Protocol for generating chemically compotent (ie. for heat shock transformation) E. coli cells for transformation (e.g. heterologous protein expression, cloning).

Distinct ROS signaling pathways initiated by singlet oxygen ((1)O2) or superoxide and hydrogen peroxide have been attributed to either cell death or acclimation, respectively. Recent studies have revealed that more complex antagonistic and synergistic relationships exist within and between these pathways. As specific chloroplastic ROS signals are difficult to study, rapid systemic signaling experiments using localized high light (HL) stress or ROS treatments were used in this study to uncouple signals required for direct HL and ROS perception and distal systemic acquired acclimation (SAA). A qPCR approach was chosen to determine local perception and distal signal reception. Analysis of a thylakoidal ascorbate peroxidase mutant (tapx), the (1)O2-retrograde signaling double mutant (ex1/ex2), and an apoplastic signaling double mutant (rbohD/F) revealed that tAPX and EXECUTER 1 are required for both HL and systemic acclimation stress perception. Apoplastic membrane-localized RBOHs were required for systemic spread of the signal but not for local signal induction in directly stressed tissues. Endogenous ROS treatments revealed a very strong systemic response induced by a localized 1 h induction of (1)O2 using the conditional flu mutant. A qPCR time course of (1)O2 induced systemic marker genes in directly and indirectly connected leaves revealed a direct vascular connection component of both immediate and longer term SAA signaling responses. These results reveal the importance of an EXECUTER-dependent (1)O2 retrograde signal for both local and long distance RBOH-dependent acclimation signaling that is distinct from other HL signaling pathways, and that direct vascular connections have a role in spatial-temporal SAA induction.

In recent years many advances have been made to obtain insight into chloroplast biogenesis and development. In plants several plastids types exist such as the proplastid (which is the progenitor of all plastids), leucoplasts (group of colourless plastids important for storage including elaioplasts (lipids), amyloplasts (starch) or proteinoplasts (proteins)), chromoplasts (yellow to orange-coloured due to carotenoids, in flowers or in old leaves as gerontoplasts), and the green chloroplasts. Chloroplasts are indispensable for plant development; not only by performing photosynthesis and thus rendering the plant photoautotrophic, but also for biochemical processes (which in some instances can also take place in other plastids types), such as the synthesis of pigments, lipids, and plant hormones and sensing environmental stimuli. Although we understand many aspects of these processes there are gaps in our understanding of the establishment of functional chloroplasts and their regulation. Why is that so? Even though chloroplast function is comparable in all plants and most of the algae, ferns and moss, detailed analyses have revealed many differences, specifically with respect to its biogenesis. As an update to our prior review on the genetic analysis of chloroplast biogenesis and development [1] herein we will focus on recent advances in Angiosperms (monocotyledonous and dicotyledonous plants) that provide novel insights and highlight the challenges and prospects for unravelling the regulation of chloroplast biogenesis specifically during the establishment of the young plants. This article is part of a Special Issue entitled: Chloroplast Biogenesis.

Plant development is regulated by external and internal factors such as light and chloroplast development. A revertant of the Arabidopsis thaliana (L.) Heyhn. chloroplast biogenesis mutant snowy cotyledon 3 (sco3-1) was isolated partially recovering the impaired chloroplast phenotype. The mutation was identified in the Phytochrome B (PhyB) gene and is a result of an amino acid change within the PAS repeat domain required for light-induced nuclear localisation. An independent phyB-9 mutation was crossed into sco3-1 mutants, resulting in the same partial reversion of sco3-1. Further analysis demonstrated that SCO3 and PhyB influence the greening process of seedlings and rosette leaves, embryogenesis, rosette formation and flowering. Interestingly, the functions of these proteins are interwoven in various ways, suggesting a complex genetic interaction. Whole-transcriptome profiling of sco3-1phyB-9 indicated that a completely distinct set of genes was differentially regulated in the double mutant compared with the single sco3-1 or phyB-9 mutants. Thus, we hypothesise that PhyB and SCO3 genetically suppress each other in plant and chloroplast development.

Reidunn Birgitta Aalen Javier Abadia Ibrokhim Y. Abdurakhmonov Ikuro Abe Steffen Abel Gian Paolo Accotto Jose Luis Acebes Keith L. Adams Joshua Adams Ahmed Afzal Khairulmazmi Ahmad Elizabeth Ainsworth Eduard Akhunov Kiba Akinori Emre Aksoy Emidio Albertini Veronica Albrecht-Borth Ruben Alcazar Jameel Al-Haddad Karine Alix James William Allwood Tancrède Alméras Maria Elena Alvarez Iraida Amaya Anna Amtmann Gynheung An Anne J. Anderson Louise E. Anderson James V. Anderson Charles T. Anderson Jill T. Anderson Mats X. Andersson John Andralojc Gerco C. Angenent Klaus Apel Miguel A. Aranda Jose Luis Araus Cris M. Argueso Shin-ichi Arimura Idoia Ariz Charles L. Armstrong Susan J. Armstrong Eva-Mari Aro Han Asard Motoyuki Ashikari Sarah M. Assmann Brian J. Atwell Sylvain Aubry Kris Audenaert Koichiro Awai Michael Axtell Thomas J. Bach Tony Bacic Eric Badel Murray Badger Stephen Baenziger Scott Baerson Sacha Baginsky Harsh Pal Bais Soren Bak Neil R. Baker Ian T. Baldwin Steven G. Ball Marilyn C. Ball Carlos L. Ballare Jennifer Baltzer Frantisek Baluska Roberto Barbato Marie Barberon Margaret M. Barbour Gianni Barcaccia Alice Barkan Fredy Barneche Cornelius Barry Bonnie Bartel Dorothea Bartels Madelaine Bartlett Kathy Barton Vitthal T. Barvkar Dudy Bar-Zvi George Bassel Gilles Basset Alfred Batschauer Petra Bauer David Baum Hermann Bauwe Ivan R. Baxter Martin Bayer Michael H. Beale Nathalie Beaudoin Frederic Beaudoin Ulrike Bechtold Dirk Becker Jörg D. Becker Diane M. Beckles Philip W. Becraft Sebastian Bednarek Pawel Bednarek Tom Beeckman Gerrit T.S. Beemster Eric P. Beers Christophe Belin Catherine Bellini Philip N. Benfey Eva Benkova Malcolm J. Bennett Tom Bennett Frederic Berger Susanne Berger John A. Berges Gerald Berkowitz Oliver Berkowitz Thomas Berleth Carl Bernacchi Mark A. Bernards Christine A. Beveridge Rishikesh Bhalerao Hongwu Bian Tatiana N. Bibikova Brad Binder James A. Birchler Kenneth D. Birnbaum Sherryl R. Bisgrove Crysten Blaby Ian Blaby Elison B. Blancaflor Mike R. Blatt Andreas Blennow Claudia Blindauer Arnold Bloom Yaroslav B. Blume Leonor C. Boavida Jens Boch Laszlo Bogre Joerg Bohlmann Cordelia Bolle Vera Bonardi Atle M. Bones Paola Bonfante Anne M. Borland Frederik Börnke Jan-Willem Borst Paul K. Boss Rebecca S. Boston Firas Bou Daher Marie Boudsocq Frédéric Bourgaud Ralph Bours Yohann Boutte Mondher Bouzayen John L. Bowman Caroline Bowsher Nanette R. Boyle Janet Braam Kent J. Bradford Siobhan M. Brady Peter M. Bramley Federica Brandizzi Benjamin Brandt Hans-Peter Braun David M. Braun Melissa Brazier-Hicks Volker Brendel Oliver Brendel Eric D. Brenner Jean-Francois Briat Winslow R. Briggs Anne Bagg Britt Myron Bruce Robert Brueggeman David A. Brummell Glenn J. Bryan Bretislav Brzobhaty Bob Buchanan Peter Buchner Thomas Buckhout Thomas N. Buckley

Chloroplasts of leaves under high light stress initiate signals to the nuclei of both exposed and distal leaves in order to acclimate against the potential threat of oxidative damage: a process known as high light systemic acquired acclimation (HL SAA). This study explores the nature of HL SAA, synergistic interactions with other environmental stresses, and the impact of repeated HL stress on the acclimation response of exposed and distal leaves. This necessitated the development of novel experimental systems to investigate the initiation, perception, and response to HL SAA. These systems were used to investigate the HL SAA response by monitoring the induction of mRNA in distal leaves not exposed to the HL stress. Acclimation to HL is induced within minutes and the response is proportionally dependent on the quality and quantity of light. HL SAA treatments in conjunction with variations in temperature and humidity reveal HL SAA is influenced by fluctuations in humidity. These treatments also result in changes in auxin accumulation and auxin-responsive genes. A key question in retrograde signaling is the extent to which transient changes in light intensity result in a “memory” of the event leading to acclimation responses. Repeated exposure to short term HL resulted in acclimation of the exposed tissue and that of emerging and young leaves (but not older leaves) to HL and oxidative stress.

A plant-specific chloroplast endopeptidase is identified that functions in protection of the photosynthetic apparatus from photo-oxidative stress. Excess light can have a negative impact on photosynthesis; thus, plants have evolved many different ways to adapt to different light conditions to both optimize energy use and avoid damage caused by excess light. Analysis of the Arabidopsis (Arabidopsis thaliana) mutant snowy cotyledon4 (sco4) revealed a mutation in a chloroplast-targeted protein that shares limited homology with CaaX-type endopeptidases. The SCO4 protein possesses an important function in photosynthesis and development, with point mutations rendering the seedlings and adult plants susceptible to photooxidative stress. The sco4 mutation impairs the acclimation of chloroplasts and their photosystems to excess light, evidenced in a reduction in photosystem I function, decreased linear electron transfer, yet increased nonphotochemical quenching. SCO4 is localized to the chloroplasts, which suggests the existence of an unreported type of protein modification within this organelle. Phylogenetic and yeast complementation analyses of SCO4-like proteins reveal that SCO4 is a member of an unknown group of higher plant-specific proteinases quite distinct from the well-described CaaX-type endopeptidases RAS Converting Enzyme1 (RCE1) and zinc metallopeptidase STE24 and lacks canonical CaaX activity. Therefore, we hypothesize that SCO4 is a novel endopeptidase required for critical protein modifications within chloroplasts, influencing the function of proteins involved in photosynthesis required for tolerance to excess light.

Excess light can have a negative impact on photosynthesis; thus, plants have evolved many different ways to adapt to different light conditions to both optimize energy use and avoid damage caused by excess light. Analysis of the Arabidopsis (Arabidopsis thaliana) mutant snowy cotyledon4 (sco4) revealed a mutation in a chloroplast-targeted protein that shares limited homology with CaaX-type endopeptidases. The SCO4 protein possesses an important function in photosynthesis and development, with point mutations rendering the seedlings and adult plants susceptible to photooxidative stress. The sco4 mutation impairs the acclimation of chloroplasts and their photosystems to excess light, evidenced in a reduction in photosystem I function, decreased linear electron transfer, yet increased nonphotochemical quenching. SCO4 is localized to the chloroplasts, which suggests the existence of an unreported type of protein modification within this organelle. Phylogenetic and yeast complementation analyses of SCO4-like proteins reveal that SCO4 is a member of an unknown group of higher plant-specific proteinases quite distinct from the well-described CaaX-type endopeptidases RAS Converting Enzyme1 (RCE1) and zinc metallopeptidase STE24 and lacks canonical CaaX activity. Therefore, we hypothesize that SCO4 is a novel endopeptidase required for critical protein modifications within chloroplasts, influencing the function of proteins involved in photosynthesis required for tolerance to excess light.

The Arabidopsis thaliana genome contains two genes with homology to the mitochondrial protein LETM1 (leucine zipper-EF-hand-containing transmembrane protein). Inactivation of both genes, Atletm1 and Atletm2, together is lethal. Plants that are hemizygous for AtLETM2 and homozygous for Atletm1 (letm1(-/-) LETM2(+/-)) displayed a mild retarded growth phenotype during early seedling growth. It was shown that accumulation of mitochondrial proteins was reduced in hemizygous (letm1(-/-) LETM2(+/-)) plants. Examination of respiratory chain proteins by Western blotting, blue native PAGE, and enzymatic activity assays revealed that the steady state level of ATP synthase was reduced in abundance, whereas the steady state levels of other respiratory chain proteins remained unchanged. The absence of a functional maternal AtLETM2 allele in an Atletm1 mutant background resulted in early seed abortion. Reciprocal crosses revealed that maternally, but not paternally, derived AtLETM2 was absolutely required for seed development. This requirement for a functional maternal allele of AtLETM2 was confirmed using direct sequencing of reciprocal crosses of Col-0 and Ler accessions. Furthermore, AtLETM2 promoter beta-glucuronidase constructs displayed exclusive maternal expression patterns.

The chloroplast is essential for photosynthesis and the production of hormones and metabolites. As a consequence its biogenesis and development needs to be coordinated with seedling growth to ensure optimal rates of photosynthesis without oxidative damage upon seedling emergence. Studies into the processes of biogenesis and development of chloroplasts have shown how important chloroplast development during germination is for plant vitality, seed set and growth. Indeed, even if chloroplast development is only impaired in cotyledons, the embryonic leaves of germinating seedlings, with normal chloroplast development in true leaves plant growth and yield can be negatively impacted. Thus, it is necessary to understand the regulation and mechanism of chloroplast development to be able to further investigate on plant vitality for improving plants. This review will concentrate on how chloroplasts differentiate giving emphasis to current insights into the regulatory factors and molecular requirements for chloroplast biogenesis and development gained from genetic and mutagenic studies.

The process of chloroplast biogenesis requires a multitude of pathways and processes to establish chloroplast function. In cotyledons of seedlings, chloroplasts develop either directly from proplastids (also named eoplasts) or, if germinated in the dark, via etioplasts, whereas in leaves chloroplasts derive from proplastids in the apical meristem and are then multiplied by division. The snowy cotyledon 2, sco2, mutations specifically disrupt chloroplast biogenesis in cotyledons. SCO2 encodes a chloroplast-localized protein disulphide isomerase, hypothesized to be involved in protein folding. Analysis of co-expressed genes with SCO2 revealed that genes with similar expression patterns encode chloroplast proteins involved in protein translation and in chlorophyll biosynthesis. Indeed, sco2-1 accumulates increased levels of the chlorophyll precursor, protochlorophyllide, in both dark grown cotyledons and leaves. Yeast two-hybrid analyses demonstrated that SCO2 directly interacts with the chlorophyll-binding LHCB1 proteins, being confirmed in planta using bimolecular fluorescence complementation (BIFC). Furthermore, ultrastructural analysis of sco2-1 chloroplasts revealed that formation and movement of transport vesicles from the inner envelope to the thylakoids is perturbed. SCO2 does not interact with the signal recognition particle proteins SRP54 and FtsY, which were shown to be involved in targeting of LHCB1 to the thylakoids. We hypothesize that SCO2 provides an alternative targeting pathway for light-harvesting chlorophyll binding (LHCB) proteins to the thylakoids via transport vesicles predominantly in cotyledons, with the signal recognition particle (SRP) pathway predominant in rosette leaves. Therefore, we propose that SCO2 is involved in the integration of LHCB1 proteins into the thylakoids that feeds back on the regulation of the tetrapyrrole biosynthetic pathway and nuclear gene expression.

Here, we describe the snowy cotyledon3 (sco3-1) mutation, which impairs chloroplast and etioplast development in Arabidopsis thaliana seedlings. SCO3 is a member of a largely uncharacterized protein family unique to the plant kingdom. The sco3-1 mutation alters chloroplast morphology and development, reduces chlorophyll accumulation, impairs thylakoid formation and photosynthesis in seedlings, and results in photoinhibition under extreme CO2 concentrations in mature leaves. There are no readily apparent changes to chloroplast biology, such as transcription or assembly that explain the disruption to chloroplast biogenesis. Indeed, SCO3 is actually targeted to another organelle, specifically to the periphery of peroxisomes. However, impaired chloroplast development cannot be attributed to perturbed peroxisomal metabolic processes involving germination, fatty acid β-oxidation or photorespiration, though there are so far undescribed changes in low and high CO2 sensitivity in seedlings and young true leaves. Many of the chloroplasts are bilobed, and some have persistent membranous extensions that encircle other cellular components. Significantly, there are changes to the cytoskeleton in sco3-1, and microtubule inhibitors have similar effects on chloroplast biogenesis as sco3-1 does. The localization of SCO3 to the periphery of the peroxisomes was shown to be dependent on a functional microtubule cytoskeleton. Therefore, the microtubule and peroxisome-associated SCO3 protein is required for chloroplast development, and sco3-1, along with microtubule inhibitors, demonstrates an unexpected role for the cytoskeleton and peroxisomes in chloroplast biogenesis.

Plants have a range of mechanisms to protect against oxidative damage induced by excess light and environmental stress. One of these processes consists of the detoxification of reactive oxygen species by the ascorbate peroxidase (APX) family of enzymes, which convert H2O2 into H2O. Two of the genes encoding APX in Arabidopsis are induced by high light, namely APX1 and APX2. We have applied a genetic approach to understanding the mechanisms of photoprotection, using APX2 as an indicator of oxidative stress. Transgenic plants containing the reporter gene luciferase linked to the APX2 promoter were EMS mutagenized and kindly provided to us by Mullineaux and colleagues. We have screened this mutagenized seed to identify mutants with aberrant photoprotection, based on altered luminescence resulting from altered luciferase activity. Here we describe the screen and steps involved in the identification of the mutations in an effort to identify novel photoprotective genes and products.

In cotyledons of etiolated seedlings light-dependent transformation of etioplasts to chloroplasts marks the transition from heterotrophic to autotrophic growth. Genetic factors required for this developmental step were identified by isolating mutants of Arabidopsis thaliana that were impaired in chloroplast development in cotyledons but not in true leaves. Several mutants with chlorophyll-deficient cotyledons were isolated and dubbed snowy cotyledon (sco). Here we describe the identification and detailed characterization of the snowy cotyledon 2 mutant. The mutated SCO2 gene was identified using a map-based cloning strategy. SCO2 was shown to encode a novel protein which contains a single DnaJ-like zinc finger domain. The SCO2 protein fused to GFP was shown to be present in chloroplasts. Inactivation of SCO2 has almost no detectable impact on the levels of transcripts encoding plastid-specific proteins but leads to a significant reduction of plastid protein levels. Even though transcripts of SCO2 have been found ubiquitously in green tissues as well as in roots phenotypic changes due to SCO2 inactivation are confined to cotyledons. The cotyledons in embryos of sco2 are unaffected in their chloroplast biogenesis. Upon precocious germination seedlings of sco2 and wild type are indistinguishable. The SCO2 mutation affects chloroplast biogenesis only at the end of dormancy during seed germination. The transition from heterotrophic to autotrophic growth is dramatically impaired in sco2 when seedlings were kept in the dark for more than 5 days prior to light exposure.

An Arabidopsis thaliana leaf-variegated mutant yellow variegated2 (var2) results from loss of FtsH2, a major component of the chloroplast FtsH complex. FtsH is an ATP-dependent metalloprotease in thylakoid membranes and degrades several chloroplastic proteins. To understand the role of proteolysis by FtsH and mechanisms leading to leaf variegation, we characterized the second-site recessive mutation fu-gaeri1 (fug1) that suppressed leaf variegation of var2. Map-based cloning and subsequent characterization of the FUG1 locus demonstrated that it encodes a protein homologous to prokaryotic translation initiation factor 2 (cpIF2) located in chloroplasts. We show evidence that cpIF2 indeed functions in chloroplast protein synthesis in vivo. Suppression of leaf variegation by fug1 is observed not only in var2 but also in var1 (lacking FtsH5) and var1 var2. Thus, suppression of leaf variegation caused by loss of FtsHs is most likely attributed to reduced protein synthesis in chloroplasts. This hypothesis was further supported by the observation that another viable mutation in chloroplast translation elongation factor G also suppresses leaf variegation in var2. We propose that the balance between protein synthesis and degradation is one of the determining factors leading to the variegated phenotype in Arabidopsis leaves.

Intracellular release of calcium ions belongs to the earliest events in cellular stress perception. The molecular mechanisms integrating signals from different environmental cues and translating them into an optimized response are largely unknown. We report here the functional characterization of CIPK1, a protein kinase interacting strongly with the calcium sensors CBL1 and CBL9. Comparison of the expression patterns indicates that the three proteins execute their functions in the same tissues. Physical interaction of CIPK1 with CBL1 and CBL9 targets the kinase to the plasma membrane. We show that, similarly to loss of CBL9 function, mutation of either CBL1 or CIPK1 renders plants hypersensitive to osmotic stress. Remarkably, in contrast to the cbl1 mutant and similarly to the cbl9 mutant, loss of CIPK1 function impairs abscisic acid (ABA) responsiveness. We therefore suggest that, by alternative complex formation with either CBL1 or CBL9, the kinase CIPK1 represents a convergence point for ABA-dependent and ABA-independent stress responses. Based on our genetic, physiological and protein-protein interaction data, we propose a general model for information processing in calcium-regulated signalling networks.

During seedling development chloroplast formation marks the transition from heterotrophic to autotrophic growth. The development and activity of chloroplasts may differ in cotyledons that initially serve as a storage organ and true leaves whose primary function is photosynthesis. A genetic screen was used for the identification of genes that affect selectively chloroplast function in cotyledons of Arabidopsis thaliana. Several mutants exhibiting pale cotyledons and green true leaves were isolated and dubbed snowy cotyledon (sco).One of the mutants, sco1, was characterized in more detail. The mutated gene was identified using map-based cloning. The mutant contains a point mutation in a gene encoding the chloroplast elongation factor G, leading to an amino acid exchange within the predicted 70S ribosome-binding domain. The mutation results in a delay in the onset of germination. At this early developmental stage embryos still contain undifferentiated proplastids, whose proper function seems necessary for seed germination. In light-grown sco1 seedlings the greening of cotyledons is severely impaired, whereas the following true leaves develop normally as in wild-type plants. Despite this apparent similarity of chloroplast development in true leaves of mutant and wild-type plants various aspects of mature plant development are also affected by the sco1 mutation such as the onset of flowering, the growth rate, and seed production. The onset of senescence in the mutant and the wild-type plants occurs, however, at the same time, suggesting that in the mutant this particular developmental step does not seem to suffer from reduced protein translation efficiency in chloroplasts.

Calcium ions represent both an integrative signal and an important convergence point of many disparate signaling pathways. Calcium-binding proteins, like calcineurin B-like (CBL) proteins, have been implicated as important relays in calcium signaling. Here, we report the in vivo study of CBL1 function in Arabidopsis. Analyses of loss-of-function as well as CBL1-overexpressing lines indicate a crucial function of this calcium sensor protein in abiotic stress responses. Mutation of CBL1 impairs plant responses to drought and salt stresses and affects gene expression of cold-regulated genes, but does not affect abscisic acid (ABA) responsiveness. Conversely, overexpression of CBL1 reduces transpirational water loss and induces the expression of early stress-responsive transcription factors and stress adaptation genes in non-stressed plants. Together, our data indicate that the calcium sensor protein CBL1 may constitute an integrative node in plant responses to abiotic stimuli and contributes to the regulation of early stress-related transcription factors of the C-Repeat-Binding Factor/dehydration-responsive element (CBF/DREB) type.

Members of the Arabidopsis calcineurin B-like Ca2+ binding protein (AtCBL) family are differentially regulated by stress conditions. One AtCBL plays a role in salt stress; another is implicated in response to other stress signals, including drought, cold, and wounding. In this study, we identified a group of novel protein kinases specifically associated with AtCBL-type Ca2+ sensors. In addition to a typical protein kinase domain, they all contain a unique C-terminal region that is both required and sufficient for interaction with the AtCBL-type but not calmodulin-type Ca2+ binding proteins from plants. Interactions between the kinases and AtCBLs require micromolar concentrations of Ca2+, suggesting that increases in cellular Ca2+ concentrations may trigger the formation of AtCBL-kinase complexes in vivo. Unlike most serine/threonine kinases, the AtCBL-interacting kinase efficiently uses Mn2+ to Mg2+ as a cofactor and may function as a Mn2+ binding protein in the cell. These findings link a new type of Ca2+ sensors to a group of novel protein kinases, providing the molecular basis for a unique Ca2+ signaling machinery in plant cells.

Summary Intracellular release of calcium ions belongs to the earliest events in cellular stress perception. The molecular mechanisms integrating signals from different environmental cues and translating them into an optimized response are largely unknown. We report here the functional characterization of CIPK1, a protein kinase interacting strongly with the calcium sensors CBL1 and CBL9. Comparison of the expression patterns indicates that the three proteins execute their functions in the same tissues. Physical interaction of CIPK1 with CBL1 and CBL9 targets the kinase to the plasma membrane. We show that, similarly to loss of CBL9 function, mutation of either CBL1 or CIPK1 renders plants hypersensitive to osmotic stress. Remarkably, in contrast to the cbl1 mutant and similarly to the cbl9 mutant, loss of CIPK1 function impairs abscisic acid (ABA) responsiveness. We therefore suggest that, by alternative complex formation with either CBL1 or CBL9, the kinase CIPK1 represents a convergence point for ABA-dependent and ABA-independent stress responses. Based on our genetic, physiological and protein-protein interaction data, we propose a general model for information processing in calcium-regulated signalling networks.