Two-component elements mediate interactions between cytokinin and salicylic acid in plant immunity.

Recent studies have revealed an important role for hormones in plant immunity. We are now beginning to understand the contribution of crosstalk among different hormone signaling networks to the outcome of plant-pathogen interactions. Cytokinins are plant hormones that regulate development and responses to the environment. Cytokinin signaling involves a phosphorelay circuitry similar to two-component systems used by bacteria and fungi to perceive and react to various environmental stimuli. In this study, we asked whether cytokinin and components of cytokinin signaling contribute to plant immunity. We demonstrate that cytokinin levels in Arabidopsis are important in determining the amplitude of immune responses, ultimately influencing the outcome of plant-pathogen interactions. We show that high concentrations of cytokinin lead to increased defense responses to a virulent oomycete pathogen, through a process that is dependent on salicylic acid (SA) accumulation and activation of defense gene expression. Surprisingly, treatment with lower concentrations of cytokinin results in increased susceptibility. These functions for cytokinin in plant immunity require a host phosphorelay system and are mediated in part by type-A response regulators, which act as negative regulators of basal and pathogen-induced SA-dependent gene expression. Our results support a model in which cytokinin up-regulates plant immunity via an elevation of SA-dependent defense responses and in which SA in turn feedback-inhibits cytokinin signaling. The crosstalk between cytokinin and SA signaling networks may help plants fine-tune defense responses against pathogens.

Environmental perception avenues: the interaction of cytokinin and environmental response pathways.

Cytokinins were discovered in the 1950s by their ability to promote cell division in cultured plant cells. Recently, there have been significant breakthroughs in our understanding of the biosynthesis, metabolism, perception and signal transduction of this phytohormone. These advances, coupled with physiological and other approaches, have enabled remarkable progress to be made in our understanding of the interactions between cytokinin function and environmental inputs. In this review, we first highlight the most recent advances in our understanding of cytokinin biosynthesis, metabolism and signalling. We then discuss how various environmental signals interact with these pathways to modulate plant growth, development and physiology.

Reduction of plastid-localized carbonic anhydrase activity results in reduced Arabidopsis seedling survivorship.

Carbonic anhydrase (CA; EC 4.2.1.1) catalyzes the interconversion of CO2 and HCO3(-) and is a major protein constituent of the C3 higher plant chloroplast where it is presumed to play a role in photosynthetic carbon assimilation. In this study, we have used both RNA antisense and gene knockout lines to specifically reduce the activity of the chloroplast betaCA1 polypeptide (At3g01500) in the model plant Arabidopsis (Arabidopsis thaliana). Although able to germinate, seedling establishment of transgenic plants is significantly reduced relative to wild-type plants when grown at ambient levels of CO2. Growth at elevated (1,500 microL L(-1)) CO2 or on plates supplemented with sucrose restores seedling establishment rates to wild-type levels. Seed from wild-type and transgenic plants exhibited no significant differences in seed protein, lipid content, or reserve mobilization during seedling growth. betaCA1-deficient seedlings do, however, exhibit reduced capacity for light-dependent 14CO2 assimilation prior to the development of true leaves. The small number of surviving seedlings able to grow and develop are phenotypically similar to wild-type plants, even when subsequently grown at subambient levels of CO2. Microarray analysis of mature leaves of betaCA1-deficient plants shows some differences in transcript abundance, particularly with genes involved in ethylene signaling and response. The data suggest that reduced levels of seedling establishment by betaCA1-deficient plants could be the result of poor cotyledon photosynthetic performance at the onset of phototrophic growth and prior to the development of true leaves.

Cytokinin regulates type-A Arabidopsis Response Regulator activity and protein stability via two-component phosphorelay.

The plant hormone cytokinin regulates many aspects of growth and development. Cytokinin signaling involves His kinase receptors that perceive cytokinin and transmit the signal via a multistep phosphorelay similar to bacterial two-component signaling systems. The final targets of this phosphorelay are a set of Arabidopsis thaliana Response Regulator (ARR) proteins containing a receiver domain with a conserved Asp phosphorylation site. One class of these, the type-A ARRs, are negative regulators of cytokinin signaling that are rapidly transcriptionally upregulated in response to cytokinin. In this study, we tested the role of phosphorylation in type-A ARR function. Our results indicate that phosphorylation of the receiver domain is required for type-A ARR function and suggest that negative regulation of cytokinin signaling by the type-A ARRs most likely involves phosphorylation-dependent interactions. Furthermore, we show that a subset of the type-A ARR proteins are stabilized in response to cytokinin in part via phosphorylation. These studies shed light on the mechanism by which type-A ARRs act to negatively regulate cytokinin signaling and reveal a novel mechanism by which cytokinin controls type-A ARR function.

A subset of Arabidopsis AP2 transcription factors mediates cytokinin responses in concert with a two-component pathway.

The plant hormone cytokinin regulates numerous growth and developmental processes. A signal transduction pathway for cytokinin has been elucidated that is similar to bacterial two-component phosphorelays. In Arabidopsis, this pathway is comprised of receptors that are similar to sensor histidine kinases, histidine-containing phosphotransfer proteins, and response regulators (ARRs). There are two classes of response regulators, the type-A ARRs, which act as negative regulators of cytokinin responses, and the type-B ARRs, which are transcription factors that play a positive role in mediating cytokinin-regulated gene expression. Here we show that several closely related members of the Arabidopsis AP2 gene family of unknown function are transcriptionally up-regulated by cytokinin through this pathway, and we have designated these AP2 genes CYTOKININ RESPONSE FACTORS (CRFs). In addition to their transcriptional regulation by cytokinin, the CRF proteins rapidly accumulate in the nucleus in response to cytokinin, and this relocalization depends on the histidine kinases and the downstream histidine-containing phosphotransfer proteins, but is independent of the ARRs. Analysis of loss-of-function mutations reveals that the CRFs function redundantly to regulate the development of embryos, cotyledons, and leaves. Furthermore, the CRFs mediate a large fraction of the transcriptional response to cytokinin, affecting a set of cytokinin-responsive genes that largely overlaps with type-B ARR targets. These results indicate that the CRF proteins function in tandem with the type-B ARRs to mediate the initial cytokinin response. Thus, the evolutionarily ancient two-component system that is used by cytokinin branches to incorporate a unique family of plant-specific transcription factors.

Cytokinin signaling.

Cytokinins influence many aspects of plant growth and development. The current model for cytokinin signaling is a multi-step phosphorelay similar to the prokaryotic two-component systems that are used in responses to environmental stimuli. Recently, progress has been made in improving our understanding of the molecular mechanism that underlies cytokinin signaling. Molecular and genetic analyses of loss-of-function mutants indicate that the two-component elements that are involved in cytokinin signaling have redundant and overlapping functions. These elements regulate both the shoot and root meristems, are required for the development of fertile flowers, and modulate the response to varying nutrient levels.

Type-A Arabidopsis response regulators are partially redundant negative regulators of cytokinin signaling.

Type-A Arabidopsis (Arabidopsis thaliana) response regulators (ARRs) are a family of 10 genes that are rapidly induced by cytokinin and are highly similar to bacterial two-component response regulators. We have isolated T-DNA insertions in six of the type-A ARRs and constructed multiple insertional mutants, including the arr3,4,5,6,8,9 hextuple mutant. Single arr mutants were indistinguishable from the wild type in various cytokinin assays; double and higher order arr mutants showed progressively increasing sensitivity to cytokinin, indicating functional overlap among type-A ARRs and that these genes act as negative regulators of cytokinin responses. The induction of cytokinin primary response genes was amplified in arr mutants, indicating that the primary response to cytokinin is affected. Spatial patterns of ARR gene expression were consistent with partially redundant function of these genes in cytokinin signaling. The arr mutants show altered red light sensitivity, suggesting a general involvement of type-A ARRs in light signal transduction. Further, morphological phenotypes of some arr mutants suggest complex regulatory interactions and gene-specific functions among family members.