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Bi-modal distribution of the second messenger c-di-GMP controls cell fate and asymmetry during the caulobacter cell cycle
No full textMany bacteria mediate important life-style decisions by varying levels of the second messenger c-di-GMP. Behavioral transitions result from the coordination of complex cellular processes such as motility, surface adherence or the production of virulence factors and toxins. While the regulatory mechanisms responsible for these processes have been elucidated in some cases, the global pleiotropic effects of c-di-GMP are poorly understood, primarily because c-di-GMP networks are inherently complex in most bacteria. Moreover, the quantitative relationships between cellular c-di-GMP levels and c-di-GMP dependent phenotypes are largely unknown. Here, we dissect the c-di-GMP network of Caulobacter crescentus to establish a global and quantitative view of c-di-GMP dependent processes in this organism. A genetic approach that gradually reduced the number of diguanylate cyclases identified novel c-di-GMP dependent cellular processes and unraveled c-di-GMP as an essential component of C. crescentus cell polarity and its bimodal life cycle. By varying cellular c-di-GMP concentrations, we determined dose response curves for individual c-di-GMP-dependent processes. Relating these values to c-di-GMP levels modeled for single cells progressing through the cell cycle sets a quantitative frame for the successive activation of c-di-GMP dependent processes during the C. crescentus life cycle. By reconstructing a simplified c-di-GMP network in a strain devoid of c-di-GMP we defined the minimal requirements for the oscillation of c-di-GMP levels during the C. crescentus cell cycle. Finally, we show that although all c-di-GMP dependent cellular processes were qualitatively restored by artificially adjusting c-di-GMP levels with a heterologous diguanylate cyclase, much higher levels of the second messenger are required under these conditions as compared to the contribution of homologous c-di-GMP metabolizing enzymes. These experiments suggest that a common c-di-GMP pool cannot fully explain spatiotemporal regulation by c-di-GMP in C. crescentus and that individual enzymes preferentially regulate specific phenotypes during the cell cycle
Locating the ‘radical’ in 'Shoot the Messenger'
No full textThis is the author's accepted manuscript. The final published article is available from the link below, copyright 2013 @ Edinburgh University Press.The 2006 BBC drama Shoot the Messenger is based on the psychological journey of a Black schoolteacher, Joe Pascale, accused of assaulting a Black male pupil. The allegation triggers Joe's mental breakdown which is articulated, through Joe's first-person narration, as a vindictive loathing of Black people. In turn, a range of common stereotypical characterisations and discourses based on a Black culture of hypocrisy, blame and entitlement is presented. The text is therefore laid wide open to a critique of its neo-conservatism and hegemonic narratives of Black Britishness. However, the drama's presentation of Black mental illness suggests that Shoot the Messenger may also be interpreted as a critique of social inequality and the destabilising effects of living with ethnicised social categories. Through an analysis of issues of representation, the article reclaims this controversial text as a radical drama and examines its implications for and within a critical cultural politics of ‘race’ and representation
Folding of the transcription factor Brinker and interactions of the bacterial second messenger c-di-GMP studied by NMR
No full textNuclear magnetic resonance (NMR) spectroscopy is a technique, which allows the non-invasive investigation of
structures, dynamics and interactions of biomolecules.
The main goal of this thesis was to elucidate the folding mechanism of the transcription factor Brinker and its
implications for DNA recognition as well as the characterization of unfolded protein states by NMR.
This constitutes the first part of this thesis.
The transcription factor Brinker is a nuclear repressor, which is involved in
cellular growth and differentiation.
In the absence of DNA, Brinker is completely disordered.
However, in the presence of DNA or at low temperatures, the Brinker DNA binding domain (BrkDBD) adopts a well-folded structure.
Thus, BrkDBD represents an extreme case of the coupling between binding and folding phenomenon.
We have aimed to elucidate this folding mechanism in order to understand its implications for DNA recognition.
From our data, it is clear that the BrkDBD folding energy landscape sharply depends on buffer anion type and concentration.
We show that folded BrkDBD always adopts the same structure irrespective of the conditions.
Our data indicate helical propensity for 3 of the 4 native helices even in unfolded BrkDBD, which may serve as
initial contact points for DNA recognition.
Resonance broadening due to conformational exchange on the micro- to millisecond time scale between folded and
unfolded BrkDBD was analyzed by NMR relaxation dispersion experiments indicating a two-state
folding mechanism.
Only few residues show a different behavior and these are all located at the DNA binding interface.
This local conformational heterogeneity may be important for DNA recognition.
Based on these findings, we propose a mechanism of DNA recognition by BrkDBD, where the electrostatics-driven folding
is a key component, accelerating the recognition process.
In addition, we have analyzed the side-chain chi1-rotamer distribution of urea-denatured ubiquitin and protein G,
revealing that individual residues show significant deviations from statistical-coil ensemble averages,
indicating local bias towards the folded state.
The second part of this thesis describes the quantitative characterization of the intermolecular interactions between monomers
of the bacterial second messenger c-di-GMP at physiologically relevant concentrations.
C-di-GMP is a bacterial second messenger, involved in many signaling events.
Its most important effect is to trigger the transition from motile to sessile bacterial life-styles which plays a major
role in biofilm formation.
In solution, c-di-GMP has been reported to form several oligomers in the presence of monovalent cations, particularly
potassium.
However, only monomeric and dimeric c-di-GMP have been observed in complexes with proteins or RNA.
We have carried out a detailed kinetic and thermodynamic analysis of c-di-GMP polymorphism in the presence of potassium,
which showed that predominantly monomers and only few dimers exist at physiological concentrations.
Additionally, we present NOE and ROE structural information on c-di-GMP oligomers, which
indicate that these are not entirely all-syn and all-anti as opposed to the literature
Formation and dimerization of the phosphodiesterase active site of the Pseudomonas aeruginosa MorA, a bi-functional c-di-GMP regulator
No full textDiguanylate cyclases (DGC) and phosphodiesterases (PDE) respectively synthesise and hydrolyse the secondary messenger cyclic dimeric GMP (c-di-GMP), and both activities are often found in a single protein. Intracellular c-di-GMP levels in turn regulate bacterial motility, virulence and biofilm formation. We report the first structure of a tandem DGC–PDE fragment, in which the catalytic domains are shown to be active. Two phosphodiesterase states are distinguished by active site formation. The structures, in the presence or absence of c-di-GMP, suggest that dimerisation and binding pocket formation are linked, with dimerisation being required for catalytic activity. An understanding of PDE activation is important, as biofilm dispersal via c-di-GMP hydrolysis has therapeutic effects on chronic infections
David C. Smith Heads Bell Community Relations
No full textDavid C. Smith, Ohio Bell residences services manager for Fremont, Sandusky and Fostoria, Ohio, has been named community relations manager for the firm in Northwest Ohio. He will be succeeded by Glenn R. Baker of Lindsey, Ohio. Smith resides in Sandusky, Ohio
The Kiel Press
No full textWeekly newspaper from Kiel, Oklahoma that includes local, territorial, and national news along with advertising
NOS in the cephalopod "cerebellum".
No full textIn short, these lobes are strikingly similar in their function, cyto-architecture and connectivity to the vertebrate cerebellum and, like the cerebellum, which contains the highest level of NOS in the mammalian brain (Rodrigo et al. 1994), they are now shown to contain high levels of NOS
BRUCE: a program for the detection of transfer-messenger RNA genes in nucleotide sequences
No full textA computer program, BRUCE, was developed for the identification of transfer‐messenger RNA (tmRNA) genes. The program employs heuristic algorithms to search for a tRNAAla‐like secondary structure surrounding a short sequence encoding the tag peptide. In the 57 completely sequenced bacterial genomes where tmRNA genes have been reported previously, BRUCE identified all with no false positives. In addition, BRUCE found 99 of the 100 tmRNAs identified previously in other bacteria, red chloroplasts and cyanelles. The output of the program reports the proposed tRNA secondary structure, the tmRNA gene sequence and the tag peptide
Binding of bacterial secondary messenger molecule c di-GMP is a STING operation
No full textInitial skirmishes between the host and pathogen result in spillage of the contents of the bacterial cell. Amongst the spillage, the secondary messenger molecule, cyclic dimeric guanosine monophosphate (c di-GMP), was recently shown to be bound by stimulator of interferon genes (STING). Binding of c di-GMP by STING activates the Tank Binding Kinase (TBK1) mediated signaling cascades that galvanize the body's defenses for elimination of the pathogen. In addition to c di-GMP, STING has also been shown to function in innate immune responses against pathogen associated molecular patterns (PAMPs) originating from the DNA or RNA of pathogens. The pivotal role of STING in host defense is exemplified by the fact that STING(-/-) mice die upon infection by HSV-1. Thus, STING plays an essential role in innate immune responses against pathogens. This opens up an exciting possibility of targeting STING for development of adjuvant therapies to boost the immune defenses against invading microbes. Similarly, STING could be targeted for mitigating the inflammatory responses augmented by the innate immune system. This review summarizes and updates our current understanding of the role of STING in innate immune responses and discusses the future challenges in delineating the mechanism of STING-mediated responses
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