Complex Assembly Factor Research Articles

Geranylgeranylated-chlorophyll-protein complexes in lhl3 mutant of the green alga Chlamydomonas reinhardtii.

Chlorophylls a and b (Chl a and b) are involved in light harvesting, photochemical reactions, and electron transfer reactions in plants and green algae. The core complexes of the photosystems (PSI and PSII) associate with Chl a, while the peripheral antenna complexes (LHCI and LHCII) bind Chls a and b. One of the final steps of Chl biosynthesis is the conversion of geranylgeranylated Chls (ChlsGG) to phytylated Chls by geranylgeranyl reductase (GGR). Here, we isolated and characterized a pale green mutant of the green alga Chlamydomonas reinhardtii that was very photosensitive and was unable to grow photoautotrophically. This mutant has a 16-bp deletion in the LHL3 gene, which resulted in the loss of LHL3 and GGR and accumulated only ChlsGG. The lhl3 mutant cells grown in the dark accumulated PSII and PSI proteins at 25-50% of WT levels, lacked PSII activity, and retained a decreased PSI activity. The PSII and PSI proteins were depleted to trace amounts in the mutant cells grown in light. In contrast, the accumulation of LHCI and LHCII was unaffected except for LHCA3. Our results suggest that the replacement of Chls with ChlsGG strongly affects the structural and functional integrity of PSII and PSI complexes but their associating LHC complexes to a lesser extent. Affinity purification of HA-tagged LHL3 confirmed the formation of a stable LHL3-GGR complex, which is vital for GGR stability. The LHL3-GGR complex contained a small amount of PSI complex assembly factors, suggesting a putative coupling between Chl synthesis and PSI complex assembly.

Characterization of Shy1, the Schizosaccharomyces pombe homolog of human SURF1

Cytochrome c oxidase (complex IV) is the terminal enzyme in the mitochondrial respiratory chain. As a rare neurometabolic disorder caused by mutations in the human complex IV assembly factor SURF1, Leigh Syndrome (LS) is associated with complex IV deficiency. In this study, we comprehensively characterized Schizosaccharomyces pombe Shy1, the homolog of human SURF1. Bioinformatics analysis revealed that Shy1 contains a conserved SURF1 domain that links to the biogenesis of complex IV and shares high structural similarity with its homologs in Saccharomyces cerevisiae and humans. Our study showed that Shy1 is required for the expression of mtDNA-encoded genes and physically interacts with structural subunits and assembly factors of complex IV. Interestingly, Rip1, the subunit of ubiquinone-cytochrome c oxidoreductase or cytochrome bc1 complex (complex III), can also co-immunoprecipitate with Shy1, suggesting Shy1 may be involved in the assembly of the mitochondrial respiratory chain supercomplexes. This conclusion is further corroborated by our BN-PAGE analysis. Unlike its homologs, deletion of shy1 does not critically disrupt respiratory chain assembly, indicating the presence of the compensatory mechanism(s) within S. pombe that ensure mitochondrial functionality. Collectively, our investigation elucidates that Shy1 plays a pivotal role in the sustainability of the regular function of mitochondria by participating in the assembly of complex IV in S. pombe.

Biallelic variants in COX18 cause a mitochondrial disorder primarily manifesting as peripheral neuropathy.

Defects in mitochondrial dynamics are a common cause of Charcot-Marie-Tooth disease (CMT), while primary deficiencies in the mitochondrial respiratory chain (MRC) are rare and atypical for this etiology. This study aims to report COX18 as a novel CMT-causing gene. This gene encodes an assembly factor of mitochondrial Complex IV (CIV) that translocates the C-terminal tail of MTCO2 across the mitochondrial inner membrane. Exome sequencing was performed in four affected individuals. The patients and available family members underwent thorough neurological and electrophysiological assessment. The impact of one of the identified variants on splicing, protein levels, and mitochondrial bioenergetics was investigated in patient-derived lymphoblasts. The functionality of the mutant protein was assessed using a Proteinase K protection assay and immunoblotting. Neuronal relevance of COX18 was assessed in a Drosophila melanogaster knockdown model. Exome sequencing coupled with homozygosity mapping revealed a homozygous splice variant c.435-6A>G in COX18 in two siblings with early-onset progressive axonal sensory-motor peripheral neuropathy. By querying external databases, we identified two additional families with rare deleterious biallelic variants in COX18 . All affected individuals presented with axonal CMT and some patients also exhibited central nervous system symptoms, such as dystonia and spasticity. Functional characterization of the c.435-6A>G variant demonstrated that it leads to the expression of an alternative transcript that lacks exon 2, resulting in a stable but defective COX18 isoform. The mutant protein impairs CIV assembly and activity, leading to a reduction in mitochondrial membrane potential. Downregulation of the COX18 homolog in Drosophila melanogaster displayed signs of neurodegeneration, including locomotor deficit and progressive axonal degeneration of sensory neurons. Our study presents genetic and functional evidence that supports COX18 as a newly identified gene candidate for autosomal recessive axonal CMT with or without central nervous system involvement. These findings emphasize the significance of peripheral neuropathy within the spectrum of primary mitochondrial disorders and the role of mitochondrial CIV in the development of CMT. Our research has important implications for the diagnostic workup of CMT patients.

A Comprehensive Pan-Cancer Analysis of Cytochrome C Oxidase Assembly Factor 1 (COA1) Reveals Instrumental Role of Mitochondrial Protein Assembly in Cancer that Modulates Disease Progression and Prognostic Outcome.

Cytochrome c oxidase assembly factor 1 (COA1), a mitochondrial respiratory chain complex assembly factor protein of inner mitochondrial membrane (IMM), is involved in translating many mitochondrial components and assembling nuclear-encoded components within mitochondria. Given the lack of extensive research on COA1 in cancer, this study undertakes a comprehensive pan-cancer analysis of COA1, which is overexpressed across various cancer types, shedding light on its multifaceted role in tumorigenesis, prognosis, and tumor microenvironment (TME) modulation. Leveraging bioinformatics tools and public databases, we elucidated its potential as a diagnostic cancer biomarker as well as a target for novel anti-cancer therapeutics. Gene expression analysis using "TIMER2.0", "UALCAN" and "GEPIA2" platforms, supported by protein expression data, revealed a significant correlation between COA1 upregulation and poor prognosis in Kaplan-Meir analysis, underscoring its clinical relevance. Additionally, genetic mutation analysis of COA1 with the help of "cBioPortal" warrants further exploration into its functional significance. Moreover, our investigation of the tumor microenvironment unveiled the interplay of COA1 with fibroblast and T cell infiltration implicating the role of COA1 in the tumor immune microenvironment. Furthermore, COA1-related gene enrichment study in "GeneMANIA" and pathway cross-talk analysis with Gene Ontology (GO) gene sets established comprehensive clarifications about the molecular pathways and protein networks associated with COA1 deregulation. Overall, this study lays a sturdy foundation to support future research endeavors targeting COA1, unraveling the molecular mechanisms underlying COA1 deregulation, and exploring its therapeutic potential in cancer.

The mitochondrial tRNA-derived fragment, mt-tRF-LeuTAA, couples mitochondrial metabolism to insulin secretion

ObjectiveThe contribution of the mitochondrial electron transfer system to insulin secretion involves more than just energy provision. We identified a small RNA fragment (mt-tRF-LeuTAA) derived from the cleavage of a mitochondrially-encoded tRNA that is conserved between mice and humans. The role of mitochondrially-encoded tRNA-derived fragments remains unknown. This study aimed to characterize the impact of mt-tRF-LeuTAA, on mitochondrial metabolism and pancreatic islet functions. MethodsWe used antisense oligonucleotides to reduce mt-tRF-LeuTAA levels in primary rat and human islet cells, as well as in insulin-secreting cell lines. We performed a joint transcriptome and proteome analysis upon mt-tRF-LeuTAA inhibition. Additionally, we employed pull-down assays followed by mass spectrometry to identify direct interactors of the fragment. Finally, we characterized the impact of mt-tRF-LeuTAA silencing on the coupling between mitochondrial metabolism and insulin secretion using high-resolution respirometry and insulin secretion assays. ResultsOur study unveils a modulation of mt-tRF-LeuTAA levels in pancreatic islets in different Type 2 diabetes models and in response to changes in nutritional status. The level of the fragment is finely tuned by the mechanistic target of rapamycin complex 1. Located within mitochondria, mt-tRF-LeuTAA interacts with core subunits and assembly factors of respiratory complexes of the electron transfer system. Silencing of mt-tRF-LeuTAA in islet cells limits the inner mitochondrial membrane potential and impairs mitochondrial oxidative phosphorylation, predominantly by affecting the Succinate (via Complex II)-linked electron transfer pathway. Lowering mt-tRF-LeuTAA impairs insulin secretion of rat and human pancreatic β-cells. ConclusionsOur findings indicate that mt-tRF-LeuTAA interacts with electron transfer system complexes and is a pivotal regulator of mitochondrial oxidative phosphorylation and its coupling to insulin secretion.

Generation of a human induced pluripotent stem cell line NTUHi004-A from a patient with Leigh syndrome harboring a homozygous missense mutation c.836 T > G (p.Met279Arg) in NDUFAF5 gene

Leigh syndrome is a rare autosomal recessive disorder showcasing a diverse range of neurological symptoms. Classical Leigh syndrome is associated with mitochondrial complex I deficiency, primarily resulting from biallelic mutations in the NDUFAF5 gene, encoding the NADH:ubiquinone oxidoreductase complex assembly factor 5. Using the Sendai virus delivery system, we generated an induced pluripotent stem cell line from peripheral blood mononuclear cells of a 47-years-old female patient who carried a homozygous NDUFAF5 c.836 T > G (p.Met279Arg) mutation. This cellular model serves as a tool for investigating the underlying pathogenic mechanisms and for the development of potential treatments for Leigh syndrome.

Age-related changes in human skeletal muscle transcriptome and proteome are more affected by chronic inflammation and physical inactivity than primary aging.

Evaluation of the influence of primary and secondary aging on the manifestation of molecular and cellular hallmarks of aging is a challenging and currently unresolved issue. Our study represents the first demonstration of the distinct role of primary aging and chronic inflammation/physical inactivity - the most important drivers of secondary aging, in the regulation of transcriptomic and proteomic profiles in human skeletal muscle. To achieve this purpose, young healthy people (n = 15), young (n = 8) and older (n = 37) patients with knee/hip osteoarthritis, a model to study the effect of long-term inactivity and chronic inflammation on the vastus lateralis muscle, were included in the study. It was revealed that widespread and substantial age-related changes in gene expression in older patients relative to young healthy people (~4000 genes regulating mitochondrial function, proteostasis, cell membrane, secretory and immune response) were related to the long-term physical inactivity and chronic inflammation rather than primary aging. Primary aging contributed mainly to the regulation of genes (~200) encoding nuclear proteins (regulators of DNA repair, RNA processing, and transcription), mitochondrial proteins (genes encoding respiratory enzymes, mitochondrial complex assembly factors, regulators of cristae formation and mitochondrial reactive oxygen species production), as well as regulators of proteostasis. It was found that proteins associated with aging were regulated mainly at the post-transcriptional level. The set of putative primary aging genes and their potential transcriptional regulators can be used as a resource for further targeted studies investigating the role of individual genes and related transcription factors in the emergence of a senescent cell phenotype.

TNF-α induced NF-κB mediated LYRM7 expression modulates the tumor growth and metastatic ability in breast cancer

Tumor microenvironment (TME) of solid tumors including breast cancer is complex and contains a distinct cytokine pattern including TNF-α, which determines the progression and metastasis of breast tumors. The metastatic potential of triple negative breast cancer subtypes is high as compared to other subtypes of breast cancer. NF-κB is key a transcription factor regulating inflammation and mitochondrial bioenergetics including oxidative phosphorylation (OXPHOS) genes which determine its oxidative capacity and generating reducing equivalents for synthesis of key metabolites for proliferating breast cancer cells. The differential metabolic adaptation and OXPHOS function of breast cancer subtypes in inflammatory conditions and its contribution to metastasis is not well understood. Here we demonstrated that different subunits of NF-κB are differentially expressed in subtypes of breast cancer patients. RELA, one of the major subunits in regulation of the NF-κB pathway is positively correlated with high level of TNF-α in breast cancer patients. TNF-α induced NF-κB regulates the expression of LYRM7, an assembly factor for mitochondrial complex III. Downregulation of LYRM7 in MDA-MB-231 cells decreases mitochondrial super complex assembly and enhances ROS levels, which increases the invasion and migration potential of these cells. Further, in vivo studies using Infliximab, a monoclonal antibody against TNF-α showed decreased expression of LYRM7 in tumor tissue. Large scale breast cancer databases and human patient samples revealed that LYRM7 levels decreased in triple negative breast cancer patients compared to other subtypes and is determinant of survival outcome in patients. Our results indicate that TNF-α induced NF-κB is a critical regulator of LYRM7, a major factor for modulating mitochondrial functions under inflammatory conditions, which determines growth and survival of breast cancer cells.

Phenotypic Heterogeneity in Patients with Mutations in the Mitochondrial Complex I Assembly Gene NDUFAF5.

Rare mutations in NADH:ubiquinone oxidoreductase complex assembly factor 5 (NDUFAF5) are linked to Leigh syndrome. We aimed to describe clinical characteristics and functional findings in a patient cohort with NDUFAF5 mutations. Patients with biallelic NDUFAF5 mutations were recruited from multi-centers in Taiwan. Clinical, laboratory, radiological, and follow-up features were recorded and mitochondrial assays were performed in patients' skin fibroblasts. Nine patients from seven unrelated pedigrees were enrolled, eight homozygous for c.836 T > G (p.Met279Arg) in NDUFAF5 and one compound heterozygous for p.Met279Arg. Onset age had a bimodal distribution. The early-onset group (age <3 years) presented with psychomotor delay, seizure, respiratory failure, and hyponatremia. The late-onset group (age ≥5 years) presented with normal development, but slowly progressive dystonia. Combing 25 previously described patients, the p.Met279Arg variant was exclusively identified in Chinese ancestry. Compared with other groups, patients with late-onset homozygous p.Met279Arg were older at onset (P = 0.008), had less developmental delay (P = 0.01), less hyponatremia (P = 0.01), and better prognosis with preserved ambulatory function into early adulthood (P = 0.01). Bilateral basal ganglia necrosis was a common radiological feature, but brainstem and spinal cord involvement was more common with early-onset patients (P = 0.02). A modifier gene analysis showed higher concomitant mutation burden in early-versus late-onset p.Met279Arg homozygous cases (P = 0.04), consistent with more impaired mitochondrial function in fibroblasts from an early-onset case than a late-onset patient. The p.Met279Arg variant is a common mutation in our population with phenotypic heterogeneity and divergent prognosis based on age at onset. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Mitochondrial UQCC3 controls embryonic and tumor angiogenesis by regulating VEGF expression

Mitochondria play important roles in angiogenesis. However, the mechanisms remain elusive. In this study, we found that mitochondrial ubiquinol-cytochrome c reductase complex assembly factor 3 (UQCC3) is a key regulator of angiogenesis. TALEN-mediated knockout of Uqcc3 in mice caused embryonic lethality at 9.5-10.5days postcoitum, and vessel density was dramatically reduced. Similarly, knockout of uqcc3 in zebrafish induced lethality post-fertilization and impaired vascular development. Knockout of UQCC3 resulted in slower tumor growth and angiogenesis. Mechanistically, UQCC3 was upregulated under hypoxia, promoted reactive oxygen species (ROS) generation, enhanced HIF-1α stability and increased VEGF expression. Finally, higher expression of UQCC3 was associated with poor prognosis in multiple types tumors, implying a role for UQCC3 in tumor progression. In conclusion, our findings highlight the important contribution of UQCC3 to angiogenesis under both physiological and pathological conditions, indicating the potential of UQCC3 as a therapeutic target for cancer.

Human Tim8a, Tim8b and Tim13 are auxiliary assembly factors of mature Complex IV.

Human Tim8a and Tim8b are paralogous intermembrane space proteins of the small TIM chaperone family. Yeast small TIMs function in the trafficking of proteins to the outer and inner mitochondrial membranes. This putative import function for hTim8a and hTim8b has been challenged in human models, but their precise molecular function(s) remains undefined. Likewise, the necessity for human cells to encode two Tim8 proteins and whether any potential redundancy exists is unclear. We demonstrate that hTim8a and hTim8b function in the assembly of cytochrome c oxidase (Complex IV). Using affinity enrichment mass spectrometry, we define the interaction network of hTim8a, hTim8b and hTim13, identifying subunits and assembly factors of the Complex IV COX2 module. hTim8-deficient cells have a COX2 and COX3 module defect and exhibit an accumulation of the Complex IV S2 subcomplex. These data suggest that hTim8a and hTim8b function in assembly of Complex IV via interactions with intermediate-assembly subcomplexes. We propose that hTim8-hTim13 complexes are auxiliary assembly factors involved in the formation of the Complex IV S3 subcomplex during assembly of mature Complex IV.

Clinical phenotypic and genotypic characterization of NPRL3-related epilepsy

As one of the assembly factors of the GATOR1 protein complex in the mechanism of rapamycin pathway, NPRL3 plays an important role in the pathogenesis of epilepsy. However, the correlation between genotype and clinical phenotype in patients with NPRL3-related epilepsy has not been clarified. A total of 11 Chinese children with NPRL3-related epilepsy were identified through whole-exome sequencing (WES). The data from the clinical presentation, laboratory data, brain imaging findings, genetic results, and treatment methods were collected. All previously reported cases with NPRL3-related epilepsy were collected and reviewed through PubMed search. Among the 11 children, eight have not been reported, and two of them presented infantile spasms (ISs) as a new phenotype of NPRL3-related epilepsy. In addition, WES identified five frameshift mutations, three nonsense mutations, two missense mutations, and one exon deletion. Based on bioinformatics analysis, it was found that two missense mutation sites were highly conserved, and the c.400G>A mutation site of the NPRL3 gene caused the alteration of the protein structure. To date, 88 patients have been reported with NPRL3-related defects, including our 11 cases. The most common presentations were sleep-related hypermotor epilepsy (SHE), frontal lobe epilepsy (FLE), and temporal lobe epilepsy. A majority of patients (70%) presented normal neuroimaging results, and focal cortical dysplasia was the most common neuroimaging abnormality (62.5%). Among the NPRL3 gene mutations, loss of function (nonsense mutations, frameshift mutations, and exons deletion) was the most common genetic variation (75%). For 73% of patients with NPRL3-related epilepsy, monotherapy of sodium channel blockers was effective. Surgery was effective for 75% of children with neuroimaging abnormalities. Two cases unresponsive to surgery or anti-seizure medications were treated with ketogenic diets (KD), which were effective. One case was treated with rapamycin at an early stage of epilepsy, which was effective as well. NPRL3-related epilepsy has high clinical and genetic heterogeneity. SHE and FLE are the most common clinical presentations. Furthermore, ISs are the new phenotypes of NPRL3-related epilepsy, while the variants c.275G>A, c.745G>A, and c.1270C>T may be the most common NPRL3 gene mutations. Sodium channel blockers, surgery, KD, and rapamycin may be the potential treatments for these patients. Our study expanded the clinical and genetic spectrum of NPRL3-related epilepsy and provided important information for the precise treatment of patients.

THADA, SDHAF4, and MACF1 Gene Polymorphisms and Placental Expression in Women with Gestational Diabetes.

Gestational diabetes mellitus (GDM) is a metabolic disorder in pregnant women leading to various complications. Consequently, factors predisposing its development are being sought. Previous studies have shown that the pathogenesis of GDM is similar to that of type 2 diabetes, and it is therefore thought that the two diseases may have a common genetic basis. The aim of this study was to examine the associations between thyroid adenoma-associated (THADA) rs7578597 T&gt;C, succinate dehydrogenase complex assembly factor 4 (SDHAF4) rs1048886 A&gt;G, and microtubule-actin crosslinking factor 1 (MACF1) rs2296172 A&gt;G gene polymorphisms and the risk of GDM development as well as selected clinical parameters in women with GDM. We also examined the expression of these genes in the placenta of women with and without GDM in association with clinical parameters. This case-control study included 272 pregnant women with GDM and 348 pregnant women with normal glucose tolerance. There were no statistically significant differences in the distribution of the THADA rs7578597 T&gt;C, SDHAF4 rs1048886 A&gt;G, and MACF1 rs2296172 A&gt;G gene polymorphisms between pregnant control women and women with GDM. The associations between clinical parameters such as body mass before pregnancy, body mass at birth, body mass increase during pregnancy, BMI before pregnancy, BMI at birth, BMI increase during pregnancy, glycated hemoglobin (HbA1c), daily insulin requirement, childbirth time, and newborn body mass and APGAR score, and the THADA rs7578597 T&gt;C, SDHAF4 rs1048886 A&gt;G, and MACF1 rs2296172 A&gt;G genotypes were statistically non-significant. We only observed lower values of body mass before pregnancy and body mass at birth in women with the SDHAF4 rs1048886 AG genotype in comparison with AA genotype carriers. There was no statistically significant difference in the expression of THADA, SDHAF4, and MACF1 genes in the placenta between women with GDM and healthy women. There were also no statistically significant correlations between THADA, SDHAF4, and MACF1 gene expression in the placenta and clinical parameters. The results of our study suggest that THADA rs7578597 T&gt;C, SDHAF4 rs1048886 A&gt;G, and MACF1 rs2296172 A&gt;G gene polymorphisms are not significant factors associated with GDM onset. In addition, SDHAF4 rs1048886 A&gt;G may be associated with body mass before pregnancy and body mass at birth in pregnant women.

Analysis of Human Clinical Mutations of Mitochondrial ND1 in a Bacterial Model System for Complex I.

The most common causes of mitochondrial dysfunction and disease include mutations in subunits and assembly factors of Complex I. Numerous mutations in the mitochondrial gene ND1 have been identified in humans. Currently, a bacterial model system provides the only method for rapid construction and analysis of mutations in homologs of human ND1. In this report, we have identified nine mutations in human ND1 that are reported to be pathogenic and are located at subunit interfaces. Our hypothesis was that these mutations would disrupt Complex I assembly. Seventeen mutations were constructed in the homologous nuoH gene in an E. coli model system. In addition to the clinical mutations, alanine substitutions were constructed in order to distinguish between a deleterious effect from the introduction of the mutant residue and the loss of the original residue. The mutations were moved to an expression vector containing all thirteen genes of the E. coli nuo operon coding for Complex I. Membrane vesicles were prepared and rates of deamino-NADH oxidase activity and proton translocation were measured. Samples were also tested for assembly by native gel electrophoresis and for expression of NuoH by immunoblotting. A range of outcomes was observed: Mutations at four of the sites allow normal assembly with moderate activity (50−76% of wild type). Mutations at the other sites disrupt assembly and/or activity, and in some cases the outcomes depend upon the amino acid introduced. In general, the outcomes are consistent with the proposed pathogenicity in humans.

An MCIA-like complex is required for mitochondrial complex I assembly and seed development in maize

During adaptive radiation, mitochondria have co-evolved with their hosts, leading to gain or loss of subunits and assembly factors of respiratory complexes. Plant mitochondrial complex I harbors ∼40 nuclear- and 9 mitochondrial-encoded subunits, and is formed by stepwise assembly during which different intermediates are integrated via various assembly factors. In mammals, the mitochondrial complex I intermediate assembly (MCIA) complex is required for building the membrane arm module. However, plants have lost almost all of the MCIA complex components, giving rise to the hypothesis that plants follow an ancestral pathway to assemble the membrane arm subunits. Here, we characterize a maize crumpled seed mutant, crk1, and reveal by map-based cloning that CRK1 encodes an ortholog of human complex I assembly factor 1, zNDUFAF1, the only evolutionarily conserved MCIA subunit in plants. zNDUFAF1 is localized in the mitochondria and accumulates in two intermediate complexes that contain complex I membrane arm subunits. Disruption of zNDUFAF1 results in severe defects in complex I assembly and activity, a cellular bioenergetic shift to aerobic glycolysis, and mitochondrial vacuolation. Moreover, we found that zNDUFAF1, the putative mitochondrial import inner membrane translocase ZmTIM17-1, and the isovaleryl-coenzyme A dehydrogenase ZmIVD1 interact each other, and could be co-precipitated from the mitochondria and co-migrate in the same assembly intermediates. Knockout of either ZmTIM17-1 or ZmIVD1 could lead to the significantly reduced complex I stability and activity as well as defective seeds. These results suggest that zNDUFAF1, ZmTIM17-1 and ZmIVD1 probably form an MCIA-like complex that is essential for the biogenesis of mitochondrial complex I and seed development in maize. Our findings also imply that plants and mammals recruit MCIA subunits independently for mitochondrial complex I assembly, highlighting the importance of parallel evolution in mitochondria adaptation to their hosts.

The terminal enzymes of (bacterio)chlorophyll biosynthesis.

(Bacterio)chlorophylls are modified tetrapyrroles that are used by phototrophic organisms to harvest solar energy, powering the metabolic processes that sustain most of the life on Earth. Biosynthesis of these pigments involves enzymatic modification of the side chains and oxidation state of a porphyrin precursor, modifications that differ by species and alter the absorption properties of the pigments. (Bacterio)chlorophylls are coordinated by proteins that form macromolecular assemblies to absorb light and transfer excitation energy to a special pair of redox-active (bacterio)chlorophyll molecules in the photosynthetic reaction centre. Assembly of these pigment–protein complexes is aided by an isoprenoid moiety esterified to the (bacterio)chlorin macrocycle, which anchors and stabilizes the pigments within their protein scaffolds. The reduction of the isoprenoid ‘tail’ and its addition to the macrocycle are the final stages in (bacterio)chlorophyll biosynthesis and are catalysed by two enzymes, geranylgeranyl reductase and (bacterio)chlorophyll synthase. These enzymes work in conjunction with photosynthetic complex assembly factors and the membrane biogenesis machinery to synchronize delivery of the pigments to the proteins that coordinate them. In this review, we summarize current understanding of the catalytic mechanism, substrate recognition and regulation of these crucial enzymes and their involvement in thylakoid biogenesis and photosystem repair in oxygenic phototrophs.

Identification of a novel gene signature predicting response to first-line chemotherapy in BRCA wild-type high-grade serous ovarian cancer patients

BackgroundHigh-grade serous ovarian cancer (HGSOC) has poor survival rates due to a combination of diagnosis at advanced stage and disease recurrence as a result of chemotherapy resistance. In BRCA1 (Breast Cancer gene 1) - or BRCA2-wild type (BRCAwt) HGSOC patients, resistance and progressive disease occur earlier and more often than in mutated BRCA. Identification of biomarkers helpful in predicting response to first-line chemotherapy is a challenge to improve BRCAwt HGSOC management.MethodsTo identify a gene signature that can predict response to first-line chemotherapy, pre-treatment tumor biopsies from a restricted cohort of BRCAwt HGSOC patients were profiled by RNA sequencing (RNA-Seq) technology. Patients were sub-grouped according to platinum-free interval (PFI), into sensitive (PFI > 12 months) and resistant (PFI < 6 months). The gene panel identified by RNA-seq analysis was then tested by high-throughput quantitative real-time PCR (HT RT-qPCR) in a validation cohort, and statistical/bioinformatic methods were used to identify eligible markers and to explore the relevant pathway/gene network enrichments of the identified gene set. Finally, a panel of primary HGSOC cell lines was exploited to uncover cell-autonomous mechanisms of resistance.ResultsRNA-seq identified a 42-gene panel discriminating sensitive and resistant BRCAwt HGSOC patients and pathway analysis pointed to the immune system as a possible driver of chemotherapy response. From the extended cohort analysis of the 42 DEGs (differentially expressed genes), a statistical approach combined with the random forest classifier model generated a ten-gene signature predictive of response to first-line chemotherapy. The ten-gene signature included: CKB (Creatine kinase B), CTNNBL1 (Catenin, beta like 1), GNG11 (G protein subunit gamma 11), IGFBP7 (Insulin-like growth factor-binding protein 7), PLCG2 (Phospholipase C, gamma 2), RNF24 (Ring finger protein 24), SLC15A3 (Solute carrier family 15 member 3), TSPAN31 (Tetraspanin 31), TTI1 (TELO2 interacting protein 1) and UQCC1 (Ubiquinol-cytochrome c reductase complex assembly factor). Cytotoxicity assays, combined with gene-expression analysis in primary HGSOC cell lines, allowed to define CTNNBL1, RNF24, and TTI1 as cell-autonomous contributors to tumor resistance.ConclusionsUsing machine-learning techniques we have identified a gene signature that could predict response to first-line chemotherapy in BRCAwt HGSOC patients, providing a useful tool towards personalized treatment modalities.

Defining the interactome of the human mitochondrial ribosome identifies SMIM4 and TMEM223 as respiratory chain assembly factors.

Human mitochondria express a genome that encodes thirteen core subunits of the oxidative phosphorylation system (OXPHOS). These proteins insert into the inner membrane co-translationally. Therefore, mitochondrial ribosomes engage with the OXA1L-insertase and membrane-associated proteins, which support membrane insertion of translation products and early assembly steps into OXPHOS complexes. To identify ribosome-associated biogenesis factors for the OXPHOS system, we purified ribosomes and associated proteins from mitochondria. We identified TMEM223 as a ribosome-associated protein involved in complex IV biogenesis. TMEM223 stimulates the translation of COX1 mRNA and is a constituent of early COX1 assembly intermediates. Moreover, we show that SMIM4 together with C12ORF73 interacts with newly synthesized cytochrome b to support initial steps of complex III biogenesis in complex with UQCC1 and UQCC2. Our analyses define the interactome of the human mitochondrial ribosome and reveal novel assembly factors for complex III and IV biogenesis that link early assembly stages to the translation machinery.

Spinal Nerve Roots Abnormalities on MRI in a Child with SURF1 Mitochondrial Disease.

Variants in SURF1, encoding an assembly factor of mitochondrial respiratory chain complex IV, cause Leigh syndrome (LS) and Charcot-Marie-Tooth type 4K in children and young adolescents. Magnetic resonance imaging (MRI) appearance of enlarged nerve roots with postcontrastographic enhancement is a distinctive feature of hypertrophic neuropathy caused by onion-bulb formation and it has rarely been described in mitochondrial diseases (MDs). Spinal nerve roots abnormalities on MRI are novel findings in LS associated with variants in SURF1. Here we report detailed neuroradiological and neurophysiologic findings in a child with LS and demyelinating neuropathy SURF1-related. Our case underlines the potential contributive role of spinal neuroimaging together with neurophysiological examination to identify the full spectrum of patterns in MDs. It remains to elucidate if these observations remain peculiar of SURF1 variants or potentially detectable in other MDs with peripheral nervous system involvement.

Ndufs4 knockout mouse models of Leigh syndrome: pathophysiology and intervention.

Mitochondria are small cellular constituents that generate cellular energy (ATP) by oxidative phosphorylation (OXPHOS). Dysfunction of these organelles is linked to a heterogeneous group of multisystemic disorders, including diabetes, cancer, ageing-related pathologies and rare mitochondrial diseases. With respect to the latter, mutations in subunit-encoding genes and assembly factors of the first OXPHOS complex (complex I) induce isolated complex I deficiency and Leigh syndrome. This syndrome is an early-onset, often fatal, encephalopathy with a variable clinical presentation and poor prognosis due to the lack of effective intervention strategies. Mutations in the nuclear DNA-encoded NDUFS4 gene, encoding the NADH:ubiquinone oxidoreductase subunit S4 (NDUFS4) of complex I, induce ‘mitochondrial complex I deficiency, nuclear type 1’ (MC1DN1) and Leigh syndrome in paediatric patients. A variety of (tissue-specific) Ndufs4 knockout mouse models were developed to study the Leigh syndrome pathomechanism and intervention testing.Here, we review and discuss the role of complex I and NDUFS4 mutations in human mitochondrial disease, and review how the analysis of Ndufs4 knockout mouse models has generated new insights into the MC1ND1/Leigh syndrome pathomechanism and its therapeutic targeting.