75 Integrating Membrane Transport with Male Gametophyte ... - TAIR

403 Association and Localization of PP2C Phosphatase/Stress-Activated MAP Kinase Complexes
in Plant Cells
Irute Meskiene, Vaiva Kazanaviciute, Alois Schweighofer
Max F. Perutz Laboratories, University of Vienna, Vienna Biocenter, Dr. Bohrgasse 9, A-1030 Vienna, Austria
Mitogen-activated protein kinase (MAPK) pathways are commonly used to transmit environmental signals and
integrate cellular responses in eukaryotes. Specific classes of protein phosphatases are employed to inactivate MAPKs,
thereby influencing MAPKs activation kinetics (1). PP2Cs are ubiquitous protein phosphatases found in all eukaryotes,
where they participate in a large array of signaling pathways. This diversity of biological functions is reflected in the
great expansion of the PP2Cs family in Arabidopsis (2). We show by complementary approaches that AP2C1, a member
of the Arabidopsis PP2C family, can specifically interact with and inactivate the stress-responsive MAPKs.
Direct protein-protein interaction in yeast and plant cells suggest high specificity in association between MAP kinases
and phosphatase. We found that specific kinase interaction motif (KIM) identified in silico in AP2C1 is responsible for
protein-protein interaction with substrate kinases. A similar KIM is found in yeast and animal MAPK interacting proteins,
such as MAPKKs or MAPK phosphatases, as well as in plant MAPKKs.
Localization of AP2C1 induction in transgenic AP2C1::GUS plants upon biotic and abiotic stress additionally supports
its role in control of stress-activated MAPK. We show that AP2C1 and MAPKs associate into a complex in the nucleus
or nucleus/cytoplasm, dependent on the MAPK involved by application of a bimolecular fluorescence complementation
(BiFC) assay based on split-YFP (3).
1. Meskiene, I., Baudouin, E., Schweighofer, A., Liwosz, A., Jonak, C., Rodriguez, P.L., Jelinek, H. and Hirt, H. (2003) The Stress-induced
protein phosphatase 2C is a negative regulator of a mitogen-activated protein kinase. J Biol Chem, 278, 18945-18945.
2. Schweighofer, A., Hirt, H. and Meskiene, I. (2004) Plant PP2C phosphatases: emerging functions in stress signaling. Trends Plant Sci, 9, 236-
243.
3. Walter, M., Chaban, C., Schutze, K., Batistic, O., Weckermann, K., Nake, C., Blazevic, D., Grefen, C., Schumacher, K., Oecking, C., Harter,
K. and Kudla, J. (2004) Visualization of protein interactions in living plant cells using bimolecular fluorescence complementation. Plant J, 40,
428-438.
404 Interactions Between Light and Auxin Signaling Networks
Jennifer Moon, Hui Shen, Ling Zhu, Nidhi Sharma, Enamul Huq
University of Texas-Austin, Austin, TX 78712
Plant growth and development is modulated by the cues plants receive from their environment. Many individual
signaling pathways, such as light or auxin signaling, have been extensively investigated. However, very little is known
about how these signaling pathways are integrated to optimize plant development. We are interested in understanding
the mechanism(s) of light and auxin signal integration. Previously, genetic and physical interaction has been shown
between phytochrome (phy) light receptors and AUX/IAA proteins, transcriptional regulators of auxin response genes.
To investigate whether the stability of AUX/IAA proteins is regulated by light signaling, we used full-length IAA3 or
IAA4 fused to firefly Luciferase (LUC) driven by a strong Cauliflower mosaic virus (CaMV35S) promoter in transient
transfection assays in Arabidopsis seedlings. We found that the luciferase activity of LUC-IAA3 and LUC-IAA4 was
strongly reduced in red (Rc) and far red (FRc) light compared to the dark control, while the LUC-GFP control remained
unchanged in all light conditions. Further, seedlings expressing only domains I and II of AUX/IAA17 fused to GUS
(IAA17NT-GUS) show reduced GUS activity in Rc and FRc compared to the dark control, suggesting light signaling,
like auxin signaling, may promote AUX/IAA degradation through domain II. Plants expressing GUS driven by an auxin
response promoter, BA3, show reduced GUS activity in Rc and FRc indicating reduced auxin level/signaling in these
light conditions. These results suggest that phy signaling modulates the abundance of AUX/IAA proteins directly or
indirectly to control seedling deetiolation in response to light, and thus suggests a potential molecular mechanism for
interactions between the light and auxin signaling pathways.