Cytokinin response factors regulate PIN-FORMED auxin transporters

Auxin and cytokinin are key endogenous regulators of plant development. Although cytokinin-mediated modulation of auxin distribution is a developmentally crucial hormonal interaction, its molecular basis is largely unknown. Here we show a direct regulatory link between cytokinin signalling and the auxin transport machinery uncovering a mechanistic framework for cytokinin-auxin cross-talk. We show that the CYTOKININ RESPONSE FACTORS (CRFs), transcription factors downstream of cytokinin perception, transcriptionally control genes encoding PIN-FORMED (PIN) auxin transporters at a specific PIN CYTOKININ RESPONSE ELEMENT (PCRE) domain. Removal of this cis-regulatory element effectively uncouples PIN transcription from the CRF-mediated cytokinin regulation and attenuates plant cytokinin sensitivity. We propose that CRFs represent a missing cross-talk component that fine-tunes auxin transport capacity downstream of cytokinin signalling to control plant development

Ethylene-independent promotion of photomorphogenesis in the dark by cytokinin requires COP1 and the CDD complex

The transition of skotomorphogenesis to photomorphogenesis is induced by the perception of light, and is characterized by the inhibition of hypocotyl elongation and opening of cotyledons. Although it is known that the plant hormone cytokinin inhibits hypocotyl elongation in dark-grown Arabidopsis plants when applied in high concentrations, it is unclear to what extent this response is the result of cytokinin alone or cytokinin-induced ethylene production. Here, we show that cytokinin-induced inhibition of hypocotyl elongation is largely independent of ethylene and suggest a close connection between the cytokinin two-component system and the light-signaling networks. We show that this cytokinin signal is mainly mediated through the cytokinin receptor ARABIDOPSIS HISTIDINE KINASE3 and the ARABIDOPSIS RESPONSE REGULATOR1 in combination with ARR12. Interestingly, mutation of CONSTITUTIVELY PHOTOMORPOGENIC1 (COP1), DE-ETIOLATED1, and CYTOKININ INSENSITIVE4/COP10 renders plants insensitive to cytokinin, and these factors are indispensable for the transcriptional response during cytokinin-induced de-etiolation, indicating that a functional light-signaling pathway is essential for this cytokinin response. In addition, the effect of cytokinin on hypocotyl elongation is strongly dependent on the light conditions, with higher light intensities causing a switch in the response to cytokinin from an inhibitor to a promoter of hypocotyl elongation

The transcription factors BEL1 and SPL are required for cytokinin and auxin signaling during ovule development in Arabidopsis

Hormones, such as auxin and cytokinin, are involved in the complex molecular network that regulates the coordinated development of plant organs. Genes controlling ovule patterning have been identified and studied in detail; however, the roles of auxin and cytokinin in ovule development are largely unknown. Here we show that key cytokinin pathway genes, such as isopentenyltransferase and cytokinin receptors, are expressed during ovule development. Also, in a cre1-12 ahk2-2 ahk3-3 triple mutant with severely reduced cytokinin perception, expression of the auxin efflux facilitator PIN-FORMED 1 (PIN1) was severely reduced. In sporocyteless/nozzle (spl/nzz) mutants, which show a similar phenotype to the cre1-12 ahk2-2 ahk3-3 triple mutant, PIN1 expression is also reduced. Treatment with the exogenous cytokinin N-6-benzylaminopurine also altered both auxin distribution and patterning of the ovule; this process required the homeodomain transcription factor BELL1 (BEL1). Thus, this article shows that cytokinin regulates ovule development through the regulation of PIN1. Furthermore, the transcription factors BEL1 and SPL/NZZ, previously described as key regulators of ovule development, are needed for the auxin and cytokinin signaling pathways for the correct patterning of the ovule

Root enhancement in cytokinin-deficient oilseed rape causes leaf mineral enrichment, increases the chlorophyll concentration under nutrient limitation and enhances the phytoremediation capacity

Background Cytokinin is a negative regulator of root growth, and a reduction of the cytokinin content or signalling causes the formation a larger root system in model plants, improves their growth under drought and nutrient limitation and causes increased accumulation of elements in the shoot. Roots are an important but understudied target of plant breeding. Here we have therefore explored whether root enhancement by lowering the cytokinin content can also be achieved in oilseed rape (Brassica napus L.) plants. Results Transgenic plants overexpressing the CKX2 gene of Arabidopsis thaliana encoding a cytokinin-degrading cytokinin oxidase/dehydrogenase showed higher CKX activity and a strongly reduced cytokinin content. Cytokinin deficiency led to the formation of a larger root system under different growth conditions, which was mainly due to an increased number of lateral and adventitious roots. In contrast, shoot growth was comparable to wild type, which caused an enhanced root-to-shoot ratio. Transgenic plants accumulated in their leaves higher concentrations of macro- and microelements including P, Ca, Mg, S, Zn, Cu, Mo and Mn. They formed more chlorophyll under Mg- and S-deficiency and accumulated a larger amount of Cd and Zn from contaminated medium and soil. Conclusions These findings demonstrate the usefulness of ectopic CKX gene expression to achieve root enhancement in oilseed rape and underpin the functional relevance of a larger root system. Furthermore, the lack of major developmental consequences on shoot growth in cytokinin-deficient oilseed rape indicates species-specific differences of CKX gene and/or cytokinin action

Cytokinin response factor 6 represses cytokinin-associated genes during oxidative stress

Cytokinin is a phytohormone that is well known for its roles in numerous plant growth and developmental processes, yet it has also been linked to abiotic stress response in a less defined manner. Arabidopsis (Arabidopsis thaliana) Cytokinin Response Factor 6 (CRF6) is a cytokinin-responsive AP2/ERF-family transcription factor that, through the cytokinin signaling pathway, plays a key role in the inhibition of dark-induced senescence. CRF6 expression is also induced by oxidative stress, and here we show a novel function for CRF6 in relation to oxidative stress and identify downstream transcriptional targets of CRF6 that are repressed in response to oxidative stress. Analysis of transcriptomic changes in wild-type and crf6 mutant plants treated with H2O2 identified CRF6-dependent differentially expressed transcripts, many of which were repressed rather than induced. Moreover, many repressed genes also show decreased expression in 35S:CRF6 overexpressing plants. Together, these findings suggest that CRF6 functions largely as a transcriptional repressor. Interestingly, among the H2O2 repressed CRF6-dependent transcripts was a set of five genes associated with cytokinin processes: (signaling) ARR6, ARR9, ARR11, (biosynthesis) LOG7, and (transport) ABCG14. We have examined mutants of these cytokinin-associated target genes to reveal novel connections to oxidative stress. Further examination of CRF6-DNA interactions indicated that CRF6 may regulate its targets both directly and indirectly. Together, this shows that CRF6 functions during oxidative stress as a negative regulator to control this cytokinin-associated module of CRF6-dependent genes and establishes a novel connection between cytokinin and oxidative stress response

Regulation of Leaf Senescence by Cytokinin, Sugars, and Light. Effects on NADH-Dependent Hydroxypyruvate Reductase

The aim of this study was to investigate the interactions between cytokinin, sugar repression, and light in the senescence-related decline in photosynthetic enzymes of leaves. In transgenic tobacco (Nicotiana tabacum) plants that induce the production of cytokinin in senescing tissue, the age-dependent decline in NADH-dependent hydroxypyruvate reductase (HPR), ribulose-1,5-bisphosphate carboxylase/oxygenase, and other enzymes involved in photosynthetic metabolism was delayed but not prevented. Glucose (Glc) and fructose contents increased with leaf age in wild-type tobacco and, to a greater extent, in transgenic tobacco. To study whether sugar accumulation in senescing leaves can counteract the effect of cytokinin on senescence, discs of wild-type leaves were incubated with Glc and cytokinin solutions. The photorespiratory enzyme HPR declined rapidly in the presence of 20 mM Glc, especially at very low photon flux density. Although HPR protein was increased in the presence of cytokinin, cytokinin did not prevent the Glc-dependent decline. Illumination at moderate photon flux density resulted in the rapid synthesis of HPR and partially prevented the negative effect of Glc. Similar results were obtained for the photosynthetic enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase. It is concluded that sugars, cytokinin, and light interact during senescence by influencing the decline in proteins involved in photosynthetic metabolism

Multiple feedback loops through cytokinin signaling control stem cell number within the Arabidopsis shoot meristem

A central unanswered question in stem cell biology, both in plants and in animals, is how the spatial organization of stem cell niches are maintained as cells move through them. We address this question for the shoot apical meristem (SAM) which harbors pluripotent stem cells responsible for growth of above-ground tissues in flowering plants. We find that localized perception of the plant hormone cytokinin establishes a spatial domain in which cell fate is respecified through induction of the master regulator WUSCHEL as cells are displaced during growth. Cytokinin-induced WUSCHEL expression occurs through both CLAVATA-dependent and CLAVATA-independent pathways. Computational analysis shows that feedback between cytokinin response and genetic regulators predicts their relative patterning, which we confirm experimentally. Our results also may explain how increasing cytokinin concentration leads to the first steps in reestablishing the shoot stem cell niche in vitro

The Effects of Cytokinin on the Transcriptional Regulation of PIN Expression in Arabidopsis thaliana

The processes of cell division and differentiation are critical to the development of any multicellular organism. During the formation of plant roots these processes take place at a region of the root tip called the meristem. Cytokinin and auxin are two plant growth hormones that influence this process. Although these two growth hormones are both necessary they also appear in many ways to have an antagonistic relationship. As meristematic root cells undergo differentiation they cease dividing. It has been proposed that the size of the root meristem and thus the overall rate of root growth are determined by the balance between the rate of cell division, determined by auxin, and the rate of cell differentiation, determined by cytokinin. One of the ways cytokinin may have an antagonistic influence on auxin regulation is by limiting auxin transport. Directed auxin transport from one part of a plant to another is controlled by a family of auxin efflux proteins called PIN proteins. To date, studies reported in the literate are inconsistent as to whether and how cytokinin influences PIN protein production at the transcriptional and/or translational level. This study was undertaken to explore the effect of cytokinin signaling on PIN gene expression using real-time quantitative PCR

Induction of Cytokinin-Independent Tobacco Tissues by Substituted Fluorenes

Two morphactins and three aminofluorenes initiated the formation of compact tissue nodules in hormone-dependent tobacco callus cultures. These nodules, upon subculture, behaved like partially transformed plant tumors. They grew on cytokinin-free media, while control callus and nonnodule tissue still required an exogenous cytokinin source. The data indicate that substituted fluorenes, including carcinogenic aminofluorenes, can cause a neoplastic growth response in cultured tobacco tissues. Of particular interest in this study is the finding that a directed and heritable cellular change is induced in tobacco tissue in which a specific (the endogenous cytokinin) biosynthetic system is regularly and persistently activated

Arabidopsis ABCG14 is essential for the root-to-shoot translocation of cytokinin.

Cytokinins are phytohormones that induce cytokinesis and are essential for diverse developmental and physiological processes in plants. Cytokinins of the trans-zeatin type are mainly synthesized in root vasculature and transported to the shoot, where they regulate shoot growth. However, the mechanism of long-distance transport of cytokinin was hitherto unknown. Here, we report that the Arabidopsis ATP-binding cassette (ABC) transporter subfamily G14 (AtABCG14) is mainly expressed in roots and plays a major role in delivering cytokinins to the shoot. Loss of AtABCG14 expression resulted in severe shoot growth retardation, which was rescued by exogenous trans-zeatin application. Cytokinin content was decreased in the shoots of atabcg14 plants and increased in the roots, with consistent changes in the expression of cytokinin-responsive genes. Grafting of atabcg14 scions onto wild-type rootstocks restored shoot growth, whereas wild-type scions grafted onto atabcg14 rootstocks exhibited shoot growth retardation similar to that of atabcg14. Cytokinin concentrations in the xylem are reduced by similar to 90% in the atabcg14 mutant. These results indicate that AtABCG14 is crucial for the translocation of cytokinin to the shoot. Our results provide molecular evidence for the long-distance transport of cytokinin and show that this transport is necessary for normal shoot development.open118380Ysciescopu