skip to main content


Title: Transcriptional profiling of the mutualistic bacterium Vibrio fischeri and an hfq mutant under modeled microgravity
Abstract

For long-duration space missions, it is critical to maintain health-associated homeostasis between astronauts and their microbiome. To achieve this goal it is important to more fully understand the host–symbiont relationship under the physiological stress conditions of spaceflight. To address this issue we examined the impact of a spaceflight analog, low-shear-modeled microgravity (LSMMG), on the transcriptome of the mutualistic bacteriumVibrio fischeri. Cultures ofV. fischeriand a mutant defective in the global regulator Hfq (∆hfq) were exposed to either LSMMG or gravity conditions for 12 h (exponential growth) and 24 h (stationary phase growth). Comparative transcriptomic analysis revealed few to no significant differentially expressed genes between gravity and the LSMMG conditions in the wild type or mutantV. fischeriat exponential or stationary phase. There was, however, a pronounced change in transcriptomic profiles during the transition between exponential and stationary phase growth in bothV. fischericultures including an overall decrease in gene expression associated with translational activity and an increase in stress response. There were also several upregulated stress genes specific to the LSMMG condition during the transition to stationary phase growth. The ∆hfqmutants exhibited a distinctive transcriptome profile with a significant increase in transcripts associated with flagellar synthesis and transcriptional regulators under LSMMG conditions compared to gravity controls. These results indicate the loss of Hfq significantly influences gene expression under LSMMG conditions in a bacterial symbiont. Together, these results improve our understanding of the mechanisms by which microgravity alters the physiology of beneficial host-associated microbes.

 
more » « less
NSF-PAR ID:
10154120
Author(s) / Creator(s):
; ; ; ; ;
Publisher / Repository:
Nature Publishing Group
Date Published:
Journal Name:
npj Microgravity
Volume:
4
Issue:
1
ISSN:
2373-8065
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. The term “microgravity” is used to describe the “weightlessness” or “zero-g” circumstances that can only be found in space beyond earth’s atmosphere. Rhodobacter sphaeroides is a gram-negative purple phototroph, used as a model organism for this study due to its genomic complexity and metabolic versatility. Its genome has been completely sequenced, and profiles of the differential gene expression under aerobic, semi-aerobic, and photosynthetic conditions were examined. In this study, we hypothesized that R. sphaeroides will show altered growth characteristics, morphological properties, and gene expression patterns when grown under simulated microgravity. To test that, we measured the optical density and colony-forming units of cell cultures grown under both microgravity and normal gravity conditions. Differences in the cell morphology were observed using scanning electron microscopy (SEM) images by measuring the length and the surface area of the cells under both conditions. Furthermore, we also identified homologous genes of R. spheroides using the differential gene expression study of Acidovorax under microgravity in our laboratory. Growth kinetics results showed that R. sphaeroides cells grown under microgravity experience a shorter log phase and early stationary phase compared to the cells growing under normal gravity conditions. The length and surface area of the cells under microgravity were significantly higher confirming that bacterial cells experience altered morphological features when grown under microgravity conditions. Differentially expressed homologous gene analysis indicated that genes coding for several COG and GO functions, such as metabolism, signal-transduction, transcription, translation, chemotaxis, and cell motility are differentially expressed to adapt and survive microgravity. 
    more » « less
  2. Plants have been recognized as key components of bioregenerative life support systems for space exploration, and many experiments have been carried out to evaluate their adaptability to spaceflight. Unfortunately, few of these experiments have involved monocot plants, which constitute most of the crops used on Earth as sources of food, feed, and fiber. To better understand the ability of monocot plants to adapt to spaceflight, we germinated and grew Brachypodium distachyon seedlings of the Bd21, Bd21-3, and Gaz8 accessions in a customized growth unit on the International Space Station, along with 1-g ground controls. At the end of a 4-day growth period, seedling organ’s growth and morphologies were quantified, and root and shoot transcriptomic profiles were investigated using RNA-seq. The roots of all three accessions grew more slowly and displayed longer root hairs under microgravity conditions relative to ground control. On the other hand, the shoots of Bd21-3 and Gaz-8 grew at similar rates between conditions, whereas those of Bd21 grew more slowly under microgravity. The three Brachypodium accessions displayed dramatically different transcriptomic responses to microgravity relative to ground controls, with the largest numbers of differentially expressed genes (DEGs) found in Gaz8 (4527), followed by Bd21 (1353) and Bd21-3 (570). Only 47 and six DEGs were shared between accessions for shoots and roots, respectively, including DEGs encoding wall-associated proteins and photosynthesis-related DEGs. Furthermore, DEGs associated with the “Oxidative Stress Response” GO group were up-regulated in the shoots and down-regulated in the roots of Bd21 and Gaz8, indicating that Brachypodium roots and shoots deploy distinct biological strategies to adapt to the microgravity environment. A comparative analysis of the Brachypodium oxidative-stress response DEGs with the Arabidopsis ROS wheel suggests a connection between retrograde signaling, light response, and decreased expression of photosynthesis-related genes in microgravity-exposed shoots. In Gaz8, DEGs were also found to preferentially associate with the “Plant Hormonal Signaling” and “MAP Kinase Signaling” KEGG pathways. Overall, these data indicate that Brachypodium distachyon seedlings exposed to the microgravity environment of ISS display accession- and organ-specific responses that involve oxidative stress response, wall remodeling, photosynthesis inhibition, expression regulation, ribosome biogenesis, and post-translational modifications. The general characteristics of these responses are similar to those displayed by microgravity-exposed Arabidopsis thaliana seedlings. However, organ- and accession-specific components of the response dramatically differ both within and between species. These results suggest a need to directly evaluate candidate-crop responses to microgravity to better understand their specific adaptability to this novel environment and develop cultivation strategies allowing them to strive during spaceflight. 
    more » « less
  3. Summary

    Nitric oxide (NO) is an important defense molecule secreted by the squidEuprymna scolopesand sensed by the bacterial symbiont,Vibrio fischeri, via the NO sensor HnoX. HnoX inhibits colonization through an unknown mechanism. The genomic location ofhnoXadjacent tohahK, a recently identified positive regulator of biofilm formation, suggested that HnoX may inhibit colonization by controlling biofilm formation, a key early step in colonization. Indeed, the deletion ofhnoXresulted in early biofilm formationin vitro, an effect that was dependent on HahK and its putative phosphotransfer residues. An allele ofhnoXthat encodes a protein with increased activity severely delayed wrinkled colony formation. Control occurred at the level of transcription of thesypgenes, which produce the polysaccharide matrix component. The addition of NO abrogated biofilm formation and diminishedsyptranscription, effects that required HnoX. Finally, anhnoXmutant formed larger symbiotic biofilms. This work has thus uncovered a host‐relevant signal controlling biofilm and a mechanism for the inhibition of biofilm formation byV. fischeri. The study ofV. fischeriHnoX permits us to understand not only host‐associated biofilm mechanisms, but also the function of HnoX domain proteins as regulators of important bacterial processes.

     
    more » « less
  4. Abstract

    The binary association between the squid,Euprymna scolopes, and its symbiont,Vibrio fischeri, serves as a model system to study interactions between beneficial bacteria and the innate immune system. Previous research demonstrated that binding of the squid's immune cells, hemocytes, toV. fischeriis altered if the symbiont is removed from the light organ, suggesting that host colonization alters hemocyte recognition ofV. fischeri. To investigate the influence of symbiosis on immune maturation during development, we characterized hemocyte binding and phagocytosis ofV. fischeriand nonsymbioticVibrio harveyifrom symbiotic (sym) and aposymbiotic (apo) juveniles, and wild‐caught and laboratory‐raised sym and apo adults. Our results demonstrate that while light organ colonization byV. fischeridid not alter juvenile hemocyte response, these cells bound a similar number ofV. fischeriandV. harveyiyet phagocytosed onlyV. harveyi. Our results also indicate that long‐term colonization altered the adult hemocyte response toV. fischeribut notV. harveyi. All hemocytes from adult squid, regardless of apo or sym state, both bound and phagocytosed a similar number ofV. harveyiwhile hemocytes from both wild‐caught and sym‐raised adults bound significantly fewerV. fischeri, although moreV. fischeriwere phagocytosed by hemocytes from wild‐caught animals. In contrast, hemocytes from apo‐raised squid bound similar numbers of bothV. fischeriandV. harveyi, although moreV. harveyicells were engulfed, suggesting that blood cells from apo‐raised adults behaved similarly to juvenile hosts. Taken together, these data suggest that persistent colonization by the light organ symbiont is required for hemocytes to differentially bind and phagocytoseV. fischeri. The cellular immune system ofE. scolopeslikely possesses multiple mechanisms at different developmental stages to promote a specific and life‐long interaction with the symbiont.

     
    more » « less
  5. Abstract

    Efficient generation of cardiomyocytes from human-induced pluripotent stem cells (hiPSCs) is important for their application in basic and translational studies. Space microgravity can significantly change cell activities and function. Previously, we reported upregulation of genes associated with cardiac proliferation in cardiac progenitors derived from hiPSCs that were exposed to space microgravity for 3 days. Here we investigated the effect of long-term exposure of hiPSC-cardiac progenitors to space microgravity on global gene expression. Cryopreserved 3D hiPSC-cardiac progenitors were sent to the International Space Station (ISS) and cultured for 3 weeks under ISS microgravity and ISS 1 G conditions. RNA-sequencing analyses revealed upregulation of genes associated with cardiac differentiation, proliferation, and cardiac structure/function and downregulation of genes associated with extracellular matrix regulation in the ISS microgravity cultures compared with the ISS 1 G cultures. Gene ontology analysis and Kyoto Encyclopedia of Genes and Genomes mapping identified the upregulation of biological processes, molecular function, cellular components, and pathways associated with cell cycle, cardiac differentiation, and cardiac function. Taking together, these results suggest that space microgravity has a beneficial effect on the differentiation and growth of cardiac progenitors.

     
    more » « less