Role In Oxidative Phosphorylation Research Articles

Mitochondrial proteins as therapeutic targets in diabetic ketoacidosis: evidence from Mendelian randomization analysis.

Diabetic ketoacidosis (DKA) is a severe and potentially fatal acute complication in diabetic patients, commonly occurring in type 1 diabetes (T1D) but also seen in type 2 diabetes (T2D). The pathogenesis of DKA involves complex physiological processes that are not fully understood, especially the role of mitochondria. Mitochondria, known as the powerhouse of cells, plays a crucial role in oxidative phosphorylation and ATP production, which is vital in various metabolic diseases, including diabetes. However, the exact causal relationship between mitochondrial dysfunction and DKA remains unclear. This study employed Mendelian randomization (MR) analysis and protein-protein interaction (PPI) networks to systematically explore the causal relationships between mitochondrial DNA copy number (mtDNA-CN) and specific mitochondrial proteins with DKA. We used bidirectional MR analysis and genome-wide association study (GWAS) data from openGWAS database to investigate the causal effects of mtDNA-CN and 64 mitochondrial-related proteins on DKA and its subtypes (T1DKA, T2DKA, unspecified-DKA). The study revealed that increased mtDNA-CN significantly reduces the risk of DKA, whereas the effect of DKA on mtDNA-CN was not significant. Mitochondrial-related proteins such as MRPL32, MRPL33, COX5B, DNAJC19, and NDUFB8 showed a negative causal relationship with DKA, indicating their potential protective roles. Conversely, ATP5F1B and COX4I2 have a positive causal relationship with DKA, indicating that excessive ATP production in diabetic patients may be detrimental to health and increase the risk of severe complications such as DKA. The results emphasize the necessity of protecting mitochondrial function in order to reduce the risk of DKA. The study offers novel perspectives on the molecular pathways involved in DKA, emphasizing the critical functions of mt-DNA and distinct proteins. These evidences not only enhance our comprehension of the implications of mitochondrial dysfunction in diabetes-related complications but also identify potential therapeutic targets for individualized treatment approaches, thereby making a substantial contribution to clinical care and public health initiatives.

A novel HMGA2/MPC-1/mTOR signaling pathway promotes cell growth via facilitating Cr (VI)-induced glycolysis

Mitochondrial Pyruvate Carrier 1 (MPC1) is localized on mitochondrial outer membrane to mediate the transport of pyruvate from cytosol to mitochondria. It is also well known to act as a tumor suppressor. Hexavalent chromium (Cr (VI)) contamination poses a global challenge due to its high toxicity and carcinogenesis. This research was intended to probe the potential mechanism of MPC1 in the effect of Cr (VI)-induced carcinogenesis. First, Cr (VI)-treatments decreased the expression of MPC1 in vitro and in vivo. Overexpression of MPC1 inhibited Cr (VI)-induced glycolysis and migration in A549 cells. Then, high mobility group A2 (HMGA2) protein strongly suppressed the transcription of MPC1 by binding to its promoter, and HMGA2/MPC1 axis played an important role in oxidative phosphorylation (OXPHOS), glycolysis and cell migration. Furthermore, endoplasmic reticulum (ER) stress made a great effect on the interaction between HMGA2 and MPC1. Finally, the mammalian target of the rapamycin (mTOR) was determined to mediate MPC1-regulated OXPHOS, aerobic glycolysis and cell migration. Collectively, our data revealed a novel HMGA2/MPC-1/mTOR signaling pathway to promote cell growth via facilitating the metabolism reprogramming from OXPHOS to aerobic glycolysis, which might be a potential therapy for cancers.

Role of oxidative phosphorylation in the antidepressant effects of arketamine via the vagus nerve-dependent spleen-brain axis

Arketamine, the (R)-enantiomer of ketamine, exhibits antidepressant-like effects in mice, though the precise molecular mechanisms remain elusive. It has been shown to reduce splenomegaly and depression-like behaviors in the chronic social defeat stress (CSDS) model of depression. This study investigated whether the spleen contributes to the antidepressant-like effects of arketamine in the CSDS model. We found that splenectomy significantly inhibited arketamine's antidepressant-like effects in CSDS-susceptible mice. RNA-sequencing analysis identified the oxidative phosphorylation (OXPHOS) pathway in the prefrontal cortex (PFC) as a key mediator of splenectomy's impact on arketamine's effects. Furthermore, oligomycin A, an inhibitor of the OXPHOS pathway, reversed the suppressive effects of splenectomy on arketamine's antidepressant-like effects. Specific genes within the OXPHOS pathways, such as COX11, UQCR11 and ATP5e, may contribute to these inhibitory effects. Notably, transforming growth factor (TGF)-β1, along with COX11, appears to modulate the suppressive effects of splenectomy and contribute to arketamine's antidepressant-like effects. Additionally, SRI-01138, an agonist of the TGF-β1 receptor, alleviated the inhibitory effects of splenectomy on arketamine's antidepressant-like effects. Subdiaphragmatic vagotomy also counteracted the inhibitory effects of splenectomy on arketamine's antidepressant-like effects in CSDS-susceptible mice. These findings suggest that the OXPHOS pathway and TGF-β1 in the PFC play significant roles in the antidepressant-like effects of arketamine, mediated through the spleen-brain axis via the vagus nerve.

Abstract B009: Isolating human RCC tumor cells details evolutionary relationships between cancer cell subsets and a role of OXPHOS in disease progression

Abstract In this study, we aimed to examine cancer-cell intrinsic features that might dictate clear cell renal cell carcinoma (ccRCC) progression. However, cellular heterogeneity within the tumor tissue complicates the analysis of pure kidney cancer cells. Therefore, we performed scRNA-seq of the whole tumor tissue to profile gene expression patterns unique to cancer cells. Next, using this information, we developed a novel FACS sorting strategy and sorted two populations: CD24loENPP3lo (CD24lo) and CD24hiENPP3hi (CD24hi). Whole genome sequencing on the isolated subsets from over 100 patients confirmed >85% tumor purity. Direct comparison with public TCGA data demonstrated that cancer cell enrichment prior to sequencing significantly increased mutation detection sensitivity highlighting the advantage of this approach. Genomic and phenotypic lineage analysis revealed a progressive relationship between CD24lo precursors and CD24hi differentiated cells. Apart from many shared clonal mutations, we detected selected subclonal events in the CD24lo that became clonal in CD24hi. CD24hi cells were characterized by highly malignant properties such as epithelial to mesenchymal transition and an increased proliferative state as measured by Ki67. In accordance with their invasive signature, increased CD24hi frequencies appeared in advanced but not early stages (T1-2) of ccRCC. Importantly, not all late-stage patients with a CD24hi population progressed after surgery. To examine the qualities of CD24hi cells that associate with disease recurrence, we performed RNA-seq on purified cancer cells from progressors or non-progressors. We found a strong link between disease progression and increased oxidative phosphorylation (OXPHOS) in CD24hi cells. Furthermore, to validate our RNA-seq we assessed mitochondrial fitness using the TMRM probe and detected increased mitochondrial potential within CD24hicells from progressors. To explore the intrinsic consequences of increased cancer cell oxidative metabolism, we examined the relationship between proliferation and OXPHOS in CD24hi cells but did not observe any significant correlations. Extrinsically, to evaluate OXPHOS effects on the tumor microenvironment we examined CD8 T cells given that increased infiltration confers a prognostic advantage in ccRCC. We found that elevated tumor oxidative metabolism inversely associated with CD8 T cells numbers and response to immunotherapy, suggesting a role for OXPHOS in anti-tumor immune control. Overall, this work used a novel human cancer cell isolation approach to identify a cancer cell differentiation path from a CD24lo precursor to an aggressive CD24hi cell. Presence alone of the CD24hi does not predict survival, instead, the oxidative dominance within these cells negatively impacts disease control. Interestingly, metabolic focus within ccRCC has overlooked OXPHOS, but we provide insight that it could be leveraged for unexplored treatment avenues in ccRCC. Citation Format: Ewelina Sobierajska, Christopher Medina, Petra Gregorova, BaoHan Vo, Rachel Greenwald, Caroline Jansen, Christian Larsen, Viraj Master, Haydn Kissick. Isolating human RCC tumor cells details evolutionary relationships between cancer cell subsets and a role of OXPHOS in disease progression [abstract]. In: Proceedings of the AACR Special Conference: Advances in Kidney Cancer Research; 2023 Jun 24-27; Austin, Texas. Philadelphia (PA): AACR; Cancer Res 2023;83(16 Suppl):Abstract nr B009.

Epitope-level profiling in children with mitochondrial disease reveals limitations in the antibacterial antibody repertoire.

Immunometabolic studies in mice have suggested the importance of oxidative phosphorylation (OXPHOS) in humoral immunity. However, there are important distinctions between murine and human immunity. Furthermore, translational studies on the role of OXPHOS in humoral immunity are nearly absent from the biomedical literature. Children with primary OXPHOS deficiency (i.e., mitochondrial disease, MtD), are an important patient population for demonstrating the functional effects of this bioenergetic defect on humoral immunity. To define whether OXPHOS deficiency extended to human B cells, we performed extracellular flux analysis on lymphoblastoid B cell lines from children with MtD and controls (N=4/group). To expand the immune phenotype of B cell OXPHOS deficiency, we conducted a cross-sectional multiplex serology study of the antibacterial antibody repertoire in children with MtD (N=16) and controls (N=16) using phage display and immunoprecipitation sequencing (PhIPseq). The PhIPseq library contained >3000 peptides (i.e., epitopes) covering >40 genera and>150 species of bacteria that infect humans. B cell lymphoblastoid cell lines from children with MtD displayed depressed baseline oxygen consumption, ATP production and reserve capacity, indicating that OXPHOS deficiency extended to these key cells in humoral immunity. Characterization of the bacterial exposome revealed comparable bacterial species between the two groups, mostly Streptococcus and Staphylococcus. The most common species of bacteria was S. pneumoniae. By interrogating the antibacterial antibody repertoire, we found that children with MtD had less robust antibody fold changes to common epitopes. Furthermore, we also found that children with MtD failed to show a direct relationship between the number of bacterial epitopes recognized and age, unlike controls. OXPHOS deficiency extends to B cells in children with MtD, leading to limitations in the antibacterial antibody repertoire. Furthermore, the timing of bacterial exposures was asynchronous, suggesting different periods of increased exposure or susceptibility. Overall, the antibacterial humoral response is distinctive in children with MtD, suggesting an important role for OXPHOS in B cell function.

A Mitochondria-Targeted Ferroptosis Inducer Activated by Glutathione-Responsive Imaging and Depletion for Triple Negative Breast Cancer Theranostics.

Ferroptosis is a new type of iron-dependent programmed cell death characterized by glutathione (GSH) depletion, selenoprotein glutathione peroxidase 4 (GPX4) inactivation, and lipid peroxides accumulation. Mitochondria, as the main source of intracellular energy supply and reactive oxygen species (ROS) generation, play a central role in oxidative phosphorylation and redox homeostasis. Therefore, targeting cancer-cell mitochondria and attacking redox homeostasis is expected to induce robust ferroptosis-mediated anticancer effects. In this work, a theranostic ferroptosis inducer (IR780-SPhF), which can simultaneously achieve the imaging and therapy of triple-negative breast cancer (TNBC) by targeting mitochondria is presented. It is developed from a mitochondria-targeting small molecule (IR780) with cancer-preferential accumulation, enabling it to react with GSH by nucleophilic substitution, resulting in mitochondrial GSH depletion and redox imbalance. More interestingly, IR780-SPhF exhibits GSH-responsive near-infrared fluorescence emission and photoacoustic imaging characteristics, further facilitating diagnosis and treatment with real-time monitoring of TNBC with a highly elevated GSH level. Both in vitro and in vivo results demonstrate that IR780-SPhF exhibits potent anticancer effect, which is significantly stronger than cyclophosphamide, a classic drug commonly recommended for TNBC patients in clinic. Hence, the reported mitochondria-targeted ferroptosis inducer may represent a promising candidate and a prospective strategy for efficient cancer treatment.

PHB2 promotes colorectal cancer cell proliferation and tumorigenesis through NDUFS1-mediated oxidative phosphorylation

The alteration of cellular energy metabolism is a hallmark of colorectal cancer (CRC). Accumulating evidence has suggested oxidative phosphorylation (OXPHOS) is upregulated to meet the demand for energy in tumor initiation and development. However, the role of OXPHOS and its regulatory mechanism in CRC tumorigenesis and progression remain unclear. Here, we reveal that Prohibitin 2 (PHB2) expression is elevated in precancerous adenomas and CRC, which promotes cell proliferation and tumorigenesis of CRC. Additionally, knockdown of PHB2 significantly reduces mitochondrial OXPHOS levels in CRC cells. Meanwhile, NADH:ubiquinone oxidoreductase core subunit S1 (NDUFS1), as a PHB2 binding partner, is screened and identified by co-immunoprecipitation and mass spectrometry. Furthermore, PHB2 directly interacts with NDUFS1 and they co-localize in mitochondria, which facilitates NDUFS1 binding to NADH:ubiquinone oxidoreductase core subunit V1 (NDUFV1), regulating the activity of complex I. Consistently, partial inhibition of complex I activity also abrogates the increased cell proliferation induced by overexpression of PHB2 in normal human intestinal epithelial cells and CRC cells. Collectively, these results indicate that increased PHB2 directly interacts with NDUFS1 to stabilize mitochondrial complex I and enhance its activity, leading to upregulated OXPHOS levels, thereby promoting cell proliferation and tumorigenesis of CRC. Our findings provide a new perspective for understanding CRC energy metabolism, as well as novel intervention strategies for CRC therapeutics.

EP14.02-006 Subtype-specific Hypersensitivity to Oxidative Phosphorylation Inhibition in Small Cell Lung Cancer

Small cell lung cancer (SCLC) is a very aggressive type of lung cancer with a poor prognosis due to a high proliferation rate and early metastasis. The current standard-of-care therapeutic treatment, a combination of a platinum-based chemotherapy and etoposide, does not take the four molecular subtypes (SCLC-A, -N, -P and -Y) into account. Oxidative phosphorylation (OXPHOS) is an integral part of cellular metabolism and is frequently altered in cancer. However, not much is known about the role of OXPHOS in SCLC.

A homozygous splice variant in ATP5PO, disrupts mitochondrial complex V function and causes Leigh syndrome in two unrelated families.

Mitochondrial complex V plays an important role in oxidative phosphorylation by catalyzing the generation of ATP. Most complex V subunits are nuclear encoded and not yet associated with recognized Mendelian disorders. Using exome sequencing, we identified a rare homozygous splice variant (c.87+3A>G) in ATP5PO, the complex V subunit which encodes the oligomycin sensitivity conferring protein, in three individuals from two unrelated families, with clinical suspicion of a mitochondrial disorder. These individuals had a similar, severe infantile and often lethal multi-systemic disorder that included hypotonia, developmental delay, hypertrophic cardiomyopathy, progressive epileptic encephalopathy, progressive cerebral atrophy, and white matter abnormalities on brain MRI consistent with Leigh syndrome. cDNA studies showed a predominant shortened transcript with skipping of exon 2 and low levels of the normal full-length transcript. Fibroblasts from the affected individuals demonstrated decreased ATP5PO protein, defective assembly of complex V with markedly reduced amounts of peripheral stalk proteins, and complex V hydrolytic activity. Further, expression of human ATP5PO cDNA without exon 2 (hATP5PO-∆ex2) in yeast cells deleted for yATP5 (ATP5PO homolog) was unable to rescue growth on media which requires oxidative phosphorylation when compared to the wild type construct (hATP5PO-WT), indicating that exon 2 deletion leads to a non-functional protein. Collectively, our findings support the pathogenicity of the ATP5PO c.87+3A>G variant, which significantly reduces but does not eliminate complex V activity. These data along with the recent report of an affected individual with ATP5PO variants, add to the evidence that rare biallelic variants in ATP5PO result in defective complex V assembly, function and are associated with Leigh syndrome.

Identification of Novel Cytochrome C1 (CYC1) Gene Expression in Oral Squamous Cell Carcinoma- An Evaluative Study.

Cytochrome C1 (CYC1) is an important subunit of mitochondrial complex III and plays a vital role in oxidative phosphorylation (OXPHOS) and reactive oxygen species generation. Overexpression of the CYC1 gene has been implicated in cancer development and its prognosis previously, but unexplored in head-and-neck squamous cell carcinomas (HNSCC), especially oral squamous cell carcinoma (OSCC). CYC1 m-RNA expression and gene alterations were assessed using the Cancer Genome Atlas dataset in HNSCC and validated in OSCC tissues using real-time polymerase chain reaction (RT-PCR). The protein-protein interaction (PPI) network and functional enrichment pathways were also analysed. A thorough analysis of the TCGA (The Cancer Genome Atlas) database revealed that CYC1 was overexpressed in the HNSCC cases and the increased expression correlated with several parameters which involve the prediction of advanced diseases such as histopathological grade, tumour-node-metastasis staging, and nodal metastases (P < 0.05). The expression of CYC1 was validated using RT-PCR showing significant upregulation (P < 0.05) in OSCC tissue samples compared to the normal tissue counterparts. PPI network and functional analysis show the prominent role of CYC1 in OXPHOS, especially in electron transport chain III complex regulation. The study revealed that CYC1 is highly expressed in HNSCC, and is validated in the OSCC patient tissue samples compared to the normal counterparts and associated with advanced disease stages and grade of the tumour. CYC1 could be a novel promising therapeutic and prognostic marker in HNSCC, especially in OSCC.

Gold(III)-P-chirogenic complex induces mitochondrial dysfunction in triple-negative breast cancer

SummaryChemical agents that specifically exploit metabolic vulnerabilities of cancer cells will be beneficial but are rare. The role of oxidative phosphorylation (OXPHOS) in promoting and maintaining triple-negative breast cancer (TNBC) growth provides new treatment opportunity. In this work, we describe AuPhos-19, a small-molecule gold(III)-based agent bearing a chiral phosphine ligand that selectively disrupts mitochondrial metabolism in murine and human TNBC cells but not normal epithelial cells. AuPhos-19 induces potent cytotoxic effect with half maximal inhibitory concentration (IC50) in the nanomolar range (220–650 nM) across different TNBC cell lines. The lipophilic cationic character of AuPhos-19 facilitates interaction with mitochondrial OXPHOS. AuPhos-19 inhibits mitochondria respiration and induces significant AMPK activation. Depolarization of the mitochondria membrane, mitochondria ROS accumulation, and mitochondria DNA depletion provided further indication that AuPhos-19 perturbs mitochondria function. AuPhos-19 inhibits tumor growth in tumor-bearing mice. This study highlights the development of gold-based compounds targeting mitochondrial pathways for efficacious cancer treatment.

Regulation of PGC1α Downstream of the Insulin Signaling Pathway Plays a Role in the Hepatic Proteotoxicity of Mutant α1-Antitrypsin Deficiency Variant Z

Insulin signaling is known to regulate essential proteostasis mechanisms. The analyses here examined effects of insulin signaling in the PiZ mouse model of α1-antitrypsin deficiency in which hepatocellular accumulation and proteotoxicity of the misfolded α1-antitrypsin Z variant (ATZ) causes liver fibrosis and cancer. We first studied the effects of breeding PiZ mice to liver-insulin-receptor knockout (LIRKO) mice (with hepatocyte-specific insulin-receptor gene disruption). The results showed decreased hepatic ATZ accumulation and liver fibrosis in PiZ x LIRKO vs PiZ mice, with reversal of those effects when we bred PiZ x LIRKO mice onto a FOXO1-deficient background. Increased intracellular degradation of ATZ mediated by autophagy was identified as the likely mechanism for diminished hepatic proteotoxicity in PiZ x LIRKO mice and the converse was responsible for enhanced toxicity in PiZ x LIRKO x FOXO1-KO animals. Transcriptomic studies showed major effects on oxidative phosphorylation and autophagy genes, and significant induction of peroxisome proliferator-activated-receptor-γ-coactivator-1α (PGC1α) expression in PiZ-LIRKO mice. Because PGC1α plays a key role in oxidative phosphorylation, we further investigated its effects on ATZ proteostasis in our ATZ-expressing mammalian cell model. The results showed PGC1α overexpression or activation enhances autophagic ATZ degradation. These data implicate suppression of autophagic ATZ degradation by down-regulation of PGC1α as one mechanism by which insulin signaling exacerbates hepatic proteotoxicity in PiZ mice, and identify PGC1α as a novel target for development of new human α1-antitrypsin deficiency liver disease therapies.

Mitochondrial respiration supports autophagy to provide stress resistance during quiescence

ABSTRACT Mitochondrial oxidative phosphorylation (OXPHOS) generates ATP, but OXPHOS also supports biosynthesis during proliferation. In contrast, the role of OXPHOS during quiescence, beyond ATP production, is not well understood. Using mouse models of inducible OXPHOS deficiency in all cell types or specifically in the vascular endothelium that negligibly relies on OXPHOS-derived ATP, we show that selectively during quiescence OXPHOS provides oxidative stress resistance by supporting macroautophagy/autophagy. Mechanistically, OXPHOS constitutively generates low levels of endogenous ROS that induce autophagy via attenuation of ATG4B activity, which provides protection from ROS insult. Physiologically, the OXPHOS-autophagy system (i) protects healthy tissue from toxicity of ROS-based anticancer therapy, and (ii) provides ROS resistance in the endothelium, ameliorating systemic LPS-induced inflammation as well as inflammatory bowel disease. Hence, cells acquired mitochondria during evolution to profit from oxidative metabolism, but also built in an autophagy-based ROS-induced protective mechanism to guard against oxidative stress associated with OXPHOS function during quiescence. Abbreviations: AMPK: AMP-activated protein kinase; AOX: alternative oxidase; Baf A: bafilomycin A1; CI, respiratory complexes I; DCF-DA: 2′,7′-dichlordihydrofluorescein diacetate; DHE: dihydroethidium; DSS: dextran sodium sulfate; ΔΨmi: mitochondrial inner membrane potential; EdU: 5-ethynyl-2’-deoxyuridine; ETC: electron transport chain; FA: formaldehyde; HUVEC; human umbilical cord endothelial cells; IBD: inflammatory bowel disease; LC3B: microtubule associated protein 1 light chain 3 beta; LPS: lipopolysaccharide; MEFs: mouse embryonic fibroblasts; MTORC1: mechanistic target of rapamycin kinase complex 1; mtDNA: mitochondrial DNA; NAC: N-acetyl cysteine; OXPHOS: oxidative phosphorylation; PCs: proliferating cells; PE: phosphatidylethanolamine; PEITC: phenethyl isothiocyanate; QCs: quiescent cells; ROS: reactive oxygen species; PLA2: phospholipase A2, WB: western blot.

Evaluation of the Ankylosing Spondylitis Transcriptome for Oxidative Phosphorylation Pathway: The Shared Pathway with Neurodegenerative Diseases.

Ankylosing spondylitis (AS) is a systemic inflammatory disorder of joints and entheses. Recent studies have reported an increased prevalence of dementia in AS patients. However, data for exploring the association between dementia and AS remain uncertain. In this study, enriched pathways and differentially expressed genes (DEGs) were identified in whole blood transcription data of AS patients obtained from the gene expression omnibus (GEO) database; using gene set enrichment analysis (GSEA) and differential expression analysis. Four pathways, including oxidative phosphorylation, Alzheimer's, Parkinson's, and Huntington's diseases were significantly enriched in AS patients compared to the controls. We identified 22 common genes among the pathways that showed an increasing trend in AS compared to the controls. Five of them including COX7B, NDUFB3, ATP5PF, UQCRB, and NDUFS4 were the most significant genes which were selected for gene expression analysis; using real-time PCR on RNA contents of peripheral blood mononuclear cells (PBMCs) of AS patients and controls (20 samples from each group). The gene expression analysis indicated considerable overexpression of COX7B (p<0.0001) and ATP5J (p=0.0001) genes in AS patients group in comparison to the control samples. The role of oxidative phosphorylation has previously been established in dementia pathogenesis. Given that AS patients have also a remarkably higher prevalence of dementia than the their healthy counterparts, hence our results may propose that the common pathway of oxidative phosphorylation can be regarded as a possible shared contributing factor in the etiopathogenesis of AS and dementia.

Abstract 2844: The role of oxidative phosphorylation (OXPHOS) in melanoma brain metastasis (MBM) formation and growth

Abstract Introduction: MBMs are a deadly complication of melanoma. We previously showed that OXPHOS is increased in MBMs, but the roles of OXPHOS in MBM formation and growth are unknown. Methods: To determine if OXPHOS predicts formation of MBMs, RNA-sequencing (RNAseq) was performed on primary melanomas from patients (pts) diagnosed with MBM (P_MBM; n=19) and from pts diagnosed with metastases (mets) solely to non-CNS sites within 3-18 months of initial diagnosis (P_ECM; n=16), as well as pts who did not develop any mets within 5 years (P_NoMet; n=19). To functionally test the role of OXPHOS in brain and lung met formation and maintenance, brain-tropic YUMM5.2-Br3 cells were treated in vitro with the direct OXPHOS inhibitor IACS-010759 (100 nM) or vehicle control for 12 hours, and then injected into the left ventricle (intracardiac; ICa) in C57BL/6 mice. Mice were randomized to in vivo treatment with vehicle or IACS-010759 (7.5 mg/kg daily) for 14 days. Mice were euthanized and ex vivo bioluminescence imaging (BLI) was performed to evaluate the extent of brain and lung mets. We also evaluated A375P and G361 human melanoma cell lines stably transfected with shRNAs targeting the OXPHOS mediator PGC1α (shPGC1α) or scrambled control (shScr). Cells were injected ICa, intracranially (ICr) or subcutaneously (SQ) in CD-1 nude mice. Lung and brain tissues were harvested for histological analysis of mets once mice became moribund. SQ tumors were measured twice weekly to assess tumor growth. Mice with ICr injection were monitored for survival and euthanized once moribund. Results: Pathway analysis of the primary tumors from pts demonstrated elevated KEGG OXPHOS gene set expression in P_MBM and P_ECM groups compared to P_NoMet (FDR q-val= 0.048 and &amp;lt;0.001, respectively), suggesting a role for OXPHOS in metastasis. However, there was no significant enrichment of OXPHOS genes in P_MBM vs. P_ECM (FDR q-val=0.516), indicating a lack of specific association with MBM risk. Treatment with IACS-010759 (in vitro and/or in vivo) in the YUMM5.2-Br3 model did not impact the incidence of lung or brain mets. However, in vivo treatment with IACS-010759 significantly decreased MBM (but not lung) tumor burden. PGC1α knockdown (shPGC1α) reduced OXPHOS in A375P and G361 compared to shScr, but less than IACS-010759 treatment. shPGC1α did not impact lung or brain met formation or burden following ICa injection of A375P and G361 cells. shPGC1α also did not inhibit tumor incidence with SQ or ICr injection, and there was no significant difference in SQ tumor growth. However, shPGC1α significantly prolonged survival of mice injected ICr with A375P (p=0.0003) and G361 (p=0.0097). Conclusions: OXPHOS was associated with increased risk of mets in pts, but not specifically MBMs. In mice, OXPHOS inhibition did not prevent MBM formation, but reduced the growth of established MBMs. Thus, OXPHOS is critical to MBM maintenance. Citation Format: Grant M. Fischer, Renato A. Guerrieri, Qianghua Hu, Fernando C. Carapeto, Aron Y. Joon, Vashisht Y. Gopal, Barbara Knighton, Wanleng Deng, Lauren E. Haydu, Jeffrey E. Gershenwald, Alexander J. Lazar, Michael T. Tetzlaff, Michael A. Davies. The role of oxidative phosphorylation (OXPHOS) in melanoma brain metastasis (MBM) formation and growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2844.

Genetic basis of negative heterosis for growth traits in chickens revealed by genome-wide gene expression pattern analysis

BackgroundHeterosis is an important biological phenomenon that has been extensively utilized in agricultural breeding. However, negative heterosis is also pervasively observed in nature, which can cause unfavorable impacts on production performance. Compared with systematic studies of positive heterosis, the phenomenon of negative heterosis has been largely ignored in genetic studies and breeding programs, and the genetic mechanism of this phenomenon has not been thoroughly elucidated to date. Here, we used chickens, the most common agricultural animals worldwide, to determine the genetic and molecular mechanisms of negative heterosis.ResultsWe performed reciprocal crossing experiments with two distinct chicken lines and found that the body weight presented widely negative heterosis in the early growth of chickens. Negative heterosis of carcass traits was more common than positive heterosis, especially breast muscle mass, which was over − 40% in reciprocal progenies. Genome-wide gene expression pattern analyses of breast muscle tissues revealed that nonadditivity, including dominance and overdominace, was the major gene inheritance pattern. Nonadditive genes, including a substantial number of genes encoding ATPase and NADH dehydrogenase, accounted for more than 68% of differentially expressed genes in reciprocal crosses (4257 of 5587 and 3617 of 5243, respectively). Moreover, nonadditive genes were significantly associated with the biological process of oxidative phosphorylation, which is the major metabolic pathway for energy release and animal growth and development. The detection of ATP content and ATPase activity for purebred and crossbred progenies further confirmed that chickens with lower muscle yield had lower ATP concentrations but higher hydrolysis activity, which supported the important role of oxidative phosphorylation in negative heterosis for growth traits in chickens.ConclusionsThese findings revealed that nonadditive genes and their related oxidative phosphorylation were the major genetic and molecular factors in the negative heterosis of growth in chickens, which would be beneficial to future breeding strategies.

Mitochondrial genome stability in human: understanding the role of DNA repair pathways.

Mitochondria are semiautonomous organelles in eukaryotic cells and possess their own genome that replicates independently. Mitochondria play a major role in oxidative phosphorylation due to which its genome is frequently exposed to oxidative stress. Factors including ionizing radiation, radiomimetic drugs and replication fork stalling can also result in different types of mutations in mitochondrial DNA (mtDNA) leading to genome fragility. Mitochondria from myopathies, dystonia, cancer patient samples show frequent mtDNA mutations such as point mutations, insertions and large-scale deletions that could account for mitochondria-associated disease pathogenesis. The mechanism by which such mutations arise following exposure to various DNA-damaging agents is not well understood. One of the well-studied repair pathways in mitochondria is base excision repair. Other repair pathways such as mismatch repair, homologous recombination and microhomology-mediated end joining have also been reported. Interestingly, nucleotide excision repair and classical nonhomologous DNA end joining are not detected in mitochondria. In this review, we summarize the potential causes of mitochondrial genome fragility, their implications as well as various DNA repair pathways that operate in mitochondria.

A Re-Assessment of Positive Selection on Mitochondrial Genomes of High-Elevation Phrynocephalus Lizards.

Due to their integral roles in oxidative phosphorylation, mitochondrially encoded proteins represent common targets of selection in response to altitudinal hypoxia across high-altitude taxa. While previous studies revealed evidence of positive selection on mitochondrial genomes of high-altitude Phrynocephalus lizards, their conclusions were restricted by out-of-date phylogenies and limited taxonomic sampling. Using topologies derived from both nuclear and mitochondrial DNA phylogenies, we re-assessed the evidence of positive selection on the mitochondrial genomes of high-altitude Phrynocephalus. We sampled representative species from all four main lineages and sequenced the mitochondrial genome of P. maculatus, a putative sister taxon to the high-altitude group. Positive selection was assessed through two widely used branch-site tests: the branch-site model in PAML and BUSTED in HyPhy. No evidence of positive selection on mitochondrial genes was detected on branches leading to two most recent common ancestors of high-altitude species; however, we recovered evidence of positive selection on COX1 on the P. forsythii branch, which represents a reversal from high- to low-elevation environments. A positively selected site therein marked a threonine to valine substitution at position 419. We suggest this bout of selection occurred as the ancestors of P. forsythii re-colonized lower altitude environments north of the Tibetan Plateau. Despite their role in oxidative phosphorylation, we posit that mitochondrial genes are unlikely to have represented historical targets of selection for high-altitude adaptation in Phrynocephalus. Consequently, future studies should address the roles of nuclear genes and differential gene expression.

The LncRNA FEZF1-AS1 promotes tumor proliferation in colon cancer by regulating the mitochondrial protein PCK2.

LncRNAs and metabolism represents two factors involved in cancer initiation and progression. However, the interaction between lncRNAs and metabolism remains to be fully explored. In this study, lncRNA FEZF1-AS1 (FEZF1-AS1) was found upregulated in colon cancer after screening all the lncRNAs of colon cancer tissues deposited in TCGA, the result of which was further confirmed by RNAscope staining on a colon tissue chip. The results obtained using FEZF1-AS1 knockout colon cancer cells (SW480 KO and HCT-116 KO) constructed using CRISPR/Cas9 system confirmed the proliferation, invasion, and migration-promoting function of FEZF1-AS1 in vitro. Mechanistically, FEZF1-AS1 associated with the mitochondrial protein phosphoenolpyruvate carboxykinase (PCK2), which plays an essential role in regulating energy metabolism in the mitochondria. Knockdown of FEZF1-AS1 greatly decreased PCK2 protein levels, broke the homeostasis of energy metabolism in the mitochondria, and inhibited proliferation, invasion, and migration of SW480 and HCT-116 cells. PCK2 overexpression in FEZF1-AS1 knockout cells partially rescued the tumor inhibitory effect on colon cancer cells both in vitro and in vivo. Moreover, PCK2 overexpression specifically rescued the abnormal accumulation of Flavin mononucleotide (FMN) and succinate, both of which play an important role in oxidative phosphorylation (OXPHOS). Overall, these results indicate that FEZF1-AS1 is an oncogene through regulating energy metabolism of the cell. This research reveals a new mechanism for lncRNAs to regulate colon cancer and provides a potential target for colon cancer diagnosis and treatment.

Mitochondrial Oxidative Stress Induces Rapid Intermembrane Space/Matrix Translocation of Apurinic/Apyrimidinic Endonuclease 1 Protein through TIM23 Complex

Mitochondria are essential cellular organelles that import the majority of proteins to sustain their function in cellular metabolism and homeostasis. Due to their role in oxidative phosphorylation, mitochondria are constantly affected by oxidative stress. Stability of mitochondrial DNA (mtDNA) is essential for mitochondrial physiology and cellular well-being and for this reason mtDNA lesions have to be rapidly recognized and repaired. Base excision repair (BER) is the main pathway responsible for repairing non-helix distorting base lesions both into the nucleus and in mitochondria. Apurinic/Apyrimidinic Endonuclease 1 (APE1) is a key component of BER pathway and the only protein that can recognize and process an abasic (AP) site. Comprehensions of the mechanisms regulating APE1 intracellular trafficking are still fragmentary. In this study we focused our attention on the mitochondrial form of APE1 protein and how oxidative stress induces its translocation to maintain mtDNA integrity. Our data proved that: (i) the rise of mitochondrial ROS determines a very rapid translocation of APE1 from the intermembrane space (IMS) into the matrix; and (ii) TIM23/PAM machinery complex is responsible for the matrix translocation of APE1. Moreover, our data support the hypothesis that the IMS, where the majority of APE1 resides, could represent a sort of storage site for the protein.