H-032. Characterization of the Histidine Kinase Partners of Enhancer Binding Proteins Important for Myxococcus xanthus Fruiting Body Development

Z. Sarwar, A. G. Garza;
Syracuse Univ., Syracuse, NY.

The social behavior and complex life cycle of Myxococcus xanthus make it an ideal model for multicellular processes such as biofilm formation. Changes in gene expression are important for the formation of organized, multicellular groups and these changes are often controlled by two-component signal transduction systems (TCS). TCS are commonly used by prokaryotes for signal transduction and they minimally contain a histidine kinase (HK) sensor protein, and a response regulator (RR), which typically modulates transcription of target genes. Many M. xanthus RRs are members of the NtrC family of transcriptional activators and, therefore, they function as enhancer binding proteins (EBPs). Two of these EBPs, Nla6 and Nla28, are key players in the multicellular process of M. xanthus fruiting body development and one of our goals is to identify their in vivo HK partners. MXAN4043 and MXAN1166 were identified as possible HK partners for Nla6 and Nla28, respectively; both HK genes are located adjacent to their respective RR genes and they encode proteins with the conserved domains found in HKs. In response to environmental changes, HKs autophosphorylate using ATP as the phosphate donor. Purified MBP-MXAN4043 and MBP-MXAN1166 fusion proteins hydrolyzed ATP at rates of 0.2 μMs-1 and 0.01 μMs-1, respectively. Furthermore, when purified MBP-MXAN4043 and MBP-MXAN1166 were incubated with [γ32P]-ATP, phosphorylation of the HKs was detected. In TCS, autophosphorylation of an HK is followed by a phosphotransfer to its cognate RR. When phosphorylated MBP-MXAN1166 was incubated with purified MBP-Nla28 (putative cognate RR), phosphorylated MBP-Nla28 was detected. Phosphotransfer from MBP-MXAN4043 to MBP-Nla6 (putative cognate RR) is now being investigated, and in vivo mutational studies of the RR genes and their corresponding HK partner genes are being performed to confirm that we have identified the in vivo HK partners of Nla6 and Nla28.