A complementary DNA encoding a salicylic acid (SA)-binding protein has been cloned. Its properties suggest involvement in SA-mediated induction of systemic acquired resistance (SAR) in plants. The sequence of the protein is similar to that of catalases and the protein exhibits catalase activity. Salicylic acid specifically inhibited the catalase activity in vitro and induced an increase in H2O2 concentrations in vivo. H2O2 or compounds, such as SA, that inhibit catalases or enhance the generation of H2O2, induced expression of defense-related genes associated with SAR. Thus, the action of SA in SAR is likely mediated by elevated amounts of H2O2.

https://science.sciencemag.org/content/sci/262/5141/1883.full.pdf

Active oxygen species in the induction of plant systemic acquired resistance by salicylic acid

■ Abstract Cytokinins are structurally diverse and biologically versatile. The chemistry and physiology of cytokinin have been studied extensively, but the regulation of cytokinin biosynthesis, metabolism, and signal transduction is still largely undefined. Recent advances in cloning metabolic genes and identifying putative receptors portend more rapid progress based on molecular techniques. This review centers on cytokinin metabolism with connecting discussions on biosynthesis and signal transduction. Important findings are summarized with emphasis on metabolic enzymes and genes. Based on the information generated to date, implications and future research directions are presented.

Cytokinin metabolism and action

After the discovery of kinetin (Miller et al. 1956, J. Am. Chem. Soc. 78: 1345–1350) there was a flurry of syntheses that led to the finding of 6-benzylaminopurine (BA), an active and easily obtainable cytokinin. Much research into cytokinin physiology was subsequently done with this substance. Further, the isolation and unequivocal identification of natural BA and the high biological activity of its meta-hydroxylated analogues stimulated the search for other natural aromatic cytokinins. Screening was accomplished by ELISA of HPLC fractions using antisera against ortho- and meta-hydroxybenzyladenosine. Subsequent isolation and decisive identification by mass spectrometry led to discovery of a broad spectrum of endogenous plant growth substances structurally similar to a highly active compound, meta-topolin (6-[3-hydroxybenzyl-amino]purine), and to its less active analogue, ortho-topolin (6-[2-hydroxybenzyl-amino]purine). The structures of such aromatic cytokinins suggest considerably different biosynthetic pathways from that of zeatin and related isoprenoid cytokinins. From a physiological viewpoint, aromatic cytokinin metabolism can be classified under four main headings analogous to isoprenoid cytokinins: interconversion, hydroxylation, conjugation, and oxidative degradation. This review attempts to put into context what is known about 9-alkyl-BAs and compares their metabolism in regard to the practical use of cytokinins in agriculture and biotechnology. The recently discovered unusual specificity of additionally C2,N9-disubstituted aromatic cytokinins toward cell cycle kinases, suggests that these cytokinin-derived growth regulators may selectively inhibit certain steps of the cell cycle. The functional overlap of the aromatic cytokinins with those of their isoprenoid counterparts and cytokinin inhibitors, in relation to growth and developmental processes in plants, has yet to be determined.

The aromatic cytokinins

Arabidopsis fusca mutants display striking purple coloration due to anthocyanin accumulation in their cotyledons. We describe six recessive fusca mutants isolated from Agrobacterium-transformed Arabidopsis families. These mutants first become defective during embryogenesis and exhibit limited seedling development. Double mutant constructs revealed that developmental defects were not simply a consequence of anthocyanin accumulation. fusca seedlings showed altered responses to several environmental and endogenous factors. Allelism tests established that three fusca loci are represented by mutants previously described as defective in light-regulated responses. To study the molecular basis of the fusca phenotype, we cloned the FUS6 gene. FUS6 encodes a novel protein that is hydrophilic, alpha-helical, and contains potential protein kinase C phosphorylation sites. The FUSCA proteins appear to act in a network of signal transduction pathways critical for plant development.

/pdf/a-fusca-gene-of-arabidopsis-encodes-a-novel-protein-50i7p8gw7c.pdf

A FUSCA gene of Arabidopsis encodes a novel protein essential for plant development.

We have genetically modified maize plants to delay leaf senescence. A senescence-enhanced promoter from maize (P(SEE1)) was used to drive expression of the Agrobacterium cytokinin biosynthesis gene IPT in senescing leaf tissue. Three maize lines expressing IPT from P(SEE1), Sg1, Sg2 and Sg3, were analysed in detail, representing mild, intermediate and extreme expression, respectively, of the delayed-senescence phenotype. Backcross populations segregating for the presence or absence of the P(SEE1Xba)IPTNOS transgene also simultaneously segregated for the senescence phenotype. At the time of ear leaf emergence, individuals of lines Sg1 and Sg2 segregating for the presence of the transgene carried about three fewer senescing leaves than control (transgene-minus) segregants, and IPT transcript levels were higher in leaves at incipient senescence than in young leaves. Leaves of transgenic Sg3 plants were significantly greener than controls and progressed directly from fully green to bleached and dead without an intervening yellowing phase. IPT transcript abundance in this line was not related to the initiation of senescence. Extended greenness was accompanied by a delay in the loss of photosynthetic capacity with leaf age. The delayed-senescence trait was associated with relatively minor changes in morphology and development. The phenotype was particularly emphasized in plants grown in low soil nitrogen. The reduced ability of the extreme transgenic line Sg3 to recycle internal nitrogen from senescing lower leaves accounted for significant chlorosis in emerging younger leaves when plants were grown in low nutrient conditions. This study demonstrates that the agronomically important delayed-senescence ('stay-green') trait can be engineered into a monocot crop, and is the first example outside Arabidopsis of senescence modification using a homologous senescence-enhanced promoter.

Leaf senescence is delayed in maize expressing the Agrobacterium IPT gene under the control of a novel maize senescence-enhanced promoter.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/0014-5793%2890%2980604-H

Isolation of a protein with cytokinin-receptor properties by means of anti-idiotype antibodies

Cytokinin signalling systems : From a whole plant to the molecular level

A role of endogenous cytokinins as the hormonal signal, by which the roots regulate leaf metabolism and prevent leaf senescence in particular, is discussed. Cytokinin signal perception and transduction in leaf cells were studied on fully expanded first leaves of 10–13-day-old barley plants (Hordeum vulgare L. cv. Viner). Their high sensitivity to exogenous cytokinins depends on dramatic decrease in endogenous cytokinin content during leaf growth. Cytokinin-binding protein of 28–30 kD was isolated from barley leaf cytosol by its affinity to synthetic cytokinin benzyladenine (BA) The 30-kD protein was involved in cytokinin-dependent activation of RNA synthesis in vitro in the system containing chromatin-bound RNA polymerase I from barley leaves. Multistage purification of the protein included affinity chromatography on trans-zeatin-Sepharose or zeatin riboside-Sepharose resulted in isolation of barley leaf cytosol 67-kD protein up to electrophoretic homogeneity. The protein was revealed as a single band in Western blot analysis developed with anti-idiotype antibodies from antiserum to zeatin. In concert with trans-zeatin the 67-kD protein activated transcription elongation directed by RNA polymerase I (in the system containing chromatin-bound RNA polymerase I from barley leaves) and by RNA polymerase II (in nuclei isolated from barley leaves). The protein effect strongly depended on cytokinin concentration. The maximum activation was observed at trans-zeatin concentration of 10-8 M. cis-Zeatin had no effect. These results together with data on reversible [3H]zeatin-binding moiety of the 67-kD protein [8] provide a definitive proof to consider this protein as one of the cytokinin receptors in barley leaf cells which is responsible for cytokinin activation of transcription elongation directed by both RNA polymerase I and RNA polymerase II.

I. E. Moshkov

Papers

Isolation of a protein with cytokinin-receptor properties by means of anti-idiotype antibodies

Cytokinin signalling systems : From a whole plant to the molecular level

Cytokinin signalling systems

Occurrence of cytokinin-controlled C-type protein kinase in barley leaves.