Search Results (219)

Complete mitochondrial DNA genome annotation of an ecologically and commercially important fish species Cirrhinus cirrhosus was executed with next-generation sequencing (NGS) for nucleotide and phylogenetic analyses. The findings of this study showed that the Cirrhinus cirrhosus mitochondrial genome contained 16,593 bp, including 13 protein-coding genes, 2 ribosomal RNA genes, 22 tRNA genes, and a D-loop region. The overall base composition was 32% adenine, 25% thiamine, 16% guanine, and 27% cytosine. This mitochondrial DNA exhibits an AT biasness, with 56% AT content in its genome. Significant fluctuations were identified in the AT and GC skew values of the ND6 gene, indicating that the selection and mutation forces acting on this gene might be different from those acting on other genes. The Ka/Ks ratios of most protein-coding genes were less than 1, indicating very strong natural selection pressure. Phylogenetic analysis of Cirrhinus cirrhosus with Cirrhinus mrigala and Bangana tungting suggested a closer evolutionary relationship among these species, which might have shared a more recent common ancestor. It has been also found that the genera Labeo and Cirrhinus are not monophyletic. Full article

Paeonia ludlowii, a critically endangered species endemic to Tibet, China, possesses significant ornamental, culinary, and medicinal value. However, its mitochondrial genome remains understudied, limiting insights into its evolutionary mechanisms and constraining conservation genetics applications and molecular breeding programs. We present the first complete assembly and comprehensive analysis of the P. ludlowii mitochondrial genome. Most remarkably, we discovered that the P. ludlowii mitogenome exhibits an atypical dual-circular structure, representing the first documented occurrence of this architectural feature within the genus Paeonia. The assembled genome spans 314,371 bp and encodes 42 tRNA genes, 3 rRNA genes, and 31 protein-coding genes, with a pronounced adenine–thymine bias. This multipartite genome structure is characterized by abundant repetitive elements (112 functionally annotated SSRs, 33 tandem repeats, and 945 dispersed repeats), which potentially drive genome rearrangements and facilitate adaptive evolution. Analyses of codon usage bias and nucleotide diversity revealed highly conserved gene expression regulation with limited variability. Phylogenetic reconstruction confirms that P. ludlowii, P. suffruticosa, and P. lactiflora form a monophyletic clade, reflecting close evolutionary relationships, while extensive syntenic collinearity with other Paeonia species underscores mitochondrial genome conservation at the genus level. Extensive inter-organellar gene transfer events, particularly from chloroplast to mitochondrion, suggest that such DNA exchanges enhance genetic diversity and promote environmental adaptation. The discovery of the dual-circular architecture provides novel insights into plant mitochondrial genome evolution and structural plasticity. This study elucidates the unique structural characteristics of the P. ludlowii mitochondrial genome and establishes a crucial genetic foundation for developing targeted conservation strategies and facilitating molecular-assisted breeding programs for this endangered species. Full article

Methylation is a biochemical process involving the addition of methyl groups to proteins, lipids, and nucleic acids (both DNA and RNA). DNA methylation predominantly occurs on cytosine and adenine nucleobases, and the resulting products—most frequently 5-methylcytosine and N⁶-methyladenine epigenetic marks—can significantly influence gene activity at the affected genomic sites without modifying the DNA sequence called nucleotide order. Various environmental factors can alter the DNA methylation pattern. Among these, methyl donor micronutrients, such as specific amino acids, choline, and several B vitamins (including folate, pyridoxine, thiamine, riboflavin, niacin, and cobalamin), primarily regulate one-carbon metabolism. This molecular pathway stimulates glutathione synthesis and recycles intracellular methionine. Glutathione plays a pivotal role during oocyte activation by protecting against oxidative stress, whereas methionine is crucial for the production of S-adenosyl-L-methionine, which serves as the universal direct methyl donor for cellular methylation reactions. Because local DNA methylation patterns at genes regulating fertility can be inherited by progeny for multiple generations even in the absence of the original disrupting factors to which the parent was exposed, and DNA methylation levels at specific genomic sites highly correlate with age and can also be passed to offspring, nutrition can influence reproduction and life span in a transgenerational manner. Full article

Introduction: Postoperative atrial fibrillation (POAF) is a common complication after heart surgery, adversely impacting clinical outcomes and healthcare costs. Little is known about the dynamics of nucleotide metabolism associated with the development of POAF at a ventricular level. We conducted a post hoc trial analysis to investigate the changes in ventricular adenine nucleotides and the clinical predictors associated with the development of AF. Methods: Using data from a randomised trial, we analysed ATP/ADP, ATP/AMP, and energy charges in left and right ventricular biopsies of patients who developed AF compared to non-AF patients. A logistic regression model was used to understand the predictors associated with the development of atrial fibrillation in this cohort. Results: We analysed adenine nucleotide levels available in 88 patients who underwent coronary artery bypass grafting (CABG) (n = 65) and aortic valve replacement (AVR) (n = 23), out of which 27 (31%) developed a new onset of AF. Seventeen (43.4%) patients in the CABG group and ten (26.15%) in the AVR group developed AF. The patients who developed postoperative AF had longer cross-clamp times for CABG (p = 0.013) and AVR (p = 0.002). The most significant predictors for AF development were age (p = 0.003) and cross-clamp time (p = 0.012). In patients undergoing CABG who developed AF, we found a significant drop in post-reperfusion ATP/ADP and ATP/AMP ratios compared to pre-reperfusion. This was not significant for the patients who underwent AVR. Furthermore, the patients who underwent CABG and developed AF had higher pre- and post-reperfusion ATP/ADP ratios and energy charges than non-AF patients, suggesting a higher reserve of cardiac nucleotides. Conclusions: The development of postoperative atrial fibrillation is associated with intraoperative changes in the ventricular adenine nucleotide metabolism of patients undergoing CABG. In the clinical analysis, age and cross-clamp time were significant predictors of AF development. Full article

Next to adrenergic signalling, purinergic pathways mediated by extracellular adenine nucleotides have been described to shape thermogenic and metabolic functions in brown adipose tissue (BAT). Here we describe high expression of P2X5 that is activated by ATP in mature adipocytes of BAT and differentiated brown adipocytes in vitro. The levels of other P2X family members were much lower, or expression was restricted to tissue-resident macrophages or endothelial cells. Global and brown adipocyte-specific P2rx5 deficiency resulted in lower expression of the uncoupling protein 1 (UCP1). However, indirect calorimetry studies showed that P2X5 did not affect systemic energy expenditure. Of note, glucose tolerance was impaired under chow and obesogenic high-fat diet conditions, which can be explained by lower glucose disposal into BAT but not into other organs. In summary, these data indicate a modulatory role of P2X5 in systemic and BAT-specific glucose metabolism. Full article

Background: Tariquidar (Tq) is an inhibitor of the multidrug resistance (MDR) proteins relevant to ATP-binding cassette transporters (ABC transporters), which suppresses the ATP-dependent efflux of a variety of hydrophilic and amphipathic compounds, including anticancer drugs. Tq is a representative of a new generation of MDR inhibitors with high affinity to ABC proteins. However, there are still no data on the possible effect of Tq on mitochondria as an important target in the regulation of cell death or survival. Methods: We investigated the influence of Tq on the Ca²⁺-dependent mitochondrial permeability transition pore (mPTP). The effect of Tq was assessed using several parameters, including the calcium load, membrane potential, and mitochondrial swelling. To evaluate the specific targets of Tq, selective inhibitors of components of the mitochondrial pore were used, including adenine nucleotides, carboxyatractylozide (Catr) and bongkrekic acid (BA), oligomycin, and cyclosporine A. Results: Tq decreased the calcium retention capacity, activated mitochondrial swelling, and lowered the influence of ADP and ATP, the inhibitors of the Ca²⁺-induced pore opening, at their low concentrations. These effects of Tq were observed in both calcium-load and swelling assays, thus mimicking the effect of Catr, a selective inhibitor of adenine nucleotide translocase (ANT). Tq also decreased the protective effect of BA, an inhibitor of ANT and mPTP, on the calcium retention capacity of mitochondria. Further, Tq dose-dependently decreased the inhibitory effect of a low ATP concentration but not of high concentrations, at which the effect of Tq was activated by oligomycin, an inhibitor of F-ATP synthase. Conclusions: The influence of Tq extends to mitochondria, specifically to the regulation of membrane permeability, promoting the activation of pore opening, probably through an interaction with ANT, a component of the pore-forming complex. The effect of Tq on the opening of mPTP is strongly dependent on the concentrations of adenine nucleotides and, consequently, on the functional state of mitochondria. The direct influence of Tq on mitochondria can be considered as a new activity that promotes the sensitization of cells to various treatments and stimuli. Full article

Although Cervus elaphus (Linnaeus, 1758) has been well studied, the subspecific taxonomy of Cervus canadensis populations in Qinghai and Gansu, China, is still controversial, and the mitochondrial characteristics of Cervus elaphus (Linnaeus, 1758) remain incompletely understood. We assembled 89 mitogenomes of C. canadensis from five geographical populations across Qinghai and Gansu. Phylogenetic analysis confirmed that the 89 individuals are taxonomically classified as C. c. kansuensis. Nucleotide compositions showed a higher abundance of adenine and cytosine compared to guanine and thymine in both complete mitogenomes and mitochondrial PCGs. Codon usage analysis revealed a strong preference towards A-ending codons (68.04% of over-represented codons, RSCU > 1.6) in mitochondrial PCGs, with systemic avoidance of G-ending codons (53.30% of unused codons, RSCU = 0). The CAMs of 13 PCGs are reported for the first time. Furthermore, the ENC plot showed that the codon usage of all PCGs was biased except for gene ATP8. The PR2 bias plot showed that gene ND6 exhibited bias towards T3 and G3, whereas the other genes preferred A3 and C3. Both the ENC-plot and PR2 bias plot suggested that natural selection played an important role in the forces driving codon usage bias in mitochondrial PCGs. Our results demonstrate the subspecific status of C. canadensis distributed in Qinghai and Gansu as C. c. kansuensis, and provide insights into the mitochondrial characteristics of C. c. kansuensis. The mitogenome sequences assembled in this study provide valuable data for further understanding of the Cervus elaphus (Linnaeus, 1758) mitogenome. Full article

We explored how the phosphorylation state of collapsin response mediator protein 2 (CRMP2) influences mitochondrial functions in cultured cortical neurons and cortical synaptic mitochondria isolated from APP-SAA KI mice, a knock-in APP mouse model of Alzheimer’s disease (AD). CRMP2 phosphorylation was increased at Thr 509/514 and Ser 522 in brain cortical lysates and cultured neurons from AD mice. The basal and maximal respiration of AD neurons were decreased. Mitochondria were hyperpolarized and superoxide anion production was increased in neurons from AD mice. In isolated synaptic AD mitochondria, ADP-stimulated and DNP-stimulated respiration were decreased, whereas ADP-induced mitochondrial depolarization was reduced and prolonged. We found that CRMP2 binds to the adenine nucleotide translocase (ANT) in a phosphorylation-dependent manner. The increased CRMP2 phosphorylation in AD mice correlated with CRMP2 dissociation from the ANT and decreased ANT activity in AD mitochondria. On the other hand, recombinant CRMP2 (rCRMP2), added to the ANT-reconstituted proteoliposomes, increased ANT activity. A small molecule (S)-lacosamide ((S)-LCM), which binds to CRMP2 and suppresses CRMP2 phosphorylation by Cdk5 and GSK-3β, prevented CRMP2 hyperphosphorylation, rescued CRMP2 binding to the ANT, improved ANT activity, and restored the mitochondrial membrane potential and respiratory responses to ADP and 2,4-dinitrophenol. Thus, our study highlights an important role for CRMP2 in regulating the mitochondrial oxidative metabolism in AD by modulating the ANT activity in a phosphorylation-dependent manner. Full article

Integration of old and recent experimental data consequences is needed to correct and help improve the hypothetical mechanism responsible for the stimulus–secretion coupling mechanism of glucose-induced insulin secretion. The main purpose of this review is to supply biochemical considerations about some of the metabolic pathways implicated in the process of insulin secretion. It is emphasized that glucose β-cells’ threshold to activate secretion (5 mM) might depend on the predominance of anaerobic glycolysis at this basal glucose concentration. This argues against the predominance of phosphoenolpyruvate (PEP) over mitochondrial pyruvate oxidation for the initiation of insulin secretion. Full quantitative and qualitative reproduction, except the threshold effect, of glucose-induced insulin release by a permeable methylated analog of succinic acid indicates that mitochondrial metabolism is enough for sustained insulin secretion. Mitochondrial PEP generation is skipped if the GABA-shunt pathway is exclusively coupled to the citric acid cycle, as proposed in the “GABA-shunt” model of stimulus–secretion coupling. Strong or maintained depolarization by KCl or sulfonylureas might induce the opening of β-cells Cx36 hemichannels, allowing the loss of adenine nucleotides and other metabolites, mimicking the effect of an excessive mitochondrial ATP demand. A few alterations of OxPhos (Oxidative Phosphorylation) regulation in human T2D islets have been described, but the responsible mechanism(s) is (are) not yet known. Finally, some experimental data arguing as proof of the relative irrelevance of the mitochondrial function in the insulin secretion coupling mechanism for the initiation and/or sustained stimulation of hormone release are discussed. Full article

Iodosulfonylation of an ethynyl group at the C2 position of 2′-deoxyadenosine or adenosine with TsI provides (E)-2-(β-iodovinyl)sulfones. The latter undergo nucleophilic substitution with amines via an addition–elimination to give β-sulfonylvinylamines (enamines). Acid-catalyzed hydrolysis of the β-sulfonylvinylamines provides 2-(β-keto)sulfones, mechanistically different probes that react with alkyl halides, resulting in α-alkylation. Adenine nucleosides with a β-ketosulfone group at C2, during conversion to their 5′-triphosphate form, undergo an unexpected conversion to 2-carboxylic acid nucleotides. The 5′-triphosphate of 2′-deoxyadenosine-2-carboxylic acid was incorporated by a human DNA polymerase into a one-nucleotide gap DNA substrate. Full article

Mitochondrial calcium (Ca²⁺) uptake plays a key role in mitochondrial physiology and disease development. This process is regulated by the mitochondrial calcium uniporter (MCU) complex. DS16570511 is a membrane-permeable drug that inhibits mitochondrial Ca²⁺ uptake, although its inhibitory mechanisms remain unclear. In this study, we evaluated the effects of DS16570511 on various mitochondrial functions through biochemical analyses. We found that DS16570511 affects multiple mitochondrial functions and exhibits variable potency in inhibiting individual processes. Specifically, DS16570511 not only inhibits MCU, its initially reported target, but also respiratory chain complexes and F_oF₁-adenosine triphosphatase/adenine nucleotide translocator, particularly respiratory chain complex II. Furthermore, the carboxyl group at the molecular terminus of DS16570511 plays a critical role in its inhibitory effects on mitochondrial Ca²⁺ uptake through respiratory chain complex II inhibition. These findings enhance our understanding of the mechanisms by which DS16570511 inhibits mitochondrial Ca²⁺ uptake and provide valuable insights for the clinical application of mitochondrial Ca²⁺ uptake inhibitors. Full article

Precisely fluorescently labeling specific nucleotide sites of RNA is critical for gaining insights into the structure and function of RNA through multiple fluorescence detection techniques. The position-selective labeling of RNA (PLOR) method provides a promising strategy to achieve this, wherein the fluorophore-modified NTPs can be co-transcriptionally introduced to specific sites of nascent RNA by using T7 RNA polymerase (T7 RNAP). However, due to steric hindrance limitations, the efficiency of T7 RNAP in recognizing and incorporating large fluorophore-modified NTPs into RNA is far from satisfactory. To overcome this challenge, in this work, we developed an efficient PLOR variant (ePLOR) for the site-specific fluorescent labeling of RNA by integrating PLOR with a post-transcriptional SPAAC (strain-promoted azido-alkyne cycloaddition) click chemistry reaction. The efficiency of the SPAAC reaction occurring on RNA is nearly 100%. Consequently, ePLOR enables the precise fluorescent labeling of designated sites across various structural regions of SAM-VI riboswitch and adenine riboswitch RNA, with labeling and synthesis efficiencies that are 2–2.5 times higher than those of PLOR. The strategy developed in this work can be used for the efficient synthesis of a broader spectrum of long-strand RNAs with site-specific fluorescent labeling and greatly facilitate the detection of the structure and function of these RNAs. Full article

Background/Objectives: Some G-quadruplex (G4)-forming nucleic acid sequences bind a hemin cofactor to enhance its peroxidase-like activity. This has been implemented in a variety of bioanalytical assays benefiting from analyte-dependent peroxidation of a chromogenic organic substrate (e.g., ABTS) to produce a color change. Adenine and cytosine nucleotides in the vicinity of the G4 hemin-binding site promote the peroxidation reaction. In this work, the effect of G4 loop and flanking nucleotides on the colorimetric signal of split hybridization probes utilizing hemin-G4 signal reporters was tested. Methods: G4s varying by loop sequences and flanking nucleotides were tested with hemin for ABTS peroxidation (A₄₂₀), and the signal was compared with that produced by the most catalytically efficient complexes reported in the literature using one-way ANOVA and post hoc pairwise comparison with Tukey’s HSD test. The best G4s were used as signal transducers in the split peroxidase deoxyribozyme (sPDz) probes for sensing two model nucleic acid analytes, as well as in a cascade system, where the analyte-dependent assembly of an RNA-cleaving deoxyribozyme 10–23 results in G4 release. Results: Intramolecular G4s (G₃T)₃G₃TC or G₃T₃G₃ATTG₃T₃G₃ were found to be the most efficient hemin PDzs. When splitting intramolecular G4 for the purpose of sPDz probe design, the addition of a flanking d(TC) sequence at one of the G4 halves or d(ATT) in a loop connecting the second and third G-tracts helps boost analyte-dependent signal intensity. However, for the cascade system, the effect of d(TC) or d(ATT) in the released G4 was not fully consistent with the data reported for intramolecular G4-hemin complexes. Conclusions: Our findings offer guidance on the design of split hybridization probes utilizing the peroxidase-like activity of G4-hemin complexes as a signal transducer. Full article

(1) Background: The insect mitogenome encodes essential genetic components and serves as an effective marker for molecular identification and phylogenetic analysis in insects due to its small size, maternal inheritance, and rapid evolution. The morphological identification of Ablabesmyia is challenging, particularly for non-experts. Thus, there is an increasing need for molecular data to improve classification accuracy and phylogenetic analysis. (2) Methods: Our analysis encompassed eight species of Ablabesmyia, a single species of Conchapelopia, one species of Denopelopia, and one species of Thienemannimyia, all originating from China. We then performed a comprehensive analysis of the nucleotide composition, sequence length, and evolutionary rate. (3) Results: All newly assembled mitogenomes displayed a negative GC-skew, indicating a cytosine bias, while most exhibited a positive AT-skew, reflecting an adenine and thymine abundance. All thirteen protein-coding genes (PCGs) featured the conventional start codon ATN, aligning closely with the typical mitochondrial start codon observed in insects. The evolutionary rates of these PCGs can be ordered as follows: ND2 > ATP8 > ND6 > ND4 > ND5 > ND3 > ND4L > ND1 > CYTB > COIII > ATP6 > COII > COI. (4) Conclusions: These newly sequenced mitogenomes exhibit structural features and nucleotide compositions that closely align with those of previously reported Chironomidae species, marking a significant expansion of the chironomid mitogenome database. Full article

The pentose phosphate pathway (PPP), traditionally recognized for its role in generating nicotinamide adenine dinucleotide phosphate (NADPH) and ribose-5-phosphate (R5P), has emerged as a critical metabolic hub with involvements in various gastrointestinal (GI) cancers. The PPP plays crucial roles in the initiation, development, and tumor microenvironment (TME) of GI cancers by modulating redox homeostasis and providing precursors for nucleotide biosynthesis. Targeting PPP enzymes and their regulatory axis has been a potential strategy in anti-GI cancer therapies. In this review, we summarize the regulatory mechanisms of PPP enzymes, elucidate the relationships between the PPP and TME’s elements, and discuss the therapeutic potential of targeting the PPP in GI cancers. Full article