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Bacterial bioluminescence regulates expression of a host cryptochrome gene in the squid-Vibrio symbiosis.

ABSTRACT: The symbiosis between the squid Euprymna scolopes and its luminous symbiont, Vibrio fischeri, is characterized by daily transcriptional rhythms in both partners and daily fluctuations in symbiont luminescence. In this study, we sought to determine whether symbionts affect host transcriptional rhythms. We identified two transcripts in host tissues (E. scolopes cry1 [escry1] and escry2) that encode cryptochromes, proteins that influence circadian rhythms in other systems. Both genes cycled daily in the head of the squid, with a pattern similar to that of other animals, in which expression of certain cry genes is entrained by environmental light. In contrast, escry1 expression cycled in the symbiont-colonized light organ with 8-fold upregulation coincident with the rhythms of bacterial luminescence, which are offset from the day/night light regime. Colonization of the juvenile light organ by symbionts was required for induction of escry1 cycling. Further, analysis with a mutant strain defective in light production showed that symbiont luminescence is essential for cycling of escry1; this defect could be complemented by presentation of exogenous blue light. However, blue-light exposure alone did not induce cycling in nonsymbiotic animals, but addition of molecules of the symbiont cell envelope to light-exposed animals did recover significant cycling activity, showing that light acts in synergy with other symbiont features to induce cycling. While symbiont luminescence may be a character specific to rhythms of the squid-vibrio association, resident microbial partners could similarly influence well-documented daily rhythms in other systems, such as the mammalian gut.

SUBMITTER: Heath-Heckman EA

PROVIDER: S-EPMC3622930 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

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Bacterial bioluminescence regulates expression of a host cryptochrome gene in the squid-Vibrio symbiosis.

Heath-Heckman Elizabeth A C EA Peyer Suzanne M SM Whistler Cheryl A CA Apicella Michael A MA Goldman William E WE McFall-Ngai Margaret J MJ

mBio 20130402 2

The symbiosis between the squid Euprymna scolopes and its luminous symbiont, Vibrio fischeri, is characterized by daily transcriptional rhythms in both partners and daily fluctuations in symbiont luminescence. In this study, we sought to determine whether symbionts affect host transcriptional rhythms. We identified two transcripts in host tissues (E. scolopes cry1 [escry1] and escry2) that encode cryptochromes, proteins that influence circadian rhythms in other systems. Both genes cycled daily i ...[more]

PMID: 23549919

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Project description:In horizontally transmitted symbioses, structural, biochemical, and molecular features both facilitate host colonization by specific symbionts and mediate their persistent carriage. In the association between the squid Euprymna scolopes and its luminous bacterial partner Vibrio fischeri, the symbionts interact with two epithelial fields; they interact (i) transiently with the superficial ciliated field that potentiates colonization and regresses within days of colonization and (ii) persistently with the cells that line the internal crypts, whose ultrastructure changes in response to the symbionts. Development of the association creates conditions that promote the symbiotic partner over the lifetime of the host. To determine whether light organ maturation requires continuous interactions with V. fischeri or only the signaling that occurs during its initiation, we compared 4-week-old squid that were uncolonized with those colonized either persistently by wild-type V. fischeri or transiently by a V. fischeri mutant that triggers early events in morphogenesis but does not persist. Microscopic analysis of the light organs showed that, while morphogenesis of the superficial ciliated field is greatly accelerated by V. fischeri colonization, its eventual outcome is largely independent of colonization state. In contrast, the symbiont-induced changes in crypt cell shape require persistent host-symbiont interaction, reflected in the similarity between uncolonized and transiently colonized animals. Transcriptomic analyses reflected the microscopy results; host gene expression at 4?weeks was due primarily to the persistent interactions of host and symbiont cells. Further, the transcriptomic signature of specific pathways reflected the daily rhythm of symbiont release and regrowth and required the presence of the symbionts. IMPORTANCE A long-term relationship between symbiotic partners is often characterized by development and maturation of host structures that harbor the symbiont cells over the host's lifetime. To understand the mechanisms involved in symbiosis maintenance more fully, we studied the mature bobtail squid, whose light-emitting organ, under experimental conditions, can be transiently or persistently colonized by Vibrio fischeri or remain uncolonized. Superficial anatomical changes in the organ were largely independent of symbiosis. However, both the microanatomy of cells with which symbionts interact and the patterns of gene expression in the mature animal were due principally to the persistent interactions of host and symbiont cells rather than to a response to early colonization events. Further, the characteristic pronounced daily rhythm on the host transcriptome required persistent V. fischeri colonization of the organ. This experimental study provides a window into how persistent symbiotic colonization influences the form and function of host animal tissues.

Project description:UnlabelledThe synergistic activity of Vibrio fischeri lipid A and the peptidoglycan monomer (tracheal cytotoxin [TCT]) induces apoptosis in the superficial cells of the juvenile Euprymna scolopes light organ during the onset of the squid-vibrio symbiosis. Once the association is established in the epithelium-lined crypts of the light organ, the host degrades the symbiont's constitutively produced TCT by the amidase activity of a peptidoglycan recognition protein (E. scolopes peptidoglycan recognition protein 2 [EsPGRP2]). In the present study, we explored the role of alkaline phosphatases in transforming the lipid A of the symbiont into a form that changes its signaling properties to host tissues. We obtained full-length open reading frames for two E. scolopes alkaline phosphatase (EsAP) mRNAs (esap1 and esap2); transcript levels suggested that the dominant light organ isoform is EsAP1. Levels of total EsAP activity increased with symbiosis, but only after the lipid A-dependent morphogenetic induction at 12 h, and were regulated over the day-night cycle. Inhibition of total EsAP activity impaired normal colonization and persistence by the symbiont. EsAP activity localized to the internal regions of the symbiotic juvenile light organ, including the lumina of the crypt spaces where the symbiont resides. These data provide evidence that EsAPs work in concert with EsPGRPs to change the signaling properties of bacterial products and thereby promote persistent colonization by the mutualistic symbiont.ImportanceThe potential for microbe-associated molecular patterns (MAMPs) to compromise host-tissue health is reflected in the often-used nomenclature for these molecules: lipopolysaccharide (LPS) is also called "endotoxin" and the peptidoglycan monomer is also called "tracheal cytotoxin" (TCT). With constant presentation of MAMPs by the normal microbiota, mechanisms to tolerate their effects have developed. The results of this contribution provide evidence that host alkaline phosphatases (APs) dephosphorylate and inactivate the symbiont MAMP lipid A. As such, APs work in synergy with a peptidoglycan recognition protein, which inactivates symbiont-exported TCT, to alter the symbiont MAMPs and promote persistence of the partnership. Not only may these activities serve to "tame" the MAMPs, but also the resulting products may themselves be important signals in persistent mutualisms. The finding of lipid A modification by APs in an invertebrate mutualism provides evidence that this specific strategy for dealing with symbiotic partners is conserved across the animal kingdom.

Project description:The squid Euprymna scolopes and the luminescent bacterium Vibrio fischeri form a highly specific beneficial light organ symbiosis. Not only does the host have to select V. fischeri from the environment, but it must also prevent subsequent colonization by non-symbiotic microorganisms. Host macrophage-like hemocytes are believed to play a role in mediating the symbiosis with V. fischeri. Previous studies have shown that the colonization state of the light organ influences the host's hemocyte response to the symbiont. To further understand the molecular mechanisms behind this process, we used two quantitative mass-spectrometry-based proteomic techniques, isobaric tags for relative and absolute quantification (iTRAQ) and label-free spectral counting, to compare and quantify the adult hemocyte proteomes from colonized (sym) and uncolonized (antibiotic-treated/cured) squid. Overall, iTRAQ allowed for the quantification of 1,024 proteins with two or more peptides. Thirty-seven unique proteins were determined to be significantly different between sym and cured hemocytes (p value < 0.05), with 20 more abundant proteins and 17 less abundant in sym hemocytes. The label-free approach resulted in 1,241 proteins that were identified in all replicates. Of 185 unique proteins present at significantly different amounts in sym hemocytes (as determined by spectral counting), 92 were more abundant and 93 were less abundant. Comparisons between iTRAQ and spectral counting revealed that 30 of the 37 proteins quantified via iTRAQ exhibited trends similar to those identified by the label-free method. Both proteomic techniques mutually identified 16 proteins that were significantly different between the two groups of hemocytes (p value < 0.05). The presence of V. fischeri in the host light organ influenced the abundance of proteins associated with the cytoskeleton, adhesion, lysosomes, proteolysis, and the innate immune response. These data provide evidence that colonization by V. fischeri alters the hemocyte proteome and reveals proteins that may be important for maintaining host-symbiont specificity.

Dataset Information

Bacterial bioluminescence regulates expression of a host cryptochrome gene in the squid-Vibrio symbiosis.

Publications

Bacterial bioluminescence regulates expression of a host cryptochrome gene in the squid-Vibrio symbiosis.

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