Factors influencing the bioactivity of natural matrices: The case of osmolarity-dependent modulation of cell viability by different dilutions of camel urines.

The widespread practice of dromedary urinotherapy as a remedy for various illnesses, including cancer, is well-established in traditional dromedary countries. Researchers attempted to demonstrate anticancer properties of camel urine through in vitro experiments with debated outcomes. Notably, two critical aspects remained unexplored in those assays: (i) the osmolarity of tested urines, which can significantly influence in vitro results; (ii) the potential morphological changes of cells, following exposure to camel urines. In this study, we addressed these gaps by evaluating the osmolarity-dependent modulation of cell viability in human renal cell lines. In this regard, we assessed the impact of hyperosmolar mannitol-based solutions and dromedary urine on the viability and morphology of human non-tumor (HK2) and tumor renal cells (Caki-1). The results indicate that cell viability or morphology in both HK2 and Caki-1 cells are not significantly affected only if mannitol-induced hyperosmolarity is lower than 500 mOsm/L. Notably, when exposed to urine solution, diluted to <500 mOsm/L, statistically significant antiproliferative effects were observed primarily in Caki-1 cells (in presence of two out of ten tested urine samples). Conversely, alterations in cell morphology were observed exclusively in HK2 cells when exposed to the same diluted camel urines. In order to investigate, at molecular level, the observed antiproliferative effects, a preliminary metabolomics analysis of the tested urine samples was performed to identify potential bioactive compounds. The Nuclear Magnetic Resonance (NMR) metabolic profiling revealed the presence of three antioxidant compounds, namely trigonelline, pyruvic acid and N-acetylglucosamine. In conclusion, our results highlight the importance of considering the critical role of osmolarity when evaluating the bioactive properties of camel urine in vitro, which should not be used to treat any illness as it is. Conversely, it can be considered the possibility to use camel urines as a source of bioactive compounds.

Uncovering a Genetic Diagnosis in a Pediatric Patient by Whole Exome Sequencing: A Modeling Investigation in Wiedemann-Steiner Syndrome.

Background: Wiedemann-Steiner syndrome (WSS), a rare autosomal-dominant disorder caused by haploinsufficiency of the KMT2A gene product, is part of a group of disorders called chromatinopathies. Chromatinopathies are neurodevelopmental disorders caused by mutations affecting the proteins responsible for chromatin remodeling and transcriptional regulation. The resulting gene expression dysregulation mediates the onset of a series of clinical features such as developmental delay, intellectual disability, facial dysmorphism, and behavioral disorders. Aim of the Study: The aim of this study was to investigate a 10-year-old girl who presented with clinical features suggestive of WSS. Methods: Clinical and genetic investigations were performed. Whole exome sequencing (WES) was used for genetic testing, performed using Illumina technology. The bidirectional capillary Sanger resequencing technique was used in accordance with standard methodology to validate a mutation discovered by WES in all family members who were available. Utilizing computational protein modeling for structural and functional studies as well as in silico pathogenicity prediction models, the effect of the mutation was examined. Results: WES identified a de novo heterozygous missense variant in the KMT2A gene KMT2A(NM_001197104.2): c.3451C>G, p.(Arg1151Gly), absent in the gnomAD database. The variant was classified as Likely Pathogenetic (LP) according to the ACMG criteria and was predicted to affect the CXXC-type zinc finger domain functionality of the protein. Modeling of the resulting protein structure suggested that this variant changes the protein flexibility due to a variation in the Gibbs free energy and in the vibrational entropy energy difference between the wild-type and mutated domain, resulting in an alteration of the DNA binding affinity. Conclusions: A novel and de novo mutation discovered by the NGS approach, enhancing the mutation spectrum in the KMT2A gene, was characterized and associated with WSS. This novel KMT2A gene variant is suggested to modify the CXXC-type zinc finger domain functionality by affecting protein flexibility and DNA binding.

Supercritical Fluid Extraction of Bioactive Components from Apple Peels and Their Modulation of Complex I Activity in Isolated Mitochondria.

Supercritical fluid extraction (SFE) was used to extract bioactive compounds from apple (Malus domestica) peel waste from three different Italian cultivars. The bioactive fractions were extracted applying a temperature of 60 °C and a pressure of 250 bar for 15 min with 20% ethanol as co-solvent, at a flow rate of 2 mL/min. The total polyphenol (TP), anthocyanin (TA), ascorbic acid (AA), and antioxidant activity contents (TACs) were measured, while chromatographic analyses were performed to highlight the differences between the extracts. The Stark cultivar had the highest levels of polyphenols, anthocyanins, and ascorbic acid, while the Royal Gala cultivar showed the highest total antioxidant activity. SFE extracts were then tested for their effect on the mitochondrial NADH-ubiquinone oxidoreductase (Complex I) activity on mitochondria isolated from human embryonic kidney cells (HEK239). The Stark extract showed the most positive response in terms of NADH oxidation. The results obtained in this work highlight the potential of apple peel waste as a source of functional phytocompounds and suggest that Stark cultivar extracts may be exploited for pharmacological applications. This study supports the circular bioeconomy by promoting the use of waste products as a valuable resource.

Nucleotide substitutions at the p.Gly117 and p.Thr180 mutational hot-spots of SKI alter molecular dynamics and may affect cell cycle.

Heterozygous deleterious variants in SKI cause Shprintzen-Goldberg Syndrome, which is mainly characterized by craniofacial features, neurodevelopmental disorder and thoracic aorta dilatations/aneurysms. The encoded protein is a member of the transforming growth factor beta signaling. Paucity of reported studies exploring the SGS molecular pathogenesis hampers disease recognition and clinical interpretation of private variants. Here, the unpublished c.349G>A, p.[Gly117Ser] and the recurrent c.539C>T, p.[Thr180Met] SKI variants were studied combining in silico and in vitro approach. 3D comparative modeling and calculation of the interaction energy predicted that both variants alter the SKI tertiary protein structure and its interactions. Computational data were functionally corroborated by the demonstration of an increase of MAPK phosphorylation levels and alteration of cell cycle in cells expressing the mutant SKI. Our findings confirmed the effects of SKI variants on MAPK and opened the path to study the role of perturbations of the cell cycle in SGS.

TNFRSF13B gene mutation in familial acute myeloid leukemia: A new piece in the complex scenario of hereditary predisposition?

TNFRSF13B mutations are widely associated with common variable immunodeficiency. TNFRSF13B was recently counted among relevant genes associated with childhood-onset of hematological malignancies; nonetheless, its role in acute myeloid leukemia (AML) remains unexplored. We report the study of a family with two cases of AML, sharing a germline TNFRSF13B mutation favoring the formation of a more stable complex with its ligand TNFSF13: a positive regulator of AML-initiating cells. Our data turn the spotlight onto the TNFRSF13B role in AML onset, inserting a new fragment into the complex scenario of a hereditary predisposition to myeloid neoplasms.

Case report: biallelic DNMT3A mutations in acute myeloid leukemia.

DNMT3A gene mutations, detected in 20-25% of de novo acute myeloid leukemia (AML) patients, are typically heterozygous. Biallelic variants are uncommon, affecting ~3% of cases and identifying a worse prognosis. Indeed, two concomitant DNMT3A mutations were recently associated with shorter event-free survival and overall survival in AML. We present an AML case bearing an unusual DNMT3A molecular status, strongly affecting its function and strangely impacting the global genomic methylation profile. A 56-year-old Caucasian male with a diagnosis of AML not otherwise specified (NOS) presented a complex DNMT3A molecular profile consisting of four different somatic variants mapping on different alleles (in trans). 3D modelling analysis predicted the effect of the DNMT3A mutational status, showing that all the investigated mutations decreased or abolished DNMT3A activity. Although unexpected, DNMT3A's severe loss of function resulted in a global genomic hypermethylation in genes generally involved in cell differentiation. The mechanisms through which DNMT3A contributes to AML remain elusive. We present a unique AML case bearing multiple biallelic DNMT3A variants abolishing its activity and resulting in an unexpected global hypermethylation. The unusual DNMT3A behavior described requires a reflection on its role in AML development and persistence, highlighting the heterogeneity of its deregulation.

The Mycotoxin Patulin Inhibits the Mitochondrial Carnitine/Acylcarnitine Carrier (SLC25A20) by Interaction with Cys136 Implications for Human Health.

The effect of mycotoxin patulin (4-hydroxy-4H-furo [3,2c] pyran-2 [6H] -one) on the mitochondrial carnitine/acylcarnitine carrier (CAC, SLC25A20) was investigated. Transport function was measured as [3H]-carnitineex/carnitinein antiport in proteoliposomes reconstituted with the native protein extracted from rat liver mitochondria or with the recombinant CAC over-expressed in E. coli. Patulin (PAT) inhibited both the mitochondrial native and recombinant transporters. The inhibition was not reversed by physiological and sulfhydryl-reducing reagents, such as glutathione (GSH) or dithioerythritol (DTE). The IC50 derived from the dose-response analysis indicated that PAT inhibition was in the range of 50 µM both on the native and on rat and human recombinant protein. The kinetics process revealed a competitive type of inhibition. A substrate protection experiment confirmed that the interaction of PAT with the protein occurred within a protein region, including the substrate-binding area. The mechanism of inhibition was identified using the site-directed mutagenesis of CAC. No inhibition was observed on Cys mutants in which only the C136 residue was mutated. Mass spectrometry studies and in silico molecular modeling analysis corroborated the outcomes derived from the biochemical assays.

Targeting mitochondrial impairment for the treatment of cardiovascular diseases: From hypertension to ischemia-reperfusion injury, searching for new pharmacological targets.

Mitochondria and mitochondrial proteins represent a group of promising pharmacological target candidates in the search of new molecular targets and drugs to counteract the onset of hypertension and more in general cardiovascular diseases (CVDs). Indeed, several mitochondrial pathways result impaired in CVDs, showing ATP depletion and ROS production as common traits of cardiac tissue degeneration. Thus, targeting mitochondrial dysfunction in cardiomyocytes can represent a successful strategy to prevent heart failure. In this context, the identification of new pharmacological targets among mitochondrial proteins paves the way for the design of new selective drugs. Thanks to the advances in omics approaches, to a greater availability of mitochondrial crystallized protein structures and to the development of new computational approaches for protein 3D-modelling and drug design, it is now possible to investigate in detail impaired mitochondrial pathways in CVDs. Furthermore, it is possible to design new powerful drugs able to hit the selected pharmacological targets in a highly selective way to rescue mitochondrial dysfunction and prevent cardiac tissue degeneration. The role of mitochondrial dysfunction in the onset of CVDs appears increasingly evident, as reflected by the impairment of proteins involved in lipid peroxidation, mitochondrial dynamics, respiratory chain complexes, and membrane polarization maintenance in CVD patients. Conversely, little is known about proteins responsible for the cross-talk between mitochondria and cytoplasm in cardiomyocytes. Mitochondrial transporters of the SLC25A family, in particular, are responsible for the translocation of nucleotides (e.g., ATP), amino acids (e.g., aspartate, glutamate, ornithine), organic acids (e.g. malate and 2-oxoglutarate), and other cofactors (e.g., inorganic phosphate, NAD+, FAD, carnitine, CoA derivatives) between the mitochondrial and cytosolic compartments. Thus, mitochondrial transporters play a key role in the mitochondria-cytosol cross-talk by leading metabolic pathways such as the malate/aspartate shuttle, the carnitine shuttle, the ATP export from mitochondria, and the regulation of permeability transition pore opening. Since all these pathways are crucial for maintaining healthy cardiomyocytes, mitochondrial carriers emerge as an interesting class of new possible pharmacological targets for CVD treatments.

Genome-Wide Identification and Validation of Gene Expression Biomarkers in the Diagnosis of Ovarian Serous Cystadenocarcinoma.

Ovarian cancer is the second most prevalent gynecologic malignancy, and ovarian serous cystadenocarcinoma (OSCA) is the most common and lethal subtype of ovarian cancer. Current screening methods have strong limits on early detection, and the majority of OSCA patients relapse. In this work, we developed and cross-validated a method for detecting gene expression biomarkers able to discriminate OSCA tissues from healthy ovarian tissues and other cancer types with high accuracy. A preliminary ranking-based approach was applied, resulting in a panel of 41 over-expressed genes in OSCA. The RNA quantity gene expression of the 41 selected genes was then cross-validated by using NanoString nCounter technology. Moreover, we showed that the RNA quantity of eight genes (ADGRG1, EPCAM, ESRP1, MAL2, MYH14, PRSS8, ST14 and WFDC2) discriminates each OSCA sample from each healthy sample in our data set with sensitivity of 100% and specificity of 100%. For the other three genes (MUC16, PAX8 and SOX17) in combination, their RNA quantity may distinguish OSCA from other 29 tumor types.

The Non-Gastric H+/K+ ATPase (ATP12A) Is Expressed in Mammalian Spermatozoa.

H+/K+ ATPase Type 2 is an heteromeric membrane protein involved in cation transmembrane transport and consists of two subunits: a specific α subunit (ATP12A) and a non-specific ß subunit. The aim of this study was to demonstrate the presence and establish the localization of ATP12A in spermatozoa from Bubalus bubalis, Bos taurus and Ovis aries. Immunoblotting revealed, in all three species, a major band (100 kDa) corresponding to the expected molecular mass. The ATP12A immunolocalization pattern showed, consistently in the three species, a strong signal at the acrosome. These results, described here for the first time in spermatozoa, are consistent with those observed for the ß1 subunit of Na+/K+ ATPase, suggesting that the latter may assemble with the α subunit to produce a functional ATP12A dimer in sperm cells. The above scenario appeared to be nicely supported by 3D comparative modeling and interaction energy calculations. The expression of ATP12A during different stages of bovine sperm maturation progressively increased, moving from epididymis to deferent ducts. Based on overall results, we hypothesize that ATP12A may play a role in acrosome reactions. Further studies will be required in order to address the functional role of this target protein in sperm physiology.

Inverse Virtual Screening for the rapid re-evaluation of the presumed biological safe profile of natural products. The case of steviol from Stevia rebaudiana glycosides on farnesoid X receptor (FXR).

Nonnutritive sweeteners (NNSs) are widely employed as dietary substitutes for classical sugars thanks to their safety profile and low toxicity. In this study, a re-evaluation of the biological effects of steviol (1), the main metabolite from Stevia rebaudiana glycosides, was performed using the Inverse Virtual Screening (IVS) target fishing computational approach. Starting from well-known pharmacological properties of Stevia rebaudiana glycosides, this computational tool was employed for predicting the putative interacting targets of 1 and, afterwards, of its five synthetic ester derivatives 2-6, accounting a large panel of proteins involved in cancer and inflammation events. Applying this methodology, the farnesoid X receptor (FXR) was identified as the putative target partner of 1-6. The predicted ligand-protein interactions were corroborated by transactivation assays, specifically disclosing the agonistic activity of 1 and the antagonistic activities of 2-6 on FXR. The reported results highlight the feasibility of IVS as a fast and potent tool for predicting the interacting targets of query compounds, addressing the re-evaluation of their bioactivity. In light of the obtained results, the presumably safe profile of known compounds, such as the case of steviol (1), is critically discussed.

Major antigen and paramyosin proteins as candidate biomarkers for serodiagnosis of canine infection by zoonotic Onchocerca lupi.

Onchocerca lupi (Spirurida: Onchocercidae) is a filarial worm parasitizing domestic carnivores and humans. Adult nematodes usually localize beneath in the sclera or in the ocular retrobulbar of infected animals, whilst microfilariae are found in the skin. Therefore, diagnosis of O. lupi is achieved by microscopic and/or molecular detection of microfilariae from skin biopsy and/or surgical removal of adults from ocular tissues of infected hosts. An urgent non-invasive diagnostic tool for the diagnosis of O. lupi in dog is mandatory. In this study, an immunoproteomic analyses was performed using a combination of immunoblotting and mass spectrometry techniques. Onchocerca lupi major antigen (Ol-MJA) and paramyosin (Ol-PARA) proteins were identified as potential biomarkers for serodiagnosis. Linear epitopes were herein scanned for both proteins using high-density peptide microarray. Sera collected from dog infected with O. lupi and healthy animal controls led to the identification of 11 immunodominant antigenic peptides (n = 7 for Ol-MJA; n = 4 for Ol-PARA). These peptides were validated using sera of dogs uniquely infected with the most important filarioids infesting dogs either zoonotic (Dirofilaria repens, Dirofilaria immitis) or not (Acanthocheilonema reconditum and Cercopithifilaria bainae). Overall, six antigenic peptides, three for Ol-MJA and for Ol-PARA, respectively, were selected as potential antigens for the serological detection of canine O. lupi infection. The molecular and proteomic dataset herein reported should provide a useful resource for studies on O. lupi toward supporting the development of new interventions (drugs, vaccines and diagnostics) against canine onchocercosis.

Targeting the mitochondrial trifunctional protein restrains tumor growth in oxidative lung carcinomas.

Metabolic reprogramming is a common hallmark of cancer, but a large variability in tumor bioenergetics exists between patients. Using high-resolution respirometry on fresh biopsies of human lung adenocarcinoma, we identified 2 subgroups reflected in the histologically normal, paired, cancer-adjacent tissue: high (OX+) mitochondrial respiration and low (OX-) mitochondrial respiration. The OX+ tumors poorly incorporated [18F]fluorodeoxy-glucose and showed increased expression of the mitochondrial trifunctional fatty acid oxidation enzyme (MTP; HADHA) compared with the paired adjacent tissue. Genetic inhibition of MTP altered OX+ tumor growth in vivo. Trimetazidine, an approved drug inhibitor of MTP used in cardiology, also reduced tumor growth and induced disruption of the physical interaction between the MTP and respiratory chain complex I, leading to a cellular redox and energy crisis. MTP expression in tumors was assessed using histology scoring methods and varied in negative correlation with [18F]fluorodeoxy-glucose incorporation. These findings provide proof-of-concept data for preclinical, precision, bioenergetic medicine in oxidative lung carcinomas.

Sweet as honey, bitter as bile: Mitochondriotoxic peptides and other therapeutic proteins isolated from animal tissues, for dealing with mitochondrial apoptosis.

Mitochondria are subcellular organelles involved in cell metabolism and cell life-cycle. Their role in apoptosis regulation makes them an interesting target of new drugs for dealing with cancer or rare diseases. Several peptides and proteins isolated from animal and plant sources are known for their therapeutic properties and have been tested on cancer cell-lines and xenograft murine models, highlighting their ability in inducing cell-death by triggering mitochondrial apoptosis. Some of those molecules have been even approved as drugs. Conversely, many other bioactive compounds are still under investigation for their proapoptotic properties. In this review we report about a group of peptides, isolated from animal venoms, with potential therapeutic properties related to their ability in triggering mitochondrial apoptosis. This class of compounds is known with different names, such as mitochondriotoxins or mitocans.

Mitochondrial Membranes of Human SH-SY5Y Neuroblastoma Cells Express Serotonin 5-HT7 Receptor.

Mitochondria in neurons contribute to energy supply, the regulation of synaptic transmission, Ca2+ homeostasis, neuronal excitability, and stress adaptation. In recent years, several studies have highlighted that the neurotransmitter serotonin (5-HT) plays an important role in mitochondrial biogenesis in cortical neurons, and regulates mitochondrial activity and cellular function in cardiomyocytes. 5-HT exerts its diverse actions by binding to cell surface receptors that are classified into seven distinct families (5-HT1 to 5-HT7). Recently, it was shown that 5-HT3 and 5-HT4 receptors are located on the mitochondrial membrane and participate in the regulation of mitochondrial function. Furthermore, it was observed that activation of brain 5-HT7 receptors rescued mitochondrial dysfunction in female mice from two models of Rett syndrome, a rare neurodevelopmental disorder characterized by severe behavioral and physiological symptoms. Our Western blot analyses performed on cell-lysate and purified mitochondria isolated from neuronal cell line SH-SY5Y showed that 5-HT7 receptors are also expressed into mitochondria. Maximal binding capacity (Bmax) obtained by Scatchard analysis on purified mitochondrial membranes was 0.081 pmol/mg of 5-HT7 receptor protein. Lastly, we evaluated the effect of selective 5-HT7 receptor agonist LP-211 and antagonist (inverse agonist) SB-269970 on mitochondrial respiratory chain (MRC) cytochrome c oxidase activity on mitochondria from SH-SY5Y cells. Our findings provide the first evidence that 5-HT7 receptor is also expressed in mitochondria.

Targeting Human Lung Adenocarcinoma with a Suppressor of Mitochondrial Superoxide Production.

Aims: REDOX signaling from reactive oxygen species (ROS) generated by the mitochondria (mitochondrial reactive oxygen species [mtROS]) has been implicated in cancer growth and survival. Here, we investigated the effect of 5-(4-methoxyphenyl)-3H-1,2-dithiole-3-thione (AOL), a recently characterized member of the new class of mtROS suppressors (S1QELs), on human lung adenocarcinoma proteome reprogramming, bioenergetics, and growth. Results: AOL reduced steady-state cellular ROS levels in human lung cancer cells without altering the catalytic activity of complex I. AOL treatment induced dose-dependent inhibition of lung cancer cell proliferation and triggered a reduction in tumor growth in vivo. Molecular investigations demonstrated that AOL reprogrammed the proteome of human lung cancer cells. In particular, AOL suppressed the determinants of the Warburg effect and increased the expression of the complex I subunit NDUFV1 which was also identified as AOL binding site using molecular modeling computer simulations. Comparison of the molecular changes induced by AOL and MitoTEMPO, an mtROS scavenger that is not an S1QEL, identified a core component of 217 proteins commonly altered by the two treatments, as well as drug-specific targets. Innovation: This study provides proof-of-concept data on the anticancer effect of AOL on mouse orthotopic human lung tumors. A unique dataset on proteomic reprogramming by AOL and MitoTEMPO is also provided. Lastly, our study revealed the repression of NDUFV1 by S1QEL AOL. Conclusion: Our findings demonstrate the preclinical anticancer properties of S1QEL AOL and delineate its mode of action on REDOX and cancer signaling.

Differential Expression of ADP/ATP Carriers as a Biomarker of Metabolic Remodeling and Survival in Kidney Cancers.

ADP/ATP carriers (AACs) are mitochondrial transport proteins playing a strategic role in maintaining the respiratory chain activity, fueling the cell with ATP, and also regulating mitochondrial apoptosis. To understand if AACs might represent a new molecular target for cancer treatment, we evaluated AAC expression levels in cancer/normal tissue pairs available on the Tissue Cancer Genome Atlas database (TCGA), observing that AACs are dysregulated in most of the available samples. It was observed that at least two AACs showed a significant differential expression in all the available kidney cancer/normal tissue pairs. Thus, we investigated AAC expression in the corresponding kidney non-cancer (HK2)/cancer (RCC-Shaw and CaKi-1) cell lines, grown in complete medium or serum starvation, for investigating how metabolic alteration induced by different growth conditions might influence AAC expression and resistance to mitochondrial apoptosis initiators, such as "staurosporine" or the AAC highly selective inhibitor "carboxyatractyloside". Our analyses showed that AAC2 and AAC3 transcripts are more expressed than AAC1 in all the investigated kidney cell lines grown in complete medium, whereas serum starvation causes an increase of at least two AAC transcripts in kidney cancer cell lines compared to non-cancer cells. However, the total AAC protein content is decreased in the investigated cancer cell lines, above all in the serum-free medium. The observed decrease in AAC protein content might be responsible for the decrease of OXPHOS activity and for the observed lowered sensitivity to mitochondrial apoptosis induced by staurosporine or carboxyatractyloside. Notably, the cumulative probability of the survival of kidney cancer patients seriously decreases with the decrease of AAC1 expression in KIRC and KIRP tissues making AAC1 a possible new biomarker of metabolic remodeling and survival in kidney cancers.

FAD/NADH Dependent Oxidoreductases: From Different Amino Acid Sequences to Similar Protein Shapes for Playing an Ancient Function.

Flavoprotein oxidoreductases are members of a large protein family of specialized dehydrogenases, which include type II NADH dehydrogenase, pyridine nucleotide-disulphide oxidoreductases, ferredoxin-NAD+ reductases, NADH oxidases, and NADH peroxidases, playing a crucial role in the metabolism of several prokaryotes and eukaryotes. Although several studies have been performed on single members or protein subgroups of flavoprotein oxidoreductases, a comprehensive analysis on structure-function relationships among the different members and subgroups of this great dehydrogenase family is still missing. Here, we present a structural comparative analysis showing that the investigated flavoprotein oxidoreductases have a highly similar overall structure, although the investigated dehydrogenases are quite different in functional annotations and global amino acid composition. The different functional annotation is ascribed to their participation in species-specific metabolic pathways based on the same biochemical reaction, i.e., the oxidation of specific cofactors, like NADH and FADH2. Notably, the performed comparative analysis sheds light on conserved sequence features that reflect very similar oxidation mechanisms, conserved among flavoprotein oxidoreductases belonging to phylogenetically distant species, as the bacterial type II NADH dehydrogenases and the mammalian apoptosis-inducing factor protein, until now retained as unique protein entities in Bacteria/Fungi or Animals, respectively. Furthermore, the presented computational analyses will allow consideration of FAD/NADH oxidoreductases as a possible target of new small molecules to be used as modulators of mitochondrial respiration for patients affected by rare diseases or cancer showing mitochondrial dysfunction, or antibiotics for treating bacterial/fungal/protista infections.

The Bacterial Protein CNF1 as a Potential Therapeutic Strategy against Mitochondrial Diseases: A Pilot Study.

The Escherichia coli protein toxin cytotoxic necrotizing factor 1 (CNF1), which acts on the Rho GTPases that are key regulators of the actin cytoskeleton, is emerging as a potential therapeutic tool against certain neurological diseases characterized by cellular energy homeostasis impairment. In this brief communication, we show explorative results on the toxin's effect on fibroblasts derived from a patient affected by myoclonic epilepsy with ragged-red fibers (MERRF) that carries a mutation in the m.8344A>G gene of mitochondrial DNA. We found that, in the patient's cells, besides rescuing the wild-type-like mitochondrial morphology, CNF1 administration is able to trigger a significant increase in cellular content of ATP and of the mitochondrial outer membrane marker Tom20. These results were accompanied by a profound F-actin reorganization in MERRF fibroblasts, which is a typical CNF1-induced effect on cell cytoskeleton. These results point at a possible role of the actin organization in preventing or limiting the cell damage due to mitochondrial impairment and at CNF1 treatment as a possible novel strategy against mitochondrial diseases still without cure.