CCaMK and CYCLOPS
Inside the nucleus: interpretation of calcium spiking
The nucleus associated calcium spiking response elicited by rhizobia and arbuscular mycorrhiza (AM) fungi is proposed to be decoded and transduced by the plant specific calcium- and calmodulin-dependent kinase CCaMK (Kosuta et al., 2008; Oldroyd and Downie, 2008). Upon symbiotic interaction, ccamk mutants initiate calcium spiking, but expression of early symbiosis genes, and thus establishment of root nodule symbiosis (RNS) and AM is impaired. The domain structure of the protein, a kinase domain linked to a calmodulin binding site and three calcium binding EF hand motifs, implies a dual, or differential mode of activation by free and calmodulin bound calcium ions, potentially enabling the protein to integrate calcium signatures derived from AM fungi and rhizobia, to mount the distinct cellular responses required for their accommodation (Gleason et al., 2006; Tirichine et al., 2006). The finding that a point mutation in the autophosphorylation site of CCaMK (snf1) triggers the development of nodules in the absence of rhizobia, identified CCaMK as a key regulatory switch controlling nodule organogenesis in the root cortex (Gleason et al., 2006; Tirichine et al., 2006). CCaMK forms a preassembled complex with CYCLOPS in the nucleus, as interaction occurs in the absence of a calcium stimulus (Yano et al., 2008). cyclops mutants are defective in AM and RNS, but show a wildtype-like calcium spiking response (Kistner et al., 2005; Miwa et al., 2006; Yano et al., 2008). The mutant name is derived from its particular phenotype characterized by a root hair curl entrapping a bacterial microcolony. At this early stage, progression of infection via an infection thread is blocked (Yano et al., 2008). Sequence analysis of CYCLOPS predicts a protein of unknown function with two nuclear localization signals and a short coiled-coil at the C-terminus. The observation that spontaneous nodules are formed in cyclops mutants transformed with gain-of-function CCaMK, suggests a bifurcation of signal transduction at, or below CCaMK. According to this model, CCaMK mediates microbial infection via CYCLOPS in the epidermis, while nodule organogenesis is initiated via yet unidentified CCaMK targets in the cortex (Yano et al., 2008).In order to understand the molecular mechanisms involved in the decoding and transduction of the respective symbiosis-specific calcium oscillations, we aim at a detailed structural and functional characterization of the nuclear CCaMK/CYCLOPS signaling complex by means of genetical, physiological and biochemical approaches. In this context we are also addressing the intriguing specificity and underlying mechanisms leading to distinct responses (e.g. nodule formation in RNS but not during AM) downstream of calcium spiking. Further, we attempt to identify additional complex components and to determine their role in symbiosis.
Gleason, C., Chaudhuri, S., Yang, T.B., Munoz, A., Poovaiah, B.W., and Oldroyd, G.E.D. (2006). Nodulation independent of rhizobia induced by a calcium-activated kinase lacking autoinhibition. Nature 441: 1149–1152.
Kistner, C., Winzer, T., Pitzschke, A., Mulder, L., Sato, S., Kaneko, T., Tabata, S., Sandal, N., Stougaard, J., Webb, K.J., Szczyglowski, K., and Parniske, M. (2005). Seven Lotus japonicus genes required for transcriptional reprogramming of the root during fungal and bacterial symbiosis. Plant Cell 17: 2217–2229.
Kosuta, S., Hazledine, S., Sun, J., Miwa, H., Morris, R.J., Downie, J.A., and Oldroyd. G.E.D. (2008). Differential and chaotic calcium signatures in the symbiosis signaling pathway of legumes. Proc. Natl. Acad. Sci. USA 105: 9823–9828.
Miwa, H., Sun, J., Oldroyd, G.E., and Downie, J.A. (2006). Analysis of Nod-factor-induced calcium signaling in root hairs of symbiotically defective mutants of Lotus japonicus. Mol. Plant Microbe Interact. 19: 914–923.
Oldroyd, G.E.D., and Downie, J.A. (2008). Coordination of nodule morphogenesis with rhizobial infection in legumes. Annu. Rev. Plant Biol. 59: 519–546.
Tirichine, L., Imaizumi-Anraku, H., Yoshida, S., Murakami, Y., Madsen, L., Miwa, H., Nakagawa, T., Sandal, N., Albrektsen, A., Kawaguchi, M., Downie, A., Sato, S., Tabata, S., Kouchi, H., Parniske, M., Kawasaki, S., and Stougaard, J. (2006). Deregulation of a Ca2+/calmodulin-dependent kinase leads to spontaneous nodule development. Nature 441: 1153-1156.
Yano, K., Yoshida, S., Müller, J., Singh, S., Banba, M., Vickers, K., Markmann, K., White, C., Schuller, B., Sato, S., Asamizu, E., Tabata, S., Murooka, Y., Perry, J., Wang, T.L., Kawaguchi, M., Imaizumi-Anraku, H., Hayashi, M., and Parniske, M. (2008). CYCLOPS, a mediator of symbiotic intracellular accommodation. Proc. Natl. Acad. Sci. USA 105: 20540–20545.