Shaping the microenvironment: evidence for the influence of a host galaxin on symbiont acquisition and maintenance in the squid-vibrio symbiosis
Summary Most bacterial species make transitions between habitats, such as switching from free living to symbiotic niches. We provide evidence that a galaxin protein, EsGal1, of the squid Euprymna scolopes participates in both: (i) selection of the specific partner Vibrio fischeri from the bacteriopl...
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| Published in | Environmental microbiology Vol. 16; no. 12; pp. 3669 - 3682 |
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| Main Authors | , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Oxford
Blackwell Publishing Ltd
01.12.2014
Blackwell Wiley Subscription Services, Inc |
| Subjects | |
| Online Access | Get full text |
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| Summary: | Summary
Most bacterial species make transitions between habitats, such as switching from free living to symbiotic niches. We provide evidence that a galaxin protein, EsGal1, of the squid Euprymna scolopes participates in both: (i) selection of the specific partner Vibrio fischeri from the bacterioplankton during symbiosis onset and, (ii) modulation of V. fischeri growth in symbiotic maintenance. We identified two galaxins in transcriptomic databases and showed by quantitative reverse‐transcriptase polymerase chain reaction that one (esgal1) was dominant in the light organ. Further, esgal1 expression was upregulated by symbiosis, a response that was partially achieved with exposure to symbiont cell‐envelope molecules. Confocal immunocytochemistry of juvenile animals localized EsGal1 to the apical surfaces of light‐organ epithelia and surrounding mucus, the environment in which V. fischeri cells aggregate before migration into the organ. Growth assays revealed that one repeat of EsGal1 arrested growth of Gram‐positive bacterial cells, which represent the cell type first ‘winnowed’ during initial selection of the symbiont. The EsGal1‐derived peptide also significantly decreased the growth rate of V. fischeri in culture. Further, when animals were exposed to an anti‐EsGal1 antibody, symbiont population growth was significantly increased. These data provide a window into how hosts select symbionts from a rich environment and govern their growth in symbiosis. |
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| Bibliography: | ark:/67375/WNG-GSK9957V-5 National Science Foundation - No. IOS 0817232 ArticleID:EMI12496 Fig. S1. Expression of esgal1 and esgal2 in symbiotic organs of adult squid. Expression, as measured by qRT-PCR, of esgal1 (A) and esgal2 (B) in the bacteria-containing epithelia of the adult light organ, or central core (CC) and the female-specific accessory nidamental gland (ANG). Data are normalized to the condition of lowest expression. Values ± SEM, and n = 3 biological replicates and 2 technical replicates per condition.Fig. S2. Sequences of the Galaxin proteins used in the alignment in Fig. 1. Full-length protein sequences of the proteins (with Genbank identifying numbers in parentheses) are given, with the repeats used for the alignment highlighted in yellow.Fig. S3. Controls for antibody staining. (A-C) Juvenile squid light organs stained with IgG as a negative control for antibody in the animals shown in Fig. 4. Shown are portions of the light organ including (A) the anterior appendage, (B) the colonized crypt spaces, and (C) the mucus outside of the light organ. No IgG staining (green) is shown, suggesting that the anti-EsGal1 staining in Figs. 4 and S4 is specific. (D), Western blot performed with the anti-EsGal1 antibody on aqueous soluble (S) and SDS-soluble, or membrane, (M) fractions. The arrow denotes a band at the predicted molecular weight. Molecular weight standards in kDa are shown at left.Fig. S4. Localization of EsGal1 in whole juvenile squid. Top, diagram of a juvenile E. scolopes, showing the tissues examined in the remainder of the figure. Multiple tissues of 24 h E. scolopes that were examined for production of the EsGal1 protein by confocal immunocytochemistry, including the tentacles, the gills, and the main haematopoietic organ known as the white body. Day-old juvenile E. scolopes were exposed to the anti-EsGal1 antibody (green), and counterstained with phalloidin (red, actin cytoskeleton) and TOTO-3 (blue, nuclei). The anti-EsGal1 antibody stained the nuclei and apical portions of epithelial cells most brightly (white arrowheads). Aa, anterior appendage; d, ducts; ge, gill epithelium; pa, posterior appendage; te, tentacle epithelium; tm, tentacle musculature; wbe, white body epithelium.Fig. S5. Dose-response of V. fischeri growth to exposure to EsGal1R3. Growth curve of V. fischeri cells exposed to 17.4 μM EsGal1R3 (orange line), 8.2 μM EsGal1R3 (red line), 4.1 μM EsGal1R3 (pink line), or to no peptide (black line). The experiment was performed with 3 biological replicates (except for the 17.4 μM condition, which had 2) and 2 technical replicates.Table S1. Primers used in this study.Table S2. Galaxin sequences and their relevant features, including predicted repeat structure and antimicrobial activity.Appendix S1. Supplementary experimental procedures. NRSA - No. T-32 GM07215 National Institutes of Health - No. R01-RR12294; No. R01-AI50661 istex:D1A5029BE0EDD7501BA735F7AE23742DE0CE386E ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 These authors contributed equally to this work Present Address: Zoological Institute, Christian-Albrechts University of Kiel, 24098 Kiel, Germany |
| ISSN: | 1462-2912 1462-2920 |
| DOI: | 10.1111/1462-2920.12496 |