Biology subjects

Matondo, M.

Publications and source records attributed to Matondo, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Biogenesis of DNA-carrying extracellular vesicles by the dominant human gut methanogenic archaeon

Extracellular vesicles (EVs) are membrane-bound particles secreted by cells from all domains of life and implicated in a variety of important processes, from intercellular communication to pathogenesis. Here, we characterize EVs produced by the dominant human gut methanogen, Methanobrevibacter smithii, which, unlike most archaea, contains a peptidoglycan cell wall. Using quantitative proteomics, we show that M. smithii EVs are enriched in various proteins responsible for chromatin structure, including histones, and DNA repair. Consistently, the M. smithii EVs carry DNA, with fragments covering the entire cellular chromosome. Notably, the EVs are strongly enriched in extrachromosomal circular DNA (eccDNA) molecules which originate from excision of a 2.9-kb chromosomal fragment and a proviral genome. The eccDNA encodes two of the key methanogenesis enzymes and could boost their expression inside the cells through the gene dosage effect. Furthermore, four of the top ten most abundant EV proteins are implicated in methanogenesis. Cryo-electron tomography (Cryo-ET) suggests that EVs are formed by budding from the cell membrane and are trapped under the cell wall prior to liberation through local disruptions in the cell wall. Collectively, our results reveal parallels with EV biogenesis in bacteria and suggest that M. smithii EVs facilitate the export of both cellular and viral DNA as well as key metabolic proteins in the gut environment, with potential impact on methane production.

microbiology↗

bioRxiv2024

In-host evolution of Yersinia enterocolitica during a chronic human infection

Following a pacemaker implantation, a 75-years-old patient suffered from five successive bacteremia episodes between in 1999 and 2013 despite long-term antibiotic treatment, with intermittent vegetation apparition on the device atrial lead. Four blood isolates, identified as Yersinia enterocolitica bioserotype 4/O:3, were further genetically and phenotypically characterized. Phylogenetic reconstruction showed that the patient was chronically infected by the same strain, which evolved within the host for 14 years. Single-nucleotide polymorphism (SNP) analysis indicates that the last two isolates evolved in parallel and formed two independent lineages within the host. Pan-genome analysis and genome comparison showed that their common evolution was characterized by 41 small insertion/deletion events, loss of three large DNA fragments and mutations in 140 genes. A phylogenetic analysis by maximum likelihood identified two genes presenting a positive selection signal, suggesting that these mutations provided a survival advantage to bacteria during chronic infection. Quinolone resistance in the last two isolates was acquired through a so far undescribed deletion in the gyrA gene. Mass-spectrometry analysis revealed a strong proteome remodeling in the last two isolates which was correlated with a truncation in the stringent response regulator DksA. A reduced carbon, energy and purine metabolism supports their severe growth defects in vitro. 3rd-generation cephalosporin resistance of the last isolate was correlated with a truncation of OmpF, the main porin translocating antibiotics through the outer-membrane, as well as an increased production of BlaA and AmpC {beta}-lactamases. This is the first report of genetic and phenotypic changes associated to within-host adaptation of a pathogenic Yersinia species under antibiotic pressure.

microbiology↗

bioRxiv2024

Metabolic imprint of an intracellular pathogen drives histone hypermethylation and tunes the host transcriptional response to infection

Chlamydia trachomatis, an intracellular bacterium, highjacks metabolites from the host cell for its own proliferation. We provide evidence of global hypermethylation of the host proteome, including histones, during the late stages of infection. Single cell analyses revealed co-occurrence of several methylated residues on histones. Histone hypermethylation correlated with bacterial load and was prevented by antibiotic treatment. Mapping of trimethylation of histone 3 at residues K4 and K9 revealed a broad distribution throughout the chromatin. Nuclear fractions of infected cells exhibited a four-fold decrease of demethylase activity towards H3K4me3 and a two-fold increase in succinate concentration, a competitive inhibitor for the demethylase co-factor -ketoglutarate. Supplementation of the culture medium with dimethyl-ketoglutarate (DMKG) or with iron, a second co-factor of histone lysine demethylases, reduced histone hypermethylation. DMKG supplementation modified the transcription of about one third of the infection-responsive genes, indicating that histone hypermethylation contributes to modulating the transcriptional response of the host to infection. Finally, chemical inhibition of histone demethylases in a mouse model of infection showed a moderate benefit regarding the outcome of infection. Overall, our data show that the metabolic pressure exerted by a pathogen with an intracellular lifestyle drives epigenetic changes in infected cells.

cell biology↗

bioRxiv2024

A multi-year longitudinal harmonization of quality controls in mass spectrometry proteomics core facilities

Quality control procedures play a pivotal role in ensuring the reliability and consistency of data generated in mass spectrometry-based proteomics laboratories. However, the lack of standardized quality control practices across laboratories poses challenges for data comparability and reproducibility. In response, we conducted a harmonization study within proteomics laboratories of the Core for Life alliance with the aim to establish a common quality control framework, which facilitates comprehensive quality assessment and identification of potential sources of performance drift. Through collaborative efforts, we developed a consensus quality control standard for longitudinal assessment and we adopted a common processing software. We generated a 4-year longitudinal dataset from multiple instruments and laboratories, which enabled us to assess intra- and inter-laboratory variability, to identify causes of performance drift, and to establish community reference values for several quality control parameters. Our study enhances data comparability and reliability and fosters a culture of collaboration and continuous improvement within the proteomics community to ensure the integrity of proteomics data.

biochemistry↗

bioRxiv2023

Dual proteomic signature of immune cells and Yersinia pestis upon blood infection

Emerging and reemerging infectious diseases represent major public health concerns. The urgent need for infection control measures requires deep understanding of molecular pathogenesis. Global approaches to study biological systems such as mass-spectrometry based proteomics benefited from groundbreaking physical and bioinformatical technological developments over recent years. However, dual proteomic study of highly pathogenic microorganisms and their hosts in complex matrices encountered during infection remains challenging due to high protein dynamic range of samples and requirements imposed in biosafety level 3 or 4 laboratories. Here, we constructed a dual proteomic pipeline of Yersinia pestis in human blood and plasma, mirroring bacteremic phase of plague. We provide the most complete Y. pestis proteome revealing a major reshaping of important bacterial path-ways such as methionine biosynthesis and iron acquisition in human plasma. Remarkably, proteomic profiling in human blood highlights a greater Yersinia outer proteins intoxication of monocytes than neutrophils. Our study unravels global expression changes and points to a specific pathogenic signature during infection, paving the way for future exploration of proteomes in the complex context of host-pathogen interactions. Subject CategoriesMicrobiology, Virology and Host Pathogen Interaction, Proteomics

systems biology↗

bioRxiv2023

HTLV-1 Rex hijacks UPF1 in a CRM1 dependent manner, leading to NMD inhibition and revealing unexpected proviral roles of UPF1

The hijacking of CRM1 export is an important step of the retroviral replication cycle. Here, we investigated the consequences of this hijacking for the host. During HTLV-1 infection, we identified that this hijacking by the viral protein Rex favours the association between CRM1 and the RNA helicase UPF1, leading to a decreased affinity of UPF1 for cellular RNA and its nuclear retention. As a consequence, we found that the nonsense mediated mRNA decay (NMD), known to have an antiviral function, was inhibited. Corroborating these results, we described a similar process with Rev, the functional homolog of Rex from HIV-1. Unexpectedly, we also found that, for HTLV-1, this process is coupled with the specific loading of UPF1 onto vRNA, independently of NMD. In this latter context, UPF1 positively regulates several steps of the viral replication cycle, from the nuclear export of vRNA to the production of mature viral particles. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/545693v2_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1e91e61org.highwire.dtl.DTLVardef@1c9951borg.highwire.dtl.DTLVardef@15c6021org.highwire.dtl.DTLVardef@1ab35ff_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO During retroviral replication, the nuclear export unspliced vRNA is conducted via the hijacking of the exportin CRM1 by the viral protein Rex. In parallel, the RNA helicase UPF1 is naturally exported in a CRM1 dependent manner. In the cytoplasm it drives NMD, whose substrates include vRNA. Here we demonstrated that HTLV-1 Rex dependent hijacking of CRM1 is associated with the nuclear accumulation of UPF1 and the stabilization of the interaction between CRM1 and UPF1 (1). In this complex, UPF1 shows a decreased affinity for cellular RNA associated to NMD inhibition (2). We also observed that UPF1 is selectively loaded onto vRNA and stimulates vRNA export (3). In this context, UPF1 is driven in the viral particles (without NMD cofactors) where it plays critical role in virion assembly, maturation (4) and ultimately viral infection (5). Created in BioRender. PROCHASSON, L. (2025) https://BioRender.com/urj0cvo". C_FIG

molecular biology↗

bioRxiv2023

Mtfp1 ablation enhances mitochondrial respiration and protects against hepatic steatosis

Hepatic steatosis is the result of an imbalance between nutrient delivery and metabolism in the liver. It is the first hallmark of Non-alcoholic fatty liver disease (NAFLD) and is characterized by the accumulation of excess lipids in the liver that can drive liver failure, inflammation, and cancer. Mitochondria control the fate and function of cells and compelling evidence implicates these multifunctional organelles in the appearance and progression of liver dysfunction, although it remains to be elucidated which specific mitochondrial functions are actually causally linked to NAFLD. Here, we identified Mitochondrial Fission Process 1 protein (MTFP1) as a key regulator of mitochondrial and metabolic activity in the liver. Deletion of Mtfp1 in hepatocytes is physiologically benign in mice yet leads to the upregulation of oxidative phosphorylation (OXPHOS) activity and mitochondrial respiration, independently of mitochondrial biogenesis. Consequently, hepatocyte-specific knockout mice are protected against high fat diet-induced hepatic steatosis and metabolic dysregulation. Additionally, we find that deletion of Mtfp1 in liver mitochondria inhibits mitochondrial permeability transition pore opening in hepatocytes, conferring protection against apoptotic liver damage in vivo and ex vivo. Our work uncovers novel functions of MTFP1 in the liver, positioning this gene as an unexpected regulator of OXPHOS and a therapeutic candidate for NAFLD.

physiology↗

bioRxiv2023

The RBPome of influenza A virus mRNA reveals a role for TDP-43 in viral replication

Recent technical advances have significantly improved our understanding of the RNA-binding protein (RBP) repertoire present within eukaryotic cells, with a particular focus on the RBPs that interact with cellular polyadenylated mRNAs. However, recent studies utilising the same technologies have begun to tease apart the RBP interactome of viral mRNAs, notably SARS-CoV-2, revealing both similarities and differences between the RBP profiles of viral and cellular mRNAs. Herein, we comprehensively identified the RBPs that associate with the NP mRNA of an influenza A virus. Moreover, we provide evidence that the viral polymerase is essential for the recruitment of RPBs to viral mRNAs through direct polymerase-RBP interactions during transcription. We show that loss of TDP-43, which associates with the viral mRNAs, results in lower levels of viral mRNAs within infected cells, and a decreased yield of infectious viral particles. Overall, our results uncover an important role for TDP-43 in the influenza A virus replication cycle via a direct interaction with viral mRNAs, and point to a role of the viral polymerase in orchestrating the assembly of viral mRNPs.

microbiology↗

bioRxiv2022

Proteomic landscape of tunneling nanotubes reveals CD9 and CD81 tetraspanins as key regulators

Tunneling nanotubes (TNTs) are open actin- and membrane-based channels, connecting remote cells and allowing direct transfer of cellular material (e.g. vesicles, mRNAs, protein aggregates) from cytoplasm to cytoplasm. Although they are important especially in pathological conditions (e.g., cancers, neurodegenerative diseases), their precise composition and their regulation were still poorly described. Here, using a biochemical approach allowing to separate TNTs from cell bodies and from extracellular vesicles and particles (EVPs), we obtained the full composition of TNTs compared to EVPs. We then focused to two major components of our proteomic data, the CD9 and CD81 tetraspanins, and further investigated their specific roles in TNT formation and function. We show that these two tetraspanins have distinct non-redundant functions: CD9 participates in stabilizing TNTs, whereas CD81 expression is required to allow the functional transfer of vesicle in the newly formed TNTs, possibly by regulating docking to or fusion with the opposing cell.

cell biology↗

bioRxiv2022

The cell wall lipoprotein CD1687 acts as a DNA binding protein during deoxycholate-induced biofilm formation in Clostridioides difficile

The ability of bacterial pathogens to establish recurrent and persistent infections is frequently associated with their ability to form biofilms. Clostridioides difficile infections have a high rate of recurrence and relapses and it is hypothesised that biofilms are involved in its pathogenicity and persistence. Biofilm formation by C. difficile is still poorly understood. It has been shown that specific molecules such as deoxycholate (DCA) or metronidazole induce biofilm formation, but the mechanisms involved remain elusive. In this study, we describe the role of the C. difficile lipoprotein CD1687 during DCA-induced biofilm formation. We showed that the expression of CD1687, which is part of an operon within the CD1685-CD1689 gene cluster, is controlled by multiple transcription starting sites and some are induced in response to DCA. Only CD1687 is required for biofilm formation and the overexpression of CD1687 is sufficient to induce biofilm formation. Using RNAseq analysis, we showed that CD1687 affects the expression of transporters and metabolic pathways and we identified several potential binding partners by pull-down assay, including transport-associated extracellular proteins. We then demonstrated that CD1687 is surface exposed in C. difficile, and that this localization is required for DCA-induced biofilm formation. Given this localization and the fact that C. difficile forms eDNA-rich biofilms, we confirmed that CD1687 binds DNA in a non-specific manner. We thus hypothesize that CD1687 is a component of the downstream response to DCA leading to biofilm formation by promoting interaction between the cells and the biofilm matrix by binding eDNA.

microbiology↗

bioRxiv2022

Rasputin/G3BP mediates o-nyong-nyong virus subversion of antiviral immunity in Anopheles coluzzii

The G3BP proteins in vertebrates and Aedes mosquito ortholog, Rasputin, are essential for alphavirus infection, but the underlying mechanism of Rasputin/G3BP proviral activity is poorly understood. It has been suggested that G3BP could influence host immune signaling, but this has not been functionally demonstrated. Here, we find that depletion of Rasputin activity in Anopheles mosquitoes, the primary vectors of the alphavirus onyong-nyong (ONNV), provokes dysregulation of the antiviral Imd, JAK/STAT and RNAi pathways, indicating that Rasputin is required for expression of normal basal immunity in uninfected mosquitoes. Depletion of Rasputin during ONNV bloodmeal infection causes increased transcript abundance of genes in the Imd pathway including positive regulator Rel2, and decreases ONNV infection in mosquitoes. Loss of Rasputin is complemented by co-depletion of Imd pathway positive regulator, Rel2, which restores normal ONNV infection levels. Thus, the presence of Rasputin is required for ONNV inhibition of Imd activity, and viral inhibition of Imd explains much of the Rasputin proviral activity. The viral non-structural protein 3 (nsP3) binds to Rasputin and alters the profile of cellular proteins binding to Rasputin. In the presence of nsP3, 48 Rasputin-binding proteins are unchanged but seven binding proteins are excluded and eight new proteins bind Rasputin. The Rasputin binding partners altered by nsP3 are candidate factors for ONNV immune manipulation and subversion through Rasputin. Overall, these results are consistent with and strongly suggest a mechanism in which ONNV, probably nsP3, co-opts the normal Rasputin function assuring basal cellular immune activity in order to inhibit antiviral immunity and promote infection. These observations may be generalizable for Rasputin function during alphavirus infection of other mosquitoes, as well as for G3BP function in the mammalian host, and could offer a target for vector-based control of arbovirus transmission.

microbiology↗

bioRxiv2022

Queuosine modification of tRNA-Tyrosine elicits translational reprogramming and enhances growth of Vibrio cholerae with aminoglycosides.

Tgt is the enzyme modifying the guanine (G) in tRNAs with GUN anticodon to queuosine (Q). tgt is required for optimal growth of Vibrio cholerae in the presence of sub-lethal aminoglycoside concentrations. We further explored here the role of the Q34 in the efficiency of codon decoding upon tobramycin exposure. We characterized its impact on the overall bacterial proteome, and elucidated the molecular mechanisms underlying the effects of Q34 modification in antibiotic translational stress response. Using molecular reporters, we showed that Q34 impacts the efficiency of decoding at tyrosine TAT and TAC codons. Proteomics analyses revealed that the anti-SoxR factor RsxA is better translated in the absence of tgt. RsxA displays a codon bias towards tyrosine TAT and overabundance of RsxA leads to decreased expression of genes belonging to SoxR oxidative stress regulon. We also identified conditions that regulate tgt expression. We propose that regulation of Q34 modification in response to environmental cues leads to translational reprogramming of transcripts bearing a biased tyrosine codon usage. In silico analysis further identified candidate genes which could be subject to such translational regulation, among which DNA repair factors. Such transcripts, fitting the definition of modification tunable transcripts, are central in the bacterial response to antibiotics.

microbiology↗

bioRxiv2022

Hemisynthetic derivatives of the natural alkaloid trilobine are fast-acting antimalarial compounds with sustained activity in multi-drug resistant P. falciparum isolates

Malaria eradication requires the development of new drugs to combat drug-resistant parasites. The search for new chemical scaffolds that target novel pathways of the human malaria parasite Plasmodium falciparum is of highest priority. We identified bisbenzylisoquinoline alkaloids isolated from Cocculus hirsutus. (trilobine derivatives) as active in the nanomolar range against P. falciparum blood stages. Synthesis of a library of 94 hemi-synthetic derivatives allowed us to identify compound 84 that kills multi-drug resistant clinical isolates in the nanomolar range (median IC50 ranging from 35-88nM). Efforts were made to obtain compounds with significantly improved preclinical properties. Out of those, compound 125 delays the onset of parasitemia in P. berghei infected mice and inhibits P. falciparum transmission stages in vitro (culture assays) and in vivo using membrane feeding assay in the Anopheles stephensi vector. Compound 125 also impairs P. falciparum development in sporozoite-infected hepatocytes, in the low micromolar range. Finally, we used a chemical pull-down strategy to identify potential protein targets of this chemical family. Mass spectrometry analysis identified the parasite interactome with trilobine derivatives, identifying protein partners belonging to metabolic pathways that have not been previously targeted by antimalarial drugs or implicated in drug-resistance mechanisms.

pathology↗

bioRxiv2022

Arp2/3 inhibition switches Eps8's network associations to favour longer actin filament formation necessary for tunneling nanotubes

Tunneling nanotubes (TNTs) connect distant cells and mediate cargo transfer for intercellular communication in physiological and pathological contexts. How cells generate these actin-mediated protrusions to span lengths beyond those attainable by canonical filopodia remains unknown. Through a combination of micropatterning, microscopy and optical tweezer-based approaches, we demonstrate that TNTs forming through the outward extension of actin (not through cellular dislodgement) achieve distances greater than the mean length of filopodia, and that branched Arp2/3-dependent pathways attenuate the extent to which actin polymerizes in nanotubes, limiting TNT occurrence. Proteomic analysis using Epidermal growth factor receptor kinase substrate 8 (Eps8) as a positive effector of TNTs showed that upon Arp2/3 inhibition, proteins enhancing filament turnover and depolymerization were reduced and Eps8 instead exhibited heightened interactions with the inverted Bin/Amphiphysin/Rvs (I-BAR) domain protein IRSp53 that provides a direct connection with linear actin polymerases. Our data reveals how common protrusion players (Eps8 and IRSp53) form TNTs, and that when competing pathways overutilizing such proteins and monomeric actin in Arp2/3 networks are inhibited, processes promoting linear actin growth dominate to favour TNT formation. Thus, this work reinforces a general principle for actin network control for cellular protrusions where simple shifts in the balance between processes that inhibit actin growth versus those that promote growth dictate protrusion formation and the ultimate length scales protrusions achieve.

cell biology↗

bioRxiv2022

Single locus phosphoproteomics reveals phosphorylation of RPA-1 is required for generation of single-strand DNA following a break at a subtelomeric locus.

Damage to the genetic material of the cell poses a universal threat to all forms of life. Central to the DNA damage response (DDR) is a phosphorylation signalling cascade that leads to the co-ordination of the cellular response to a DNA break. Identifying the proteins that are phosphorylated is crucial to understanding the mechanisms underlying this DDR. We have used SILAC-based quantitative phosphoproteomics to profile changes in phosphorylation site abundance following a single double strand break (DSB) at a chromosome internal locus and the subtelomeric bloodstream form expression site in Trypanosoma brucei. We report >6500 phosphorylation sites, including a core set of 211 DSB responsive phosphorylation sites. Along with phosphorylation of canonical DNA damage factors, we find that there is a striking distinction between the proteins phosphorylated in response to a chromosome internal DSB and one at the active BES and describe a single phosphorylation event on Replication factor A (RPA) 1 that is required for efficient resection at a bloodstream form expression site.

molecular biology↗

bioRxiv2022

Pneumococcus triggers NFkB degradation in COMMD2 aggresome-like bodies.

NF-{kappa}B driven cellular immunity is essential for both pro- and anti-inflammatory responses to microbes, which makes it one of the most frequently targeted pathways by bacteria during pathogenesis. How NF-{kappa}B tunes the epithelial response to Streptococcus pneumoniae across the spectrum of commensal to pathogenic outcomes is not fully understood. In this study, we compare a commensal-like 6B ST90 strain to an invasive TIGR4 isolate and demonstrate, through comparative mass spectrometry of the p65 interactome, TIGR4 challenge triggers a novel interaction of COMMD2 with p65 and p62. Mechanistically, we show this complex mediates export of p65 for degradation and COMMD2 is necessary for altering host cellular immunity. With these results, we reveal for the first time a new bacterial pathogenesis mechanism to repress host inflammatory response though COMMD2 and p65 degradation while presenting a paradigm for diverging NF-{kappa}B responses to pneumococcus.

microbiology↗

bioRxiv2022

MSclassifR: an R package for supervised classification of mass spectra with machine learning methods

MSclassifR is an R package that has been specifically designed to improve the classification of mass spectra obtained from MALDI-TOF mass spectrometry. It offers a comprehensive range of functions that are focused on processing mass spectra, identifying discriminant m/z values, and making accurate predictions. The package introduces innovative algorithms for selecting discriminating m/z values and making predictions. To assess the effectiveness of these methods, extensive tests were conducted using challenging real datasets, including bacterial subspecies of the Mycobacterium abscessus complex, virulent and avirulent phenotypes of Escherichia coli, different species of Streptococci and nasal swabs from individuals infected and uninfected with SARS-CoV-2. Additionally, multiple datasets of varying sizes were created from these real datasets to evaluate the robustness of the algorithms. The results demonstrated that the Machine Learning-based pipelines in MSclassifR achieved high levels of accuracy and Kappa values. On an in-house dataset, some pipelines even achieved more than 95% mean accuracy, whereas commercial system only achieved 62% mean accuracy. Certain methods showed greater resilience to changes in dataset sizes when constructing Machine Learning-based pipelines. These simulations also helped determine the minimum sizes of training sets required to obtain reliable results. The package is freely available online, and its open-source nature encourages collaborative development, customization, and fosters innovation within the community focused on improving diagnosis based on MALDI-TOF spectra. Key pointsO_LIMSclassifR is a comprehensive R package enabling the construction of data analysis pipelines for the precise classification of mass spectra. C_LIO_LIOur R package contains an innovative method for variable selection from random forests, which delivered excellent results on real data. C_LIO_LIIn-depth analysis of various machine learning-based pipelines using our package allowed us to make conclusions about the optimal m/z selection and prediction methods depending on the size of the training dataset. C_LIO_LIUsing a publicly available dataset of mass spectra obtained from various MALDI-TOF instruments across different countries, MSclassifR is able to build robust pipelines capable of adapting to different instruments in an automatic way. C_LIO_LIWhen tested on an in-house dataset, MSclassifR pipelines consistently outperformed a commercial software in terms of prediction accuracy. C_LI

bioinformatics↗

bioRxiv2021

Mitochondrial fission process 1 (MTFP1) controls bioenergetic efficiency and prevents inflammatory cardiomyopathy and heart failure in mice

Mitochondria are paramount to the metabolism and survival of cardiomyocytes. Here we show that Mitochondrial Fission Process 1 (MTFP1) is essential for cardiac structure and function. Constitutive knockout of cardiomyocyte MTFP1 in mice resulted in adult-onset dilated cardiomyopathy (DCM) characterized by sterile inflammation and cardiac fibrosis that progressed to heart failure and middle-aged death. Failing hearts from cardiomyocyte-restricted knockout mice displayed a general decline in mitochondrial gene expression and oxidative phosphorylation (OXPHOS) activity. Pre-DCM, we observed no defects in mitochondrial morphology, content, gene expression, OXPHOS assembly nor phosphorylation dependent respiration. However, knockout cardiac mitochondria displayed reduced membrane potential and increased non-phosphorylation dependent respiration, which could be rescued by pharmacological inhibition of the adenine nucleotide translocase ANT. Primary cardiomyocytes from pre-symptomatic knockout mice exhibited normal excitation-contraction coupling but increased sensitivity to programmed cell death (PCD), which was accompanied by an opening of the mitochondrial permeability transition pore (mPTP). Intriguingly, mouse embryonic fibroblasts deleted for Mtfp1 recapitulated PCD sensitivity and mPTP opening, both of which could be rescued by pharmacological or genetic inhibition of the mPTP regulator Cyclophilin D. Collectively, our data demonstrate that contrary to previous in vitro studies, the loss of the MTFP1 promotes mitochondrial uncoupling and increases cell death sensitivity, causally mediating pathogenic cardiac remodeling.

physiology↗