Exploring new links between phosphoinositides and phytohormone signaling in Arabidopsis thaliana
Zusammenfassung der Projektergebnisse
Phosphoinositides (PIs) are minor but essential lipid constituents of eukaryotic membranes. The components of a PI signaling system in plants have long been known. However, up to now it has remained unclear what cellular functions PIs or PI-derived second messengers have in plants and whether such functions of PIs would include plant-specific roles not found in other eukyryotic models. Therefore, this project was initiated to determine whether there are links between the PI signaling system and plant-typical phytohormones, such as auxin or jasmonic acid (JA). At the beginning of this study it was established that PI-derived signaling molecules are essential for typical auxin- or JA-dependent plant responses, such as gravitropic curvature or wound-responses, respectively. Also it had just been determined that the auxin receptor protein, TIR1, contained PI- derived inositol hexakisphosphate (InsP6) bound in its crystal structure. Because TIR1 is structurally related to the JA-receptor protein, COI1, it was one goal of this project to determine, whether InsP6 or another inositol polyphosphate was required for the function of TIR1 and/or COI1 in the context of gravitropic curvature and defence responses to wounding. In order to test these two responses for involvement of inositol polyphosphates, two plant lines defective in PI metabolism were compared to wild type controls. InsP 5-ptase plants with globally reduced PI intermediates were used to test for general involvement of PIs in each process. Furthermore, ipk1-1 plants defective in the production of InsP6 were used to test whether TIR1 or COI1 would function in plants with reduced InsP6 levels. Both plant lines were first physiologically characterized in the context of auxin- and JA-dependent responses. Both lines exhibited reduced gravitropic bending. When testing for wounding and defense responses, in InsP 5-ptase plants defense responses upon wounding were attenuated, whereas to our surprise in ipk1-1 plants defense responses were increased. Yeast-two-hybrid experiments revealed that in ipk1-1 plants accumulation of the InsP6-precursor, inositol pentakisphosphate (InsP5), promoted the interaction of COI1 with its target, JAZ9. The enhanced defensive capabilities of ipk1-1 plants might, thus, be due to enhanced degradation of the JAZ9 transcriptional repressor and increased defense gene expression and defense as well as general hypersensitivity of ipk1-1 plants to JA. An alanine substitution screen targeting amino acid residues of COI1 likely involved in the interaction of COI1 with its inositol polyphosphate cofactor yielded coi1 variants with progressively lower capability to interact with JAZ9 in yeast-two-hybrid tests and similarly lower capability to rescue the coi1 mutant phenotype in transgenic Arabidopsis plants. The data are consistent with the notion that InsP5 - not InsP6 - is important for COI1 function. The focus on defense responses and JA-signaling in consequence of only one position receiving support so far precluded us from obtaining more conclusive data on the possible relevance of InsP6 for TIR1 function in auxin responses. In a parallel non-directed global approach transcript arrays were used to determine transcripts induced or repressed in a PI-dependent manner upon gravistimulation or wounding. To this end, wild type controls, InsP 5-ptase plants and ipk1-1 plants were gravistimulated or wounded, and transcript abundance was monitored. The data indicate a large number of stimulus-specific genes that were induced in wild type controls but not or only weakly in InsP 5-ptase plants or in ipk1-1 plants. The transcript changes observed represent the direct read-out of TIR1 and COI1 function in the respective lines and support the results from the targeted approach. The transcript patterns furthermore suggest numerous new gene clusters linked to PI-signaling that invite further study. Overall, the data obtained in this project indicate that PI-derived inositol polyphosphates have an important role as cofactors for plant-typical phytohormone receptors, such as TIR1 and COI1, and are important for stimulus-induced transcript accumulation. The observation by our group and others that TIR1 contains InsP6 as a cofactor, whereas COI1 requires InsP5 to function suggests that the balance between InsP6 and InsP5 is important to the way plants perceive and respond to their environment. Experiments conceived for the planned extension of this project will aim at elucidating this question in the context of plant responses to pathogens, which involve both auxin and JA signaling. Furthermore, we are excited to have found new links of PI-signaling in the non-targeted approach, e.g. to signalling factors controlling stem-cell differentiation, that we hope to explore in the future.
Projektbezogene Publikationen (Auswahl)
- (2008) Phosphoinositide and inositolpolyphosphate-signalling in defence responses of Arabidopsis thaliana challenged by mechanical wounding. Mol Plant 1: 249-261
Mosblech A, König S, Stenzel I, Grzeganek P, Feussner I, Heilmann I
- (2009) Oxylipins: Structurally diverse metabolites from fatty acid oxidation. Plant Physiol Biochem 47: 511-517
Mosblech A, Feussner I, Heilmann I
- (2009) Reassessing the role of phospholipase D in the Arabidopsis wounding response. Plant Cell Environ 32: 837-850
Bargmann BO, Laxalt AM, Riet B, Testerink C, Merquiol E, Mosblech A, Reyes AL, Pieterse CM, Haring MA, Heilmann I, Bartels D, Munnik T
- (2010) Oxylipin signaling and plant growth. In Plant Lipid Signaling, Plant Cell Monographs, T Munnik, ed., Springer Academic Publishers, Berlin/Heidelberg, pp. 277-291
Mosblech A, Feussner I, Heilmann I
- (2011) Jasmonic acid perception by COI1 involves inositol polyphosphates in Arabidopsis thaliana. Plant J 65: 949-957
Mosblech A, Thurow C, Gatz C, Feussner I, Heilmann I
