Subtle selectivity in a pheromone sensor triumvirate desynchronizes competence and predation in a human gut commensal

Constantly surrounded by kin or alien organisms in nature, eukaryotes and prokaryotes developed various communication systems to coordinate adaptive multi-entity behavior. In complex and overcrowded environments, they require to discriminate relevant signals in a myriad of pheromones to execute appropriate responses. In the human gut commensal Streptococcus salivarius, the cytoplasmic Rgg/RNPP regulator ComR couples competence to bacteriocin-mediated predation. Here, we describe a paralogous sensor duo, ScuR and SarF, which circumvents ComR in order to disconnect these two physiological processes. We highlighted the recurring role of Rgg/RNPP in the production of antimicrobials and designed a robust genetic screen to unveil potent/optimized peptide pheromones. Further mutational and biochemical analyses dissected the modifiable selectivity toward their pheromone and operating sequences at the subtle molecular level. Additionally, our results highlight how we might mobilize antimicrobial molecules while silencing competence in endogenous populations of human microflora and temper gut disorders provoked by bacterial pathogens

Study on late competence proteins involved in natural transformation of Bacillus subtilis

The following study comprises in vivo and in vitro data on several of the so-called late competence proteins, which are involved in natural transformation of Bacillus subtilis. The gram-positive bacterium B. subtilis belongs to those bacteria, who are able to take up DNA from their environment and incorporate the foreign DNA by homologous recombination into their own chromosome; a feature named competence. This fascinating ability is carried out by only a portion of the bacterial culture, expressing specific proteins, encoded by the late competence operons. If exogenous double-stranded DNA is about to be taken up from the environment, it needs to first cross the thick cell wall of B. subtilis, with a width of ~40 nm. In case of B. subtilis, this first border is crossed by a putative pseudopilus who transfers the DNA inside of the cell. The energy for this particular process is probably provided by the assembly/disassembly ATPase ComGA. The taken-up DNA is then further transferred into the cytosol by the so-called competence complex or competence machinery. The complex consists out of specific competence proteins, which assemble at the membrane, including a DNA-binding transmembrane protein (ComEA) and an aqueous channel protein (ComEC). In the following thesis, the unknown role of the protein ComEB has been further elucidated in the context of competence, and its enzymatic function was analysed in vitro. It was found that the protein carries out deaminase activity, which is not essential for transformation. In case of ComEC, an amino acid, D573, has been identified as essential for transformation. Truncations of the protein, supposed to carry out an exonuclease function, were heterologously expressed and purified as GST-tag fusions, but, probably due to aggregations of the proteins, no enzymatic activity was detected. The intracellular diffusion of fluorophore fusions of several competence proteins, namely ComEB-mV, ComGA-mV, ComEC-mV and mV-ComEA was analysed via single-molecule tracking, in the presence and absence of exogeneous DNA. In case of ComGA, a C-terminal fusion to mVenus was analysed and it was found that the protein becomes more dynamic in the presence of DNA. Further, the localization and diffusion of a fluorescently labeled PCR product inside of competent Bacillus cells was analysed for the first time. The diffusive behaviour and localization of the stained DNA resembles the diffusion of mV-ComEA. This led to the hypothesis that ComEA serves as a reservoir for taken-up DNA, similar to what is already known for orthologues of ComEA from other naturally competent bacteria

Application of advanced technology to space automation

Automated operations in space provide the key to optimized mission design and data acquisition at minimum cost for the future. The results of this study strongly accentuate this statement and should provide further incentive for immediate development of specific automtion technology as defined herein. Essential automation technology requirements were identified for future programs. The study was undertaken to address the future role of automation in the space program, the potential benefits to be derived, and the technology efforts that should be directed toward obtaining these benefits

Vnitrobuněčná a mezibuněčná regulace genové exprese u Gram-pozitivních bakterií

Bakterie patří mezi nejčetnější organismy na Zemi a jsou součástí našeho každodenního života. Symbiotické bakterie, které se nachází například v trávicím traktu živočichů, mají většinou příznivý vliv na organismus. Existují však také patogenní druhy bakterií, které jsou původci méně či více závažných onemocnění po celém světě. Abychom mohli s patogeny účinně bojovat, je zapotřebí co nejvíce pochopit strategie chování bakteriální populace a molekulární mechanismy, kterými tyto organismy reagují na okolní prostředí. Práce je rozdělena na dvě hlavní části. První část charakterizuje bakteriální genovou expresi v modelových organismech Bacillus subtilis a Mycobacterium smegmatis. DNA-dependentní RNA polymeráza (RNAP) je enzym zodpovědný za přepis nukleových kyselin (z DNA do RNA) a má tedy klíčovou roli v expresi genů. V Disertační práci je řešena struktura RNAP a důležité faktory (proteiny, nebo RNA), které s tímto enzymem asociují a ovlivňují jeho funkci. Druhá část je zaměřena na bakteriální mezibuněčnou komunikaci a faktory/mechanismy (zahrnující genovou expresi), které tento proces ovlivňují. Jeden z typů bakteriální komunikace je označen jako bakteriální nanotrubičky (NT). NT byly v minulosti popsány jako mezibuněčné kanály, kterými si bakterie mohou předávat metabolity, mRNA, proteiny a dokonce...Bacteria, the most dominant organisms on Earth, are an everyday presence in our lives. Symbiotic bacteria, which are present in the digestive tract of animals, usually have a beneficial effect on the body. On the opposite side of the spectrum are pathogenic species that cause more or less serious diseases around the world. In order to fight pathogens effectively, it is necessary to learn as much as possible about the molecular mechanisms by which bacteria respond to their environment, and also about the types of communication within bacterial populations that allow them to react to environmental changes as "multicellular" organisms. This Thesis consists of two main parts. In the first part, selected aspects of bacterial gene expression are characterized, using Bacillus subtilis and Mycobacterium smegmatis as model organisms. DNA-dependent RNA polymerase (RNAP) is the enzyme that is responsible for transcription of DNA into RNA, and thus plays a key role in gene expression. This Thesis deals with the structure of bacterial RNAP and important auxiliary factors (proteins and RNA) that associate with this enzyme and modulate its function. In the second part, the focus is on cell-to-cell communication, revealing which factors/mechanisms, including gene expression, affect this process in B. subtilis....Department of Genetics and MicrobiologyKatedra genetiky a mikrobiologiePřírodovědecká fakultaFaculty of Scienc

ADAPTATION OF MICROBIAL COMMUNITIES AND THEIR FUNCTIONS TO ENVIRONMENTAL PRESSURES IN AN URBAN WATERWAYS SYSTEM

Ph.DDOCTOR OF PHILOSOPH

Sixth Annual Workshop on Space Operations Applications and Research (SOAR 1992), volume 2

This document contains papers presented at the Space Operations, Applications, and Research Symposium (SOAR) hosted by the U.S. Air Force (USAF) on 4-6 Aug. 1992. The symposium was cosponsored by the Air Force Material Command and by NASA/JSC. Key technical areas covered during the symposium were robotics and telepresence, automation and intelligent systems, human factors, life sciences, and space maintenance and servicing. The SOAR differed from most other conferences in that it was concerned with Government-sponsored research and development relevant to aerospace operations. Symposium proceedings include papers covering various disciplines presented by experts from NASA, the USAF, universities, and industry

The Advocate - Jan. 10, 1963

Original title (1951-1987)--The Advocate: official publication of the Archdiocese of Newark (N.J.)

College of Engineering

Cornell University Courses of Study Vol. 96 2004/200