Quantitative Proteomic Analysis of Bacillus pumilus from Spores that Survived in Outer Space Conditions

The hardy spores of Bacillus species are well known for their resistance to unfavorable conditions such as UV and gamma radiation, hydrogen peroxide desiccation, chemical disinfection, or starvation. In particular, Bacillus pumilus strain SAFR-032 that was originally recovered from the Jet Propulsion Lab Spacecraft Assembly Facility has shown to exhibit unusually high resistance to UV radiation and peroxide treatment compared to other Bacillus species. To further understand the resistance of bacterial endospores to relevant outer space environments, spores of B. pumilus SAFR-032 were exposed for 1.5 years to selected parameters of space on board of the International Space Station (ISS). Here we are using a quantitative proteomics approach to gain insights into the resistance mechanism of B. pumilus

Proteomic and Metabolomic Characteristics of Extremophilic Fungi Under Simulated Mars Conditions

Filamentous fungi have been associated with extreme habitats, including nuclear power plant accident sites and the International Space Station (ISS). Due to their immense adaptation and phenotypic plasticity capacities, fungi may thrive in what seems like uninhabitable niches. This study is the first report of fungal survival after exposure of monolayers of conidia to simulated Mars conditions (SMC). Conidia of several Chernobyl nuclear accident-associated and ISS-isolated strains were tested for UV-C and SMC sensitivity, which resulted in strain-dependent survival. Strains surviving exposure to SMC for 30 min, ISSFT-021-30 and IMV 00236-30, were further characterized for proteomic, and metabolomic changes. Differential expression of proteins involved in ribosome biogenesis, translation, and carbohydrate metabolic processes was observed. No significant metabolome alterations were revealed. Lastly, ISSFT-021-30 conidia re-exposed to UV-C exhibited enhanced UV-C resistance when compared to the conidia of unexposed ISSFT-021

Effects of Simulated Microgravity on the Proteome and Secretome of the Polyextremotolerant Black Fungus Knufia chersonesos.

Black fungi are a group of melanotic microfungi characterized by remarkable polyextremotolerance. Due to a broad ecological plasticity and adaptations at the cellular level, it is predicted that they may survive in a variety of extreme environments, including harsh niches on Earth and Mars, and in outer space. However, the molecular mechanisms aiding survival, especially in space, are yet to be fully elucidated. Based on these premises, the rock-inhabiting black fungus Knufia chersonesos (Wt) and its non-melanized mutant (Mut) were exposed to simulated microgravity-one of the prevalent features characterizing space conditions-by growing the cultures in high-aspect-ratio vessels (HARVs). Qualitative and quantitative proteomic analyses were performed on the mycelia and supernatant of culture medium (secretome) to assess alterations in cell physiology in response to low-shear simulated microgravity (LSSMG) and to ultimately evaluate the role of cell-wall melanization in stress survival. Differential expression was observed for proteins involved in carbohydrate and lipid metabolic processes, transport, and ribosome biogenesis and translation via ribosomal translational machinery. However, no evidence of significant activation of stress components or starvation response was detected, except for the scytalone dehydratase, enzyme involved in the synthesis of dihydroxynaphthalene (DNH) melanin, which was found to be upregulated in the secretome of the wild type and downregulated in the mutant. Differences in protein modulation were observed between K. chersonesos Wt and Mut, with several proteins being downregulated under LSSMG in the Mut when compared to the Wt. Lastly, no major morphological alterations were observed following exposure to LSSMG. Similarly, the strains' survivability was not negatively affected. This study is the first to characterize the response to simulated microgravity in black fungi, which might have implications on future astrobiological missions

Effects of Simulated Microgravity on the Proteome and Secretome of the Polyextremotolerant Black Fungus Knufia chersonesos.

Black fungi are a group of melanotic microfungi characterized by remarkable polyextremotolerance. Due to a broad ecological plasticity and adaptations at the cellular level, it is predicted that they may survive in a variety of extreme environments, including harsh niches on Earth and Mars, and in outer space. However, the molecular mechanisms aiding survival, especially in space, are yet to be fully elucidated. Based on these premises, the rock-inhabiting black fungus Knufia chersonesos (Wt) and its non-melanized mutant (Mut) were exposed to simulated microgravity-one of the prevalent features characterizing space conditions-by growing the cultures in high-aspect-ratio vessels (HARVs). Qualitative and quantitative proteomic analyses were performed on the mycelia and supernatant of culture medium (secretome) to assess alterations in cell physiology in response to low-shear simulated microgravity (LSSMG) and to ultimately evaluate the role of cell-wall melanization in stress survival. Differential expression was observed for proteins involved in carbohydrate and lipid metabolic processes, transport, and ribosome biogenesis and translation via ribosomal translational machinery. However, no evidence of significant activation of stress components or starvation response was detected, except for the scytalone dehydratase, enzyme involved in the synthesis of dihydroxynaphthalene (DNH) melanin, which was found to be upregulated in the secretome of the wild type and downregulated in the mutant. Differences in protein modulation were observed between K. chersonesos Wt and Mut, with several proteins being downregulated under LSSMG in the Mut when compared to the Wt. Lastly, no major morphological alterations were observed following exposure to LSSMG. Similarly, the strains' survivability was not negatively affected. This study is the first to characterize the response to simulated microgravity in black fungi, which might have implications on future astrobiological missions

Characterization of Aspergillus niger Isolated from the International Space Station.

The initial characterization of the Aspergillus niger isolate JSC-093350089, collected from U.S. segment surfaces of the International Space Station (ISS), is reported, along with a comparison to the extensively studied strain ATCC 1015. Whole-genome sequencing of the ISS isolate enabled its phylogenetic placement within the A. niger/welwitschiae/lacticoffeatus clade and revealed that the genome of JSC-093350089 is within the observed genetic variance of other sequenced A. niger strains. The ISS isolate exhibited an increased rate of growth and pigment distribution compared to a terrestrial strain. Analysis of the isolate's proteome revealed significant differences in the molecular phenotype of JSC-093350089, including increased abundance of proteins involved in the A. niger starvation response, oxidative stress resistance, cell wall modulation, and nutrient acquisition. Together, these data reveal the existence of a distinct strain of A. niger on board the ISS and provide insight into the characteristics of melanized fungal species inhabiting spacecraft environments. IMPORTANCE A thorough understanding of how fungi respond and adapt to the various stimuli encountered during spaceflight presents many economic benefits and is imperative for the health of crew. As A. niger is a predominant ISS isolate frequently detected in built environments, studies of A. niger strains inhabiting closed systems may reveal information fundamental to the success of long-duration space missions. This investigation provides valuable insights into the adaptive mechanisms of fungi in extreme environments as well as countermeasures to eradicate unfavorable microbes. Further, it enhances understanding of host-microbe interactions in closed systems, which can help NASA's Human Research Program maintain a habitat healthy for crew during long-term manned space missions

International Space Station conditions alter genomics, proteomics, and metabolomics in Aspergillus nidulans.

The first global genomic, proteomic, and secondary metabolomic characterization of the filamentous fungus Aspergillus nidulans following growth onboard the International Space Station (ISS) is reported. The investigation included the A. nidulans wild-type and three mutant strains, two of which were genetically engineered to enhance secondary metabolite production. Whole genome sequencing revealed that ISS conditions altered the A. nidulans genome in specific regions. In strain CW12001, which features overexpression of the secondary metabolite global regulator laeA, ISS conditions induced the loss of the laeA stop codon. Differential expression of proteins involved in stress response, carbohydrate metabolic processes, and secondary metabolite biosynthesis was also observed. ISS conditions significantly decreased prenyl xanthone production in the wild-type strain and increased asperthecin production in LO1362 and CW12001, which are deficient in a major DNA repair mechanism. These data provide valuable insights into the adaptation mechanism of A. nidulans to spacecraft environments

The International Space Station Environment Triggers Molecular Responses in Aspergillus niger.

Due to immense phenotypic plasticity and adaptability, Aspergillus niger is a cosmopolitan fungus that thrives in versatile environments, including the International Space Station (ISS). This is the first report of genomic, proteomic, and metabolomic alterations observed in A. niger strain JSC-093350089 grown in a controlled experiment aboard the ISS. Whole-genome sequencing (WGS) revealed that ISS conditions, including microgravity and enhanced irradiation, triggered non-synonymous point mutations in specific regions, chromosomes VIII and XII of the JSC-093350089 genome when compared to the ground-grown control. Proteome analysis showed altered abundance of proteins involved in carbohydrate metabolism, stress response, and cellular amino acid and protein catabolic processes following growth aboard the ISS. Metabolome analysis further confirmed that space conditions altered molecular suite of ISS-grown A. niger JSC-093350089. After regrowing both strains on Earth, production of antioxidant-Pyranonigrin A was significantly induced in the ISS-flown, but not the ground control strain. In summary, the microgravity and enhanced irradiation triggered unique molecular responses in the A. niger JSC-093350089 suggesting adaptive responses

\u3ci\u3eDrosophila\u3c/i\u3e Muller F Elements Maintain a Distinct Set of Genomic Properties Over 40 Million Years of Evolution

The Muller F element (4.2 Mb, ~80 protein-coding genes) is an unusual autosome of Drosophila melanogaster; it is mostly heterochromatic with a low recombination rate. To investigate how these properties impact the evolution of repeats and genes, we manually improved the sequence and annotated the genes on the D. erecta, D. mojavensis, and D. grimshawi F elements and euchromatic domains from the Muller D element. We find that F elements have greater transposon density (25–50%) than euchromatic reference regions (3–11%). Among the F elements, D. grimshawi has the lowest transposon density (particularly DINE-1: 2% vs. 11–27%). F element genes have larger coding spans, more coding exons, larger introns, and lower codon bias. Comparison of the Effective Number of Codons with the Codon Adaptation Index shows that, in contrast to the other species, codon bias in D. grimshawi F element genes can be attributed primarily to selection instead of mutational biases, suggesting that density and types of transposons affect the degree of local heterochromatin formation. F element genes have lower estimated DNA melting temperatures than D element genes, potentially facilitating transcription through heterochromatin. Most F element genes (~90%) have remained on that element, but the F element has smaller syntenic blocks than genome averages (3.4–3.6 vs. 8.4–8.8 genes per block), indicating greater rates of inversion despite lower rates of recombination. Overall, the F element has maintained characteristics that are distinct from other autosomes in the Drosophila lineage, illuminating the constraints imposed by a heterochromatic milieu