Developing tardigrade-inspired material: Track membranes functionalized with Dsup protein for cell-free DNA isolation.

When developing functionalized biomaterials, the proteins from extremophilic organisms, in particular unique tardigrade disordered proteins, are of great value. The damage suppressor protein (Dsup), initially discovered in the tardigrade Ramazzottius varieornatus and found to be an efficient DNA protector under oxidative and irradiation stress, has been hypothesized to possess a good potential for the development of the material, which can isolate cell-free DNA. With this in mind, DNA-nonadsorbing polyethylene terephthalate track membranes have been functionalized using the Dsup protein via covalent bonding with glutaraldehyde. The filtration experiments have verified the ability of track membranes with the immobilized Dsup protein to adsorb cell-free DNA, with an accumulation capacity of 70 ± 19 mg m-2. The resulting track membrane-based biomaterial might be used in various devices for filtration and separation of cell-free DNA molecules from biological solutions and environmental samples, and also for their accumulation, storage, and further manipulation.

The tardigrade Dsup protein enhances radioresistance in Drosophila melanogaster and acts as an unspecific repressor of transcription.

The tardigrade-unique damage suppressor protein (Dsup) can protect DNA from ionizing radiation and reactive oxygen species (ROS). In this study, we generated Dsup-expressing lines of Drosophila melanogaster and demonstrated that Dsup increased the survival rate after γ-ray irradiation and hydrogen peroxide treatment in flies too, but reduced the level of their locomotor activity. The transcriptome analyses of Dsup-expressing lines revealed a significant number of DEGs, >99% of which were down-regulated. Moreover, Dsup could bind RNA. These findings suggest that Dsup can act not only as a DNA protector but also as a non-specific transcriptional repressor and RNA-binding protein, that may lead to disturbance of a number of biological processes in D. melanogaster. The obtained data demonstrate features of the Dsup protein action in non-tardigrade organisms and can be used to understand the impact of other unspecific DNA/RNA-binding proteins on ROS and radiation resistance, gene expression, and epigenetic processes.

Cytobacillus pseudoceanisediminis sp. nov., A Novel Facultative Methylotrophic Bacterium with High Heavy Metal Resistance Isolated from the Deep Underground Saline Spring.

In this study, we present the characterization of the BNO1T bacterial strain isolated from the deep subsurface saline spring at the Baksan Neutrino Observatory INR RAS (Kabardino-Balkaria, Russia). The complete genome sequence of the strain BNO1T is 5,347,902 bp, with a GC content 41 and 49%. The cell wall peptidoglycan contains meso-diaminopimelic acid. The major isoprenoid quinone is MK-7 and the polar lipids are diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine. The major fatty acids are anteiso-C15:0 (23.34%), iso-C15:0 (20.10%), C16:0 (11.96%), iso-C16:0 (10.88%), and anteiso-C17:0 (10.79%). The 16S rRNA gene sequence clearly demarcated the strain as belonging to Cytobacillus genera. Based on the phylogenetic analysis, ANI (average nucleotide identity) and dDDH (digital DNA-DNA hybridization) assessments we propose to assign the strain BNO1T and other related strains to new species and to name it Cytobacillus pseudoceanisediminis sp. nov. (The values of ANI and dDDH between BNO1T and Cytobacillus oceanisediminis CGMCC 1.10115 T are 80.65% and 24.7%, respectively; values of ANI and dDDH between BNO1T and Cytobacillus firmus NCTC 10335 T are 89% and 38%, respectively). Genomic analysis of strain BNO1T revealed pathways for C1 compounds oxidation and two pathways for C1 compounds assimilation: serine and ribulose monophosphate pathways. In addition, strain BNO1T contains a plasmid (342,541 bp) coding multiple genes involved in heavy metal ion balance. Moreover, heavy metal toxicity testing confirmed the high potential of the strain BNO1T as a source of metal resistance genes and enzymes. The type strain is BNO1T (= BIM B-1921 T = VKM B-3664 T).

First transcriptome profiling of D. melanogaster after development in a deep underground low radiation background laboratory.

Natural background radiation is a permanent multicomponent factor. It has an influence on biological organisms, but effects of its deprivation still remain unclear. The aim of our work was to study for the first time responses of D. melanogaster to conditions of the Deep Underground Low-Background Laboratory DULB-4900 (BNO, INR, RAS, Russia) at the transcriptome level by RNA-seq profiling. Overall 77 transcripts demonstrated differential abundance between flies exposed to low and natural background radiation. Enriched biological process functional categories were established for all genes with differential expression. The results showed down-regulation of primary metabolic processes and up-regulation of both the immune system process and the response to stimuli. The comparative analysis of our data and publicly available transcriptome data on D. melanogaster exposed to low and high doses of ionizing radiation did not reveal common DEGs in them. We hypothesize that the observed changes in gene expression can be explained by the influence of the underground conditions in DULB-4900, in particular, by the lack of stimuli. Thus, our study challenges the validity of the LNT model for the region of background radiation doses below a certain level (~16.4 nGy h-1) and the presence of a dose threshold for D. melanogaster.

Transcriptome analysis of Drosophila melanogaster laboratory strains of different geographical origin after long-term laboratory maintenance.

Positive selection may be the main factor of the between-population divergence in gene expression. Expression profiles of two Drosophila melanogaster laboratory strains of different geographical origin and long-term laboratory maintenance were analyzed using microchip arrays encompassing probes for 18,500 transcripts. The Russian strain D18 and the North American strain Canton-S were compared. A set of 223 known or putative genes demonstrated significant changes in expression levels between these strains. Differentially expressed genes (DEG) were enriched in response to DDT (p = .0014), proteolysis (p = 2.285E-5), transmembrane transport (p = 1.03E-4), carbohydrate metabolic process (p = .0317), protein homotetramerization (p = .0444), and antibacterial humoral response (p = 425E-4). The expression in subset of genes from different categories was verified by qRT-PCR. Analysis of transcript abundance between Canton-S and D18 strains allowed to select several genes to estimate their participation in latitude adaptation. Expression of selected genes was analyzed in five D. melanogaster lines of different geographic origins by qRT-PCR, and we found two candidate genes that may be associated with latitude adaptation in adult flies-smp-30 and Cda9. Quite possible that several alleles of these genes may be important for insect survival in the environments of global warming. It is interesting that the number of genes involved in local adaptation demonstrates expression level appropriate to their geographical origin even after decades of laboratory maintenance.

The Oxidation-Induced Autofluorescence Hypothesis: Red Edge Excitation and Implications for Metabolic Imaging.

Endogenous autofluorescence of biological tissues is an important source of information for biomedical diagnostics. Despite the molecular complexity of biological tissues, the list of commonly known fluorophores is strictly limited. Still, the question of molecular sources of the red and near-infrared excited autofluorescence remains open. In this work we demonstrated that the oxidation products of organic components (lipids, proteins, amino acids, etc.) can serve as the molecular source of such red and near-infrared excited autofluorescence. Using model solutions and cell systems (human keratinocytes) under oxidative stress induced by UV irradiation we demonstrated that oxidation products can contribute significantly to the autofluorescence signal of biological systems in the entire visible range of the spectrum, even at the emission and excitation wavelengths higher than 650 nm. The obtained results suggest the principal possibility to explain the red fluorescence excitation in a large class of biosystems-aggregates of proteins and peptides, cells and tissues-by the impact of oxidation products, since oxidation products are inevitably presented in the tissue. The observed fluorescence signal with broad excitation originated from oxidation products may also lead to the alteration of metabolic imaging results and has to be taken into account.