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In the present study, leaves of different plant species were girdled by the hot wax collar method to prevent export of assimilates. Photosynthetic activity of girdled and control leaves was evaluated 3 to 7 days later by two methods: (a) carbon exchange rate (CER) of attached leaves was determined under ambient CO2 concentrations using a closed gas system, and (b) maximum photosynthetic capacity (Amax) probably not directly responsible for the end-product inhibition of photosynthesis. The occurrence of strong end-product inhibition appears to be correlated with high acid-invertase activity in fully expanded leaves. The inhibition may be related to the nature of soluble sugar metabolism in the extrachloroplastic compartment and may be caused by a metabolite that has different rates of accumulation and turnover in sucrose storers and other plants.

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Grafting, an old plant propagation practice, is still widely used with fruit trees and in recent decades also with vegetables. Taxonomic proximity is a general prerequisite for successful graft-take and long-term survival of the grafted, composite plant. However, the mechanisms underlying interspecific graft incompatibility are as yet insufficiently understood. Hormonal signals, auxin in particular, are believed to play an important role in the wound healing and vascular regeneration within the graft union zone. Incomplete and convoluted vascular connections impede the vital upward and downward whole plant transfer routes. Long-distance protein, mRNA and small RNA graft-transmissible signals currently emerge as novel mechanisms which regulate nutritional and developmental root/top relations and may play a pivotal role in grafting physiology. Grafting also has significant pathogenic projections. On one hand, stock to scion mechanical contact enables the spread of diseases, even without a complete graft union. But, on the other hand, grafting onto resistant rootstocks serves as a principal tool in the management of fruit tree plagues and vegetable soil-borne diseases. The ‘graft hybrid’ historic controversy has not yet been resolved. Recent evidence suggests that epigenetic modification of DNA-methylation patterns may account for certain graft-transformation phenomena. Root grafting is a wide spread natural phenomenon; both intraspecific and interspecific root grafts have been recorded. Root grafts have an evolutionary role in the survival of storm-hit forest stands as well as in the spread of devastating diseases. A more fundamental evolutionary role is hinted by recent findings that demonstrate plastid and nuclear genome transfer between distinct Nicotiana species in the graft union zone, within a tissue culture system. This has led to the formation of alloploid cells that, under laboratory conditions, gave rise to a novel, alloploid Nicotiana species, indicating that natural grafts may play a role in plant speciation, under certain circumstances.

Summary We report on the isolation, functional expression and characterization of a cDNA encoding chlorophyllase, the enzyme catalyzing the first step in the chlorophyll breakdown pathway. The Chlase1 cDNA from Valencia Orange (Citrus sinensis cv. Valencia) was obtained by RT–PCR using degenerate primers based on the amino acid sequence of the previously purified protein. Chlase1 encodes a protein of 329 amino acids, including a sequence domain characterizing serine‐lipases and a putative chloroplast‐directing transit peptide. The Chlase1 gene encodes an active chlorophyllase enzyme which catalyzes the dephytylation of chlorophyll as shown by in vitro recombinant enzyme assays. Chlorophyllase expression at the transcript level in Valencia orange peel was found to be low and constitutive during natural fruit development without significant increase towards color‐break and ripening. However, ethylene treatment induced an increase in chlorophyllase transcript at all stages of development. An enhanced response to ethylene treatment was observed during the months of October and November, corresponding to the time of natural color‐break. The senescence‐delaying regulator gibberellin‐A3 (GA3) inhibited the effect of ethylene on chlorophyllase transcript accumulation. The data presented suggest that chlorophyllase may not be the regulator of chlorophyll breakdown during natural fruit ripening but is consistent with the notion that chlorophyll is gradually degraded during ripening due to a negative balance between synthesis and breakdown. According to this model, exogenous application of ethylene accelerates chlorophyll breakdown due to increased de novo synthesis of chlorophyllase. Further experi‐ mentation on the regulation and role of chlorophyllase in planta will be facilitated by the gene tools established in this work.

Biology of Citrus provides a concise and comprehensive discussion of all major developmental, genetic and horticultural aspects of citriculture in an easily readable text. The book deals with the history, distribution and climatic adaptation of the crop, followed by taxonomy and systematics, including a horticultural classification of edible citrus species. Subsequent chapters cover tree structure and function, reproductive physiology, including flowering, fruiting, productivity, ripening, post-harvest and fruit constituents. The main aspects of cultivated citrus, such as rootstocks, irrigation, pests, viruses and diseases are dealt with, leading to a concluding chapter that considers genetic improvement, including the use of tissue culture and plant biotechnology. The book includes many specially produced original illustrations and the extensive reading lists will make it invaluable for students and citrus specialists.

Chlorophyll is a central player in harvesting light energy for photosynthesis, yet the rate-limiting steps of chlorophyll catabolism and the regulation of the catabolic enzymes remain unresolved. To study the role and regulation of chlorophyllase (Chlase), the first enzyme of the chlorophyll catabolic pathway, we expressed precursor and mature versions of citrus (Citrus sinensis) Chlase in two heterologous plant systems: (1) squash (Cucurbita pepo) plants using a viral vector expression system; and (2) transiently transformed tobacco (Nicotiana tabacum) protoplasts. Expression of full-length citrus Chlase resulted in limited chlorophyll breakdown in protoplasts and no visible leaf phenotype in whole plants, whereas expression of a Chlase version lacking the N-terminal 21 amino acids (ChlaseDN), which corresponds to the mature protein, led to extensive chlorophyll breakdown in both tobacco protoplasts and squash leaves. ChlaseDN-expressing squash leaves displayed a dramatic chlorotic phenotype in plants grown under low-intensity light, whereas under natural light a lesion-mimic phenotype occurred, which was demonstrated to follow the accumulation of chlorophyllide, a photodynamic chlorophyll breakdown product. Fulllength and mature citrus Chlase versions were localized to the chloroplast membrane fraction in expressing tobacco protoplasts, where processing of the N-terminal 21 amino acids appears to occur. Results obtained in both plant systems suggest that Chlase functions as a rate-limiting enzyme in chlorophyll catabolism controlled via posttranslational regulation.

One important reaction of chlorophyll (chl) breakdown during plant senescence is the removal of the lipophilic phytol moiety by chlorophyllase. AtCLH1 and AtCLH2 were considered to be required for this reaction in Arabidopsis thaliana. Here we present evidence against this assumption. Using green fluorescent protein fusions, neither AtCLH isoform localizes to chloroplasts, the predicted site of chlorophyll breakdown. Furthermore, clh1 and clh2 single and double knockout lines are still able to degrade chlorophyll during senescence. From our data we conclude that AtCLHs are not required for senescence-related chlorophyll breakdown in vivo and propose that genuine chlorophyllase has not yet been molecularly identified.