Identifizierung und Charakterisierung zweier für die Entwicklung essentieller Serin/Threonin-Proteinkinasen in Myxococcus xanthus

The ability of Myxococcus xanthus to form fruiting bodies and myxospores mainly depends on the exchange of intercellular signals. The C-signal plays an important role in this process. The recognition of this 17 kDa protein (CsgA) depends on direct cell-cell-contact. Therefore, a receptor in the outer membrane is assumed, which transduces the signal across the cell envelope into the cytoplasm, leading to the expression of various developmentally regulated genes. The aim of this work was the identification of proteins, that are involved in the recognition and transduction of the C-signal, in particular the identification of the so far unknown C-signal receptor. We wanted to purify CsgA as a native protein to use it for the identification of its interaction partners. For this purpose, CsgA was produced with N- and C-terminal affinity tags in M. xanthus. This was successfully possible by using an N-terminal His-tag. The csgA- mutant was functionally complemented by this construct. There is evidence that Ser/Thr protein kinases might be involved in the transduction of the C-signal. Based on a bioinformatic analysis, several Ser/Thr protein kinases that are encoded in putative transcription units with proteins of the outer or inner membranes have been chosen for a mutagenesis screening. Two kinases that are essential for fruiting body formation (MXAN4557 und MXAN6570) were identified by in-frame deletion of those candidate genes. Inactivation of any of the two genes led to a complete loss of the ability to aggregate and sporulate. MXAN4557 is co-transcribed with a TonB-dependent receptor. An upstream transcription unit encodes for a Ser/Thr specific protein phosphatase. However, those two proteins are not essential for development. MXAN6570 forms a transcription unit with a protein that contains an FHA domain, and which is not essential for development either. The analysis of characteristic developmental markers in the deletion mutants of the kinases showed, that early developmental processes are not affected. The results indicate that both proteins act downstream of the C-signal and upstream of the branching point of aggregation and sporulation, where they may represent additional checkpoints for the progression of development.