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Molecular basis of diseases caused by the mtDNA mutation m.8969G>A in the subunit a of ATP synthase
Get PDFThe ATP synthase which provides aerobic eukaryotes with ATP, organizes into a membrane-extrinsic catalytic domain, where ATP is generated, and a membrane-embedded FO domain that shuttles protons across the membrane. We previously identified a mutation in the mitochondrial MT-ATP6 gene (m.8969G>A) in a 14-year-old Chinese female who developed an isolated nephropathy followed by brain and muscle problems. This mutation replaces a highly conserved serine residue into asparagine at amino acid position 148 of the membrane-embedded subunit a of ATP synthase. We showed that an equivalent of this mutation in yeast (aS175N) prevents FO-mediated proton translocation. Herein we identified four first-site intragenic suppressors (aN175D, aN175K, aN175I, and aN175T), which, in light of a recently published atomic structure of yeast FO indicates that the detrimental consequences of the original mutation result from the establishment of hydrogen bonds between aN175 and a nearby glutamate residue (aE172) that was proposed to be critical for the exit of protons from the ATP synthase towards the mitochondrial matrix. Interestingly also, we found that the aS175N mutation can be suppressed by second-site suppressors (aP12S, aI171F, aI171N, aI239F, and aI200M), of which some are very distantly located (by 20-30 Å) from the original mutation. The possibility to compensate through long-range effects the aS175N mutation is an interesting observation that holds promise for the development of therapeutic molecules
Methane‑yielding microbial communities processing lactate‑rich substrates: a piece of the anaerobic digestion puzzle.
Get PDFBackground: Anaerobic digestion, whose final products are methane and carbon dioxide, ensures energy flow and
circulation of matter in ecosystems. This naturally occurring process is used for the production of renewable energy from biomass. Lactate, a common product of acidic fermentation, is a key intermediate in anaerobic digestion of biomass in the environment and biogas plants. Effective utilization of lactate has been observed in many experimental approaches used to study anaerobic digestion. Interestingly, anaerobic lactate oxidation and lactate oxidizers as a physiological group in methane-yielding microbial communities have not received enough attention in the context of the acetogenic step of anaerobic digestion. This study focuses on metabolic transformation of lactate during the acetogenic and methanogenic steps of anaerobic digestion in methane-yielding bioreactors.
Results: Methane-yielding microbial communities instead of pure cultures of acetate producers were used to
process artificial lactate-rich media to methane and carbon dioxide in up-flow anaerobic sludge blanket reactors. The
media imitated the mixture of acidic products found in anaerobic environments/digesters where lactate fermentation
dominates in acidogenesis. Effective utilization of lactate and biogas production was observed. 16S rRNA profiling was
used to examine the selected methane-yielding communities. Among Archaea present in the bioreactors, the order
Methanosarcinales predominated. The acetoclastic pathway of methane formation was further confirmed by analysis
of the stable carbon isotope composition of methane and carbon dioxide. The domain Bacteria was represented by
Bacteroidetes, Firmicutes, Proteobacteria, Synergistetes, Actinobacteria, Spirochaetes, Tenericutes, Caldithrix, Verrucomicrobia, Thermotogae, Chloroflexi, Nitrospirae, and Cyanobacteria. Available genome sequences of species and/or genera identified in the microbial communities were searched for genes encoding the lactate-oxidizing metabolic machinery
homologous to those of Acetobacterium woodii and Desulfovibrio vulgaris. Furthermore, genes for enzymes of the reductive acetyl-CoA pathway were present in the microbial communities.
Conclusions: The results indicate that lactate is oxidized mainly to acetate during the acetogenic step of AD and this
comprises the acetotrophic pathway of methanogenesis. The genes for lactate utilization under anaerobic conditions
are widespread in the domain Bacteria. Lactate oxidation to the substrates for methanogens is the most energetically
attractive process in comparison to butyrate, propionate, or ethanol oxidation
Intrinsically disordered N-terminal domain of the Helicoverpa armigera Ultraspiracle stabilizes the dimeric form via a scorpion-like structure
Get PDFNuclear receptors (NRs) are a family of ligand-dependent transcription factors activated by lipophilic compounds. NRs share a common structure comprising three domains: a variable N-terminal domain (NTD), a highly conserved globular DNA-binding domain and a ligand-binding domain. There are numerous papers describing the molecular details of the latter two globular domains. However, very little is known about the structure-function relationship of the NTD, especially as an intrinsically disordered fragment of NRs that may influence the molecular properties and, in turn, the function of globular domains. Here, we investigated whether and how an intrinsically disordered NTD consisting of 58 amino acid residues affects the functions of the globular domains of the Ultraspiracle protein from Helicoverpa armigera (HaUsp). The role of the NTD was examined for two well-known and easily testable NR functions, i.e., interactions with specific DNA sequences and dimerization. Electrophoretic mobility shift assays showed that the intrinsically disordered NTD influences the interaction of HaUsp with specific DNA sequences, apparently by destabilization of HaUsp-DNA complexes. On the other hand, multi-angle light scattering and sedimentation velocity analytical ultracentrifugation revealed that the NTD acts as a structural element that stabilizes HaUsp homodimers. Molecular models based on small-angle X-ray scattering indicate that the intrinsically disordered NTD may exert its effects on the tested HaUsp functions by forming an unexpected scorpion-like structure, in which the NTD bends towards the ligand-binding domain in each subunit of the HaUsp homodimer. This structure may be crucial for specific NTD-dependent regulation of the functions of globular domains in NR
Thioloxidoreductase HP0231 of Helicobacter pylori impacts HopQ-dependent CagA translocation
Get PDFThioloxidoreductase HP0231 of Helicobacter pylori plays essential roles in gastric colonization and related gastric pathology. Comparative proteomics and analysis of complexes between HP0231 and its protein substrates suggested that several Hop proteins are its targets. HP0231 is a dimeric oxidoreductase that functions in an oxidizing Dsb (disulfide bonds) pathway of H. pylori. H. pylori HopQ possesses six cysteine residues, which generate three consecutive disulfide bridges. Comparison of the redox state of HopQ in wild-type cells to that in hp0231-mutated cells clearly indicated that HopQ is a substrate of HP0231. HopQ binds CEACAM1, 3, 5 and 6 (carcinoembryonic antigen-related cell adhesion molecules). This interaction enables T4SS-mediated translocation of CagA into host cells and induces host signaling. Site directed mutagenesis of HopQ (changing cysteine residues into serine) and analysis of the functioning of HopQ variants showed that HP0231 influences the delivery of CagA into host cells, in part through its impact on HopQ redox state. Introduction of a C382S mutation into HopQ significantly affects its reaction with CEACAM receptors, which disturbs T4SS functioning and CagA delivery. An additional effect of HP0231 on other adhesins and their redox state, resulting in their functional impairment, cannot be excluded
Advanced adenoid cystic carcinoma (ACC) is featured by SWI/SNF chromatin remodeling complex aberrations
Get PDFAbstract Purpose Adenoid cystic carcinoma (ACC) is a rare neurotropic cancer with slow progression occurring in salivary glands and less frequently in other body parts. ACC is featured by hyperchromatic nuclei and various mutations in genes encoding chromatin-related machineries. The ACC treatment is mainly limited to the radical surgery and radiotherapy while the chemotherapy remains ineffective. As the knowledge about molecular basis of ACC development is limited, we investigated here the molecular features of this disease. Patients and methods This study included 50 patients with ACC. Transcript profiling of available ACC samples vs normal salivary gland tissue, quantitative real-time PCR (qRT-PCR) transcript level measurements and the immunohistochemistry (IHC) for SWI/SNF chromatin remodeling complex (CRC) subunits and androgen receptor on surgery-derived paraffinembedded samples were performed. Results Transcriptomic study followed by Gene Ontology classification indicated alteration of chromatin-related processes, including downregulated transcript levels of main SWI/SNF CRC subunits and elevated expression of BRM ATPase-coding SMARCA2 gene in ACC. Subsequent IHC indicated broad accumulation of BRM ATPase and several SWI/SNF subunits, suggesting affected control of their protein level in ACC. The IHC revealed ectopic, heterogeneous expression of androgen receptor (AR) in some ACC cells. Conclusions Our study indicated that ACC features aberrant expression of genes controlling chromatin status and structure. We found that the balance between SWI/SNF classes is moved towards the BRM ATPase-containing complex in ACC. As BRM is known to be involved in chemoresistance in cancer cells, this observation may be the likely explanation for ACC chemoresistance
Characterization of mucus-related properties of Streptococcus thermophilus: from adhesion to induction
Get PDFMucus is a major component of the intestinal barrier involved both in the protection of the host and the fitness of commensals of the gut. Streptococcus thermophilus is consumed world-wide in fermented dairy products and is also recognized as a probiotic, as its consumption is associated with improved lactose digestion. We determined the overall effect of S. thermophilus on the mucus by evaluating its ability to adhere, degrade, modify, or induce the production of mucus and/or mucins. Adhesion was analyzed in vitro using two types of mucins (from pig or human biopsies) and mucus-producing intestinal HT29-MTX cells. The induction of mucus was characterized in two different rodent models, in which S. thermophilus is the unique bacterial species in the digestive tract or transited as a sub-dominant bacterium through a complex microbiota. S. thermophilus LMD-9 and LMG18311 strains did not grow in sugars used to form mucins as the sole carbon source and displayed weak binding to mucus/mucins relative to the highly adhesive TIL448 Lactococcus lactis. The presence of S. thermophilus as the unique bacteria in the digestive tract of gnotobiotic rats led to accumulation of lactate and increased the number of Alcian-Blue positive goblet cells and the amount of the mucus-inducer KLF4 transcription factor. Lactate significantly increased KLF4 protein levels in HT29-MTX cells. Introduction of S. thermophilus via transit as a sub-dominant bacterium (103 CFU/g feces) in a complex endogenous microbiota resulted in a slight increase in lactate levels in the digestive tract, no induction of overall mucus production, and moderate induction of sulfated mucin production. We thus show that although S. thermophilus is a poor mucus-adhesive bacterium, it can promote mucus pathway at least in part by producing lactate in the digestive tract
Grain Size Distribution of Bedload Transport in a Glaciated Catchment (Baranowski Glacier, King George Island, Western Antarctica)
Get PDFFemale Fabry disease patients and X-chromosome inactivation
Get PDFFabry disease is an X-linked inherited lysosomal storage disorder caused by mutations in the gene encoding α-
galactosidase A (GLA). Once it was thought to affect only hemizygous males. Over the last fifteen years, research
has shown that most females carrying mutated allele also develop symptoms, demonstrating a wide range of
disease severity, from a virtually asymptomatic to more classical profile, with cardiac, renal, and cerebrovascular
manifestations. This variable expression in females is thought to be influenced by the process of X-chromosome
inactivation (XCI). The aim of this study was to assess severity of the clinical phenotype, to analyze XCI patterns,
and to estimate their effect on disease manifestation in twelve female Fabry disease patients from five unrelated
Polish families. Our analyses revealed that patients presented with the broad range of disease expression - from
mild to severe, and their clinical involvement did not correlate with XCI profiles. Female carriers of the mutation
in the GLA gene with the random XCI may present with the wide range of disease signs and symptoms. Thus, XCI
is not a main factor in the phenotype variability of Fabry disease manifestation in heterozygous females
