Bacteria achieve directed motion through their environments by integrating propulsion with chemical detection in the process of chemotaxis. Central to this process are the macromolecular protein structures of the flagellar motor and the chemoreceptor arrays, which are responsible for motility and chemical sensing, respectively. These protein complexes localise to different discrete subcellular positions in different bacterial species, and their correct subcellular localisation is often essential to their function. In the monotrichous α‐proteobacterium Rhodobacter sphaeroides, the flagellum is subpolar and two distinct sets of chemotaxis proteins localise to discrete polar and cytoplasmic positions within the cell. In this study, the development of software for the analysis of fluorescent microscopy images allowed cellular morphologies and the localisation and distribution of the chemoreceptor arrays of R.sphaeroides to be characterised in detail, showing that protein partitioning at cell division results in an asymmetric separation of both cytoplasmic and membrane‐bound protein components between daughter cells. The design of a fluorescence‐based assay for the analysis of gene expression assisted in demonstrating that expression of both the chemotaxis and motor genes of R.sphaeroides is regulated by the sigma factor, FliA, and its inhibitor, FlgM. FliA was then used to achieve varying expression of the chemotaxis genes, and the concentration dependence of array clustering was explored in microscopy images, revealing important differences between cluster formation in R.sphaeroides and other species. Additionally, FliA was identified as a regulator of flagellar number in R.sphaeroides, controlling a negative feedback‐loop in the hierarchy of flagellar assembly that represses flagellar formation upon secretion of FlgM. The complex regulatory pathway controlling R.sphaeroides flagellar assembly is the first identified system where completion of a single flagellum directly inhibits the production of a second, a mechanism that may be important to many monotrichous bacterial species.

Im Fokus dieser Arbeit stand die Analyse von Faktoren, die den Zusammenbau des Flagellenapparates von Legionella pneumophila (Lp) regulieren. Mit einem kombinierten Replikations-/ Überlebensversuchs mit Lp Corby oder Lp Paris und ihren zugehörigen Regulationsmutanten wurde eine verminderte Fitness für dfliA und erstmals für drpoN, dfleQ defiziente Stämme nachgewiesen. Zur Validierung von Microarray-Daten aus Lp Paris wurden wachstumsphasenabhängige Transkriptions- und Translationsanalysen mit Lp Corby Wildtyp und drpoN, dfleQ, dfliA und dflaA defizienten Stämmen durchgeführt. Es wurde gezeigt, dass die basale Expression von fliA in den späteren Phasen unabhängig von RpoN und FleQ stattfindet. In dieser Arbeit konnte erstmals der Transkriptionsstartpunkt des Hauptregulators FliA bestimmt werden. Es zeigte sich eine putative RpoD (S70) Bindungsstelle. Ein Modell zur Regulation der fliA Expression wurde weiterentwickelt. Demnach kommt es in der exponentiellen Phase durch das Zusammenwirken von RpoD und DksA, aber unabhängig von FleQ, zur basalen fliA Promotoraktivität. Durch den Übergang in die transmissive Phase und direkte oder indirekte Interaktion mit FleQ sowie dem Alarmon ppGpp scheint es zu einem Austausch des Sigmafaktors S70 gegen SS und zu einer Aktivierung der fliA Expression zu kommen. Elektronenmikroskopische Studien zeigten, dass drpoN und dfleQ defiziente Mutanten wahrscheinlich aufgrund des fehlenden Basalkörpers nicht flagelliert sind. Mutanten für dfliA, dflaA und dfliD hatten ebenfalls keine Flagelle, zeigten aber eine ungewöhnliche, gerade Hook Struktur, die den Zusammenbau des Basalkörpers demonstriert. Weiterhin wurden durch in silico Studien 15 Legionella Spezies in Bezug auf das Flagellensystem und ein putatives Chemotaxissystem untersucht. So konnte L. oakridgensis als erste Art ohne beide Systeme sequenziert werden. Andererseits konnten mit LLAP12, L. bozemanii, L. gormanii und L. lytica Stämme beschrieben werden, die beide Systeme tragen. / This work focused on the analysis of factors contributing to the regulation of the flagellum self-assembly of Legionella pneumophila (Lp). With a combined replication/survival assay with Lp Corby or Lp Paris and their corresponding regulatory mutants a reduced fitness could be verified for dfliA and for the first time for drpoN, dfleQ deficient strains. For validation of microarray-data for Lp Paris with strain Lp Corby a growth phase dependent analysis of transcription and translation rates was done with wild-type and the drpoN, dfleQ, dfliA and dflaA deficient strains. A regulation of basal fliA expression independently from RpoN and FleQ was shown in the later growth phases. Furthermore the transcriptional start site of fliA could be shown for the first time. A RpoD (S70) binding site could be identified. According to a further developed model for the regulation of the fliA expression RpoD and DksA lead to a basal fliA promotor activity, independently from FleQ. Most likely, during transition to stationary phase, direct or indirect interaction with FleQ and the alarmone ppGpp results in the exchange of the sigma factor S70 and the binding of RpoS. This leads to the activation of fliA expression. Electron microscopic studies revealed that drpoN and dfleQ deficient mutants are not flagellated caused by the missing basal body. Mutants of dfliA, dflaA and dfliD were also aflagellated, but there was a uncommon straight hook structure visible which demonstrates a filament-independent assembly of the basal body. Furthermore, in silico analysis was done with 15 Legionella species with regard to the flagellum regulation system and a putative chemotaxis system. Analysis revealed that the strain L. oakridgensis is the first strain lacking both systems. On the other hand the strains LLAP12, L. bozemanii, L. gormanii and L. lytica could be characterized as strains carrying both systems.