Control of biofilm formation in Bacillus subtilis
by Jan Gerwig
Date of Examination:2015-01-20
Date of issue:2015-05-12
Advisor:Prof. Dr. Jörg Stülke
Referee:Prof. Dr. Jörg Stülke
Referee:Dr. Fabian Moritz Commichau
Persistent Address:
http://dx.doi.org/10.53846/goediss-5066
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Abstract
English
Biofilms can be considered as one main lifestyle of many bacteria in their natural environment. A bacterial biofilm is a cell community that is surrounded by a self-produced extracellular matrix. This matrix usually consists of polysaccharides, protein, lipids and extracellular DNA. Within this matrix the cells are protected from harmful components in the environment such as antibiotics or from predators and phages. Furthermore, the biofilm matrix enables the cells to float on liquid surfaces as a community or to cover solid surfaces such as plant roots, human tissue or even medical devices. Thus, the formation of biofilms by pathogenic bacteria can also serve as a virulence factor and needs to be considered as a threat to human health. The Gram-positive model organism Bacillus subtilis also forms biofilms in its natural environment. In the laboratory environment it forms wrinkled colonies on agar plates and structured floating biofilms, so-called pellicles, on the top of liquid medium. The regulation of matrix gene expression is highly complex and was subject of many studies. The main protein components of the matrix are encoded in the tapA-sipW-tasA operon and the bslA gene, whereas the machinery for exopolysaccharide synthesis and export is encoded within the epsA-O operon. One aim of this work was to study the function of the first two genes of the epsA-O operon, namely epsA and epsB that encode a tyrosine kinase modulator and the cognate kinase in the regulation of exopolysaccharide production. In this work, it was shown that the EpsB kinase and its modulator directly interact with each other and that deletion of the two genes reduces the biofilm structure suggesting a defect in exopolysaccharide production. Simultaneous deletion of the genes for EpsB and the only other known tyrosine kinase PtkA led to a complete loss of complex colony formation due to impaired exopolysaccharide production. The same was observed in the absence of both modulator proteins demonstrating that tyrosine kinases are essential for the formation of biofilms in B. subtilis. The expression of biofilm matrix and motility genes is mutually exclusive in B. subtilis. In the absence of the YmdB protein biofilm matrix gene expression is inhibited and instead all cells express motility genes. Thus, the ymdB mutant does not form complex colonies and pellicles anymore. This phenotype is due to the phosphodiesterase activity of the YmdB protein. In this work, it was demonstrated that impaired biofilm formation is due increased amounts of the master regulator of biofilm formation SinR. Interestingly, the formation of spontaneous suppressor mutations within the sinR gene restored biofilm matrix gene expression and enabled the cells to switch between sessile and motile life styles. However, the target of the YmdB phosphodiesterase remains unclear but the interplay with the major endoribonuclease RNase Y seems to be important. RNA sequencing of the ymdB mutant revealed potential processing targets for further research.
Keywords: BY-kinases; phosphodiesterases; tyrosine phosphorylation
