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  • 1
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    Massachusetts Institute of Technology and Woods Hole Oceanographic Institution
    Publication Date: 2022-05-25
    Description: Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at the Massachusetts Institute of Technology and the Woods Hole Oceanographic Institution June 1999
    Description: In the marine bacterium Vibrio fischeri two intercellular homoserine-Iactone signal molecules (luxI-dependent 30C6-HSL and the ainS-dependent C8-HSL) and the transcriptional activator LuxR regulate the luminescence system in a cell-density dependent manner by a process termed quorum sensing. In this study, five additional proteins whose production is regulated by quorum sensing are described, and the genes encoding four of the five proteins, denoted as QsrP, RibB, QsrV, and AcfA, are analyzed. Each protein is positively regulated by 30C6-HSL and LuxR and negatively regulated at low population density by C8-HSL. Probable LuxR/autoinducer binding sites are found in the promoter region of each. QsrP and RibB are encoded monocistronically, whereas AcfA and QsrV appear to be encoded by a two-gene operon. On the basis of sequence similarity to proteins of known function from other organisms, RibB is believed to be an enzyme that catalyzes the transformation of ribulose 5-phosphate to 3,4-dihydroxy-2- butanone 4-phosphate, a precursor for the xylene ring of riboflavin; AcfA is believed to be a pilus subunit; and the functions of QsrP and QsrV are unknown at this time. A qsrP mutant was reduced in its ability to colonize its symbiotic partner, Euprymna scolopes when placed in competition with the parent strain. On the other hand, a mutant strain of V. fischeri containing an insertion in acfA, which is believed to be polar with respect to qsrV, displayed enhanced colonization competence in a competition assay. A ribB mutant grew well on media not supplemented with additional riboflavin and displayed normal induction of luminescence. Both phenotypes suggest that the lack of a functional ribB gene is complemented by another gene of similar function in the mutant. Oriented divergently from acfA are open reading frames that code for two putative proteins that are similar in sequence to members of the LysR family of transcriptional regulators. Organization of the two divergent sets of genes and the shared promoter region suggests that transcription of acfA and qsrV may be regulated by one or both of these divergently transcribed proteins. This work defines a quorum-sensing regulon in V. fischeri. A model describing its regulation is presented.
    Description: Woods Hole Oceanographic Institution, including The J. Seward Johnson Fund, for contributing financially
    Keywords: Vibrio fischeri ; Bioluminescence ; Cellular signal transduction ; Genetic transcription ; Luminous bacteria
    Repository Name: Woods Hole Open Access Server
    Type: Thesis
    Format: application/pdf
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  • 2
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Science, Ltd
    Molecular microbiology 40 (2001), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The gene hetN encodes a putative oxidoreductase that is known to suppress heterocyst differentiation when present on a multicopy plasmid in Anabaena sp. PCC 7120. To mimic the hetN null phenotype and to examine where HetN acts in the regulatory cascade that controls heterocyst differentiation, we replaced the native chromosomal hetN promoter with the copper-inducible petE promoter. In the presence of copper, heterocyst formation was suppressed in undifferentiated filaments. When hetN expression was turned off by transferring cells to media lacking copper, the filaments initially displayed the wild-type pattern of single heterocysts but, 48 h after the induction of heterocyst formation, a pattern of multiple contiguous heterocysts predominated. Suppression of heterocyst formation by HetN appears to occur both upstream and downstream of the positive regulator HetR: overexpression of hetN in undifferentiated filaments prevents the wild-type pattern of hetR expression as well as the multiheterocyst phenotype normally observed when hetR is expressed from an inducible promoter. Green fluorescent protein fusions show that the expression of hetN in wild-type filaments normally occurs primarily in heterocysts. We propose that HetN is normally involved in the maintenance of heterocyst spacing after the initial heterocyst pattern has been established, but ectopic expression of hetN can also block the initial establishment of the pattern.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: In the filamentous cyanobacterium Anabaena sp. PCC 7120 patS and hetN suppress the differentiation of vegetative cells into nitrogen-fixing heterocysts to establish and maintain a pattern of single heterocysts separated by approximately 10 undifferentiated vegetative cells. Here we show that the patS- and hetN-dependent suppression pathways are the only major factors that prevent vegetative cells from differentiating into heterocysts when a source of ammonia is not present. The patS and hetN pathways are independent of each other, and inactivation of both patS and hetN leads to differentiation of almost all cells of a filament in the absence of a source of fixed nitrogen, compared with approximately 9% in the wild type. Complete differentiation of filaments also occurs when nitrate is supplied as a source of fixed nitrogen, conditions that do not induce differentiation of wild-type filaments. However, ammonia is still capable of suppressing differentiation. The percentage of cells that differentiate into heterocysts appears to be a function of time when a source of fixed nitrogen is absent or a function of growth phase when nitrate is supplied. Although differentiation proceeds unchecked in the absence of patS and hetN expression, differentiation is asynchronous and non-random.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-072X
    Keywords: Key wordsVibrio fischeri ; Spontaneous variant ; Pleiotropic variant ; Dimorphism ; Symbiosis ; Sepiolid squid ; Euprymna scolopes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Vibrio fischeri strains isolated from light organs of the sepiolid squid Euprymna scolpes are non-visibly luminous and fast growing in laboratory culture, whereas in the symbiosis they are visibly luminous and slow growing. A spontaneous, visibly luminous, slow-growing variant was isolated from a laboratory culture of the squid-symbiotic V. fischeri strain ES114. Taxonomic and DNA-homology analyses demonstrated that the variant was V. fischeri and was very similar to the original form. However, the variant grew at one-fourth the rate of the original form, produced 30,000-fold more luminescence, induced luminescence at a lower cell density, and produced a higher level of V. fischeri luminescence autoinducer. Regulation of luminescence, nonetheless, was similar in the two forms and typical of V. fischeri with respect to responses to autoinducer, glucose, the iron chelator ethylenediamine-di(o-hydroxyphenyl acetic acid), and 3′:5′-cyclic AMP. Compared to the original form, cells of the variant were smaller, exhibited from zero to two polar, sheathed flagella instead of a tuft of three to eight flagella, produced a deeper yellow-orange pigment, did not acidify media containing glycerol, and produced a more distinct pellicle. The two forms also differed in the levels of several outer membrane and soluble proteins. These results establish a distinctive physiological, morphological, and biochemical dimorphism in V. fischeri ES114 in which the variant exhibits several traits similar to V. fischeri cells in the symbiotic state. The variant and its conversion from the original form in laboratory culture may provide insight into the properties of V. fischeri cells in the symbiosis and may serve as a model for elucidating the mechanism for their pleiotropic conversion upon colonization of the squid.
    Type of Medium: Electronic Resource
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