of hospitalization, morbidity and mortality in pediatric patients, especially in the neonatal or pediatric intensive care unit (NICU/PICU). In recent years, rapid genome-wide sequencing (exome or whole genome sequencing) has been applied in the NICU/PICU. However, mtDNA sequencing is not routinely available in rapid genetic diagnosis programs, which may fail to diagnose mtDNA mutation-associated diseases. Herein, we explored the clinical utility of rapid exome sequencing combined with mtDNA sequencing in critically ill pediatric patients with suspected genetic disorders. Rapid clinical exome sequencing (CES) was performed as a first-tier test in 40 critically ill pediatric patients (aged from 6 days to 15 years) with suspected genetic conditions. Blood samples were also collected from the parents for trio analysis. Twenty-six patients presented with neuromuscular abnormalities or other systemic abnormalities, suggestive of suspected mitochondrial diseases or the necessity for a differential diagnosis of other diseases, underwent rapid mtDNA sequencing concurrently. A diagnosis was made in 18 patients (45.0%, 18/40); three cases with de novo autosomal dominant variants, ten cases with homozygous or compound heterozygous variants, three cases with hemizygous variants inherited from mother, three cases with heterozygous variants inherited from either parent, and one case with a mtDNA mutation. The 18 patients were diagnosed with metabolic (n = 7), immunodeficiency (n = 4), cardiovascular (n = 2), neuromuscular (n = 2) disorders, and others. Genetic testing reports were generated with a median time of 5 days (range, 3-9 days). Thirteen patients that were diagnosed had an available medical treatment and resulted in a positive outcome. We propose that rapid exome sequencing combined with mitochondrial DNA sequencing should be available to patients with suspected mitochondrial diseases or undefined clinical features necessary for making a differential diagnosis of other diseases.

Keywords:  genetic disorders; mitochondrial diseases; mtDNA sequencing; pediatric patients; rapid exome sequencing

DOI:  https://doi.org/10.3389/fgene.2021.725259
Brain Behav Immun. 2021 Sep 06. pii: S0889-1591(21)00542-0. [Epub ahead of print]

Low peripheral mitochondrial DNA copy number during manic episodes of bipolar disorders is associated with disease severity and inflammation.

Loïc Angrand, Wahid Boukouaci, Mohamed Lajnef, Jean-Romain Richard, Ana Andreazza, Chieng-Lien Wu, Jihène Bouassida, Ismail Rafik, Marianne Foiselle, Esma Mezouad, Soumia Naamoune, Leila Chami, Ons Mihoub, Sofiane Salah, Arij Benchaaben, Philippe Le Corvoisier, Caroline Barau, Bruno Costes, Robert Yolken, Guillemette Crepeaux, Marion Leboyer, Ryad Tamouza.

  Mitochondria (Mt) are intra-cellular components essential for cellular energy processes whose dysfunction may induce premature cellular senescence and/or inflammation, both observed in bipolar disorders (BD). We investigated mitochondrial DNA copy number (mtDNAcn) levels in patients with BD being in manic, depressive or euthymic phase and in healthy controls (HC) both characterized for the levels of blood-based inflammatory markers and stigma of pathogens. 312 patients with BD were compared to 180 HC. mtDNAcn were measured using a digital droplet PCR. Serum levels of 14 inflammatory molecules and 3 anti-infectious IgG stigma were respectively evaluated by electro-chemiluminescence, ELISA and dedicated immunoassays. The statistical analyses were performed using Spearman's correlation, Wilcoxon signed-rank and Kruskal-Wallis rank sum tests. P-values were adjusted for multiple testing with Benjamini-Hochberg method. We found low levels of mtDNAcn in BD patients as compared to HC (P=0.008) especially during manic episodes (P=0.0002). We also observed that low levels of mtDNAcn are negatively correlated with mood and psychotic scales (PANSS, YMRS and CGI) (adjusted P (Adj P) = 0.02, 0.003 and 0.05 respectively) and positively with the GAF severity scale (Adj P= 0.002). They were also correlated with high levels of both intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 (Adj P =0.003 and 0.001) along with a trend toward increased IL-2, IL-10 and B2M circulating levels (Adj P= 0.05). Here, we report correlations between marker of mitochondria functioning and both clinical scales and inflammatory markers in BD patients experiencing manic episodes. If replicated, these finding might allow to predict transition between disease phases and to design accurate therapeutic options.

Keywords:  Bipolar disorders (BD), Mania, Mitochondria (mt); Mitochondrial DNA copy number (mtDNAcnv); inflammation, intercellular adhesion molecule (ICAM)-1; vascular cell adhesion molecule (VCAM)-1

DOI:  https://doi.org/10.1016/j.bbi.2021.09.003
Int Microbiol. 2021 Sep 06.

Digging into the lesser-known aspects of CRISPR biology.

Noemí M Guzmán, Belén Esquerra-Ruvira, Francisco J M Mojica.

  A long time has passed since regularly interspaced DNA repeats were discovered in prokaryotes. Today, those enigmatic repetitive elements termed clustered regularly interspaced short palindromic repeats (CRISPR) are acknowledged as an emblematic part of multicomponent CRISPR-Cas (CRISPR associated) systems. These systems are involved in a variety of roles in bacteria and archaea, notably, that of conferring protection against transmissible genetic elements through an adaptive immune-like response. This review summarises the present knowledge on the diversity, molecular mechanisms and biology of CRISPR-Cas. We pay special attention to the most recent findings related to the determinants and consequences of CRISPR-Cas activity. Research on the basic features of these systems illustrates how instrumental the study of prokaryotes is for understanding biology in general, ultimately providing valuable tools for diverse fields and fuelling research beyond the mainstream.

Keywords:  Adaptive immunity; CRISPR; CRISPR regulation; Cas proteins; Non-canonical CRISPR roles; RNA-guided transposition

DOI:  https://doi.org/10.1007/s10123-021-00208-7
Anal Chem. 2021 Sep 10.

Mitokyne: A Ratiometric Raman Probe for Mitochondrial pH.

Liam T Wilson, William J Tipping, Corinna Wetherill, Zoë Henley, Karen Faulds, Duncan Graham, Simon P Mackay, Nicholas C O Tomkinson.

Mitochondrial pH (pHmito) is intimately related to mitochondrial function, and aberrant values for pHmito are linked to several disease states. We report the design, synthesis, and application of mitokyne 1-the first small molecule pHmito sensor for stimulated Raman scattering (SRS) microscopy. This ratiometric probe can determine subtle changes in pHmito in response to external stimuli and the inhibition of both the electron transport chain and ATP synthase with small molecule inhibitors. In addition, 1 was also used to monitor mitochondrial dynamics in a time-resolved manner with subcellular spatial resolution during mitophagy providing a powerful tool for dissecting the molecular and cell biology of this critical organelle.

DOI: https://doi.org/10.1021/acs.analchem.1c03075
Integr Zool. 2021 Sep 08.

Genomic landscape of mitochondrial DNA insertions in 23 bat genomes: characteristics, loci, phylogeny, and polymorphism.

Guojun Zhang, Deqi Geng, Qiulin Guo, Wei Liu, Shufen Li, Wujun Gao, Yongfei Wang, Min Zhang, Yilin Wang, Yanzhen Bu, Hongxing Niu.

  The transfer of mitochondrial DNA to the nuclear genome gives rise to the nuclear DNA sequences of mitochondrial origin (NUMTs), considered as a driving force in genome evolution. In this study, NUMTs in 23 bat genomes were investigated and compared systematically. The results showed that NUMTs existed in 22 genomes except for Noctilio leporinus, suggesting that mitochondrial fragment insertion in the nuclear genome was a common event in bat genomes. However, remarkable variations in NUMTs number, cumulative length, and proportion in the nuclear genome were discovered across bat species. Further orthologous NUMT loci analysis of the Phyllostomidae family indicated that the NUMTs insertion events in bat genomes were homoplasy-free. The NUMTs were mainly inserted into the intergenic regions, particularly, co-localized with repetitive sequences (especially transposable elements). However, several NUMTs were inserted into genes, some of which were in the exon region of functional genes. One NUMT in the genome of Pteropus alecto surprisingly matched with cDNA of ATP8B3 that provided evidence of NUMTs with coding function. Phylogenic analysis on NUMTs originating from COXI and COXII loci highlighted two clusters of Yinpterochiroptera and Yangochiroptera for Chiroptera. Seven NUMTs from Rhinolophus ferrumequinum were amplified, and the sequencing results confirmed the reliability of the NUMT analysis. One of them was polymorphic for the presence or absence of the NUMT insertion, and each genotype of NUMT loci showed a distinct regional distribution pattern. The information obtained in this study provides novel insights into the NUMT organization and features in bat genomes and establishes a basis for further studying of the evolution of bat species. This article is protected by copyright. All rights reserved.

Keywords:  NUMTs; bat; homoplasy-free; mitochondrial DNA (mtDNA); polymorphism

DOI:  https://doi.org/10.1111/1749-4877.12582
Biol Chem. 2021 Sep 09.

Two-photon fluorescent turn-on probes for highly efficient detection and profiling of thiols in live cells and tissues.

Congzhen Shen, Duoteng Zhang, Fang Xu, Yang Yang, Yi Tan, Qian Zhao, Lin Li, Ke Ding, Zhengqiu Li.

  Thiols are important units in amino acids such as cysteine and peptides like glutathione. Development of chemical sensors capable of precise detection of thiols is important in cancer diagnosis and therapy. We have developed novel two-photon fluorescent turn-on probes for selective detection of thiols. The probes displayed excellent sensitivity and low detection limits. The dual-purpose probes have been demonstrated to be suitable for simultaneous imaging and proteome profiling in live cells and tumor tissues. The unique turn-on design endows the probes with excellent selectivity toward thiols in vitro and in situ, and can be further developed to support a thiol-quantification assay.

Keywords:  cysteine residue; fluorescence probe; proteome profiling; thiol group; two-photon probes

DOI:  https://doi.org/10.1515/hsz-2021-0189