In mammals, cadmium is widely considered as a non-genotoxic carcinogen acting through a methylation-dependent epigenetic mechanism. Here, the effects of Cd treatment on the DNA methylation patten are examined together with its effect on chromatin reconfiguration in Posidonia oceanica. DNA methylation level and pattern were analysed in actively growing organs, under short- (6 h) and long- (2 d or 4 d) term and low (10 mM) and high (50 mM) doses of Cd, through a Methylation-Sensitive Amplification Polymorphism technique and an immunocytological approach, respectively. The expression of one member of the CHROMOMETHYLASE (CMT) family, a DNA methyltransferase, was also assessed by qRT-PCR. Nuclear chromatin ultrastructure was investigated by transmission electron microscopy. Cd treatment induced a DNA hypermethylation, as well as an up-regulation of CMT, indicating that de novo methylation did indeed occur. Moreover, a high dose of Cd led to a progressive heterochromatinization of interphase nuclei and apoptotic figures were also observed after long-term treatment. The data demonstrate that Cd perturbs the DNA methylation status through the involvement of a specific methyltransferase. Such changes are linked to nuclear chromatin reconfiguration likely to establish a new balance of expressed/repressed chromatin. Overall, the data show an epigenetic basis to the mechanism underlying Cd toxicity in plants.

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In Posidonia oceanica cadmium induces changes in DNA methylation and chromatin patterning

Alterations in plant development are frequently observed following pathogen infection. Infection by virus frequently results in stunting of growth, and the chlorosis and abscission of leaves; infection by fungi is often notable for green island formation and growth malformations; and infection by some bacteria results in the formation of galls. While the area of plant-pathogen interactions is currently receiving considerable attention and some plant-pathogen interactions are well characterised with both cytokinins and auxins being implicated (infection by Agrobacterium tumefaciens being the obvious example), there has been relatively little published in the recent literature pertaining to the involvement of cytokinins and auxins in viral, fungal and other forms of bacterial pathogenesis. This overview focuses on what is known concerning the strategies utilised by gall-forming bacteria, and fungal and viral phytopathogens to manipulate the endogenous cytokinin and/or auxin content of their host plant.

Cytokinins and auxins in plant-pathogen interactions – An overview

The cytokinins have been implicated in many facets of plant growth and development including cell division and differentiation, shoot and root growth, apical dominance, senescence, fruit and seed development, and the response to biotic and abiotic stressors. Cytokinin levels are regulated by a balance between biosynthesis [isopentenyl transferase (IPT)], activation [Lonely Guy (LOG)], inactivation (O-glucosyl transferase), re-activation (β-glucosidase), and degradation [cytokinin oxidase/dehydrogenase (CKX)]. During senescence, the levels of active cytokinins decrease, with premature senescence leading to a decrease in yield. During the early stages of fruit and seed development, cytokinin levels are transiently elevated, and coincide with nuclear and cell divisions which are a determinant of final seed size. Exogenous application of cytokinin, ectopic expression of IPT, or down-regulation of CKX have, on occasions, led to increased seed yield, leading to the suggestion that cytokinin may be limiting yield. However, manipulation of cytokinins is complex, not only because of their pleiotropic nature but also because the genes coding for biosynthesis and metabolism belong to multigene families, the members of which are themselves spatially and temporally differentiated. Previous research on yield of rice showed that plant breeders could directly target the cytokinins. Modern genome editing tools could be employed to target and manipulate cytokinin levels to increase seed yield with the concurrent aim of maintaining quality. However, how the cytokinin level is modified and whether IPT or CKX is targeted may depend on whether the plant is considered to be in a source-limiting environment or to be sink limited.

Cytokinin: a key driver of seed yield

Cytokinin oxidase (CK oxidase) is widely distributed in plants and is the only enzyme that has been shown unequivocally to catalyze the catabolism of specific cytokinins (CKs) to inactive products that lack the N6-unsaturated side chain Thus, the enzyme is thought to play a major role in controlling the level or species of CKs in plant tissues However, despite its discovery more than 25 years ago, little attention has been given to the elucidation of its role and function in plant growth and development This review seeks to bring in to context the current state of knowledge regarding the biochemical and molecular properties, regulation in undifferentiated and differentiated tissues, and recent results from studies using transgenic plants in an attempt to provide a more comprehensive understanding of the physiological significance of the enzyme in plants Notwithstanding species, tissue and other specific differences, in general, CK oxidase appears to contribute to CK homeostasis in plants However, complete clarity as to its function awaits purification of the protein to homogeneity and the ultimate development of requisite molecular probes

Role and function of cytokinin oxidase in plants

CPPU application on size and quality of hardy kiwifruit

The significance of changes in cytokinin content during early fruit growth was examined in the kiwifruit (Actinidia deliciosa var. deliciosa cv. Hayward). Fruit growth was modified by the reduction of seed number or by the application of the synthetic phenylurea cytokinin N-(2-chloro-4-pyridyl)-N'-phenylurea (CPPU). The influence of these treatments on cell division was monitored by flow cytometry and changes in the endogenous cytokinins were measured at days 10 and 20 after anthesis, using high-performance liquid chromatography and radioimmunoassay. Total cytokinin levels appeared not to be limiting growth since the highest total cytokinin concentration was detected in unpollinated fruit, which abscised by day 25 after anthesis. However, compared with control fruit which had the highest concentration of zeatin (Z) 10 days post anthesis, Z levels were low in unpollinated fruit. It is hypothesised that an increase in Z is the critical change in cytokinin metabolism required for the initiation of cell division and fruit growth. The synthetic cytokinin CPPU promoted fruit development, but there was a decrease in the endogenous cytokinin concentration. Zeatin was not detected in CPPU-treated fruit. Cell division was reduced in unpollinated fruitlets but there was no significant difference (P > 0.05) between the other treatments. Differences in final fruit size appeared to be due to cell expansion.

Murray E. Hopping

Papers

Cytokinins and fruit development in the kiwifruit (Actinidia deliciosa). II. Effects of reduced pollination and CPPU application