SummaryDisproportionating enzyme (D-enzyme) is a plastidial a-1,4-glucanotransferase but its role in starch metabolism is unclear. Using a reverse genetics approach we have isolated a mutant of Arabidopsis thaliana in which the gene encoding this enzyme (DPE1) is disrupted by a T-DNA insertion. While Denzyme activity is eliminated in the homozygous dpe1±1 mutant, changes in activities of other enzymes of starch metabolism are relatively small. During the diurnal cycle, the amount of leaf starch is higher in dpe1±1 than in wild type and the amylose to amylopectin ratio is increased, but amylopectin structure is unaltered. The amounts of starch synthesised and degraded are lower in dpe1±1 than in wild type. However, the lower amount of starch synthesised and the higher proportion of amylose are both eliminated when plants are completely de-starched by a period of prolonged darkness prior to the light period. During starch degradation, a large accumulation of malto-oligosaccharides occurs in dpe1±1 but not in wild type. These data show that D-enzyme is required for malto-oligosaccharide metabolism during starch degradation. The slower rate of starch degradation in dpe1±1 suggests that maltooligosaccharides affect an enzyme that attacks the starch granule, or that D-enzyme itself can act directly on starch. The effects on starch synthesis and composition in dpe1±1 under normal diurnal conditions are probably a consequence of metabolism at the start of the light period, of the high levels of maltooligosaccharides generated during the dark period. We conclude that the primary function of D-enzyme is in starch degradation.

SummaryHeteroblasty refers to the changes in leaf shape and size (allometry) along stems. Although evolutionary changes involving heteroblasty might contribute to leaf diversity, little is known of the extent to which heteroblasty differs between species or how it might relate to other aspects of allometry or other developmental transitions.Here, we develop a computational model that can quantify differences in leaf allometry between Antirrhinum (snapdragon) species, including variation in heteroblasty. It allows the underlying genes to be mapped in inter-species hybrids, and their effects to be studied in similar genetic backgrounds.Heteroblasty correlates with overall variation in leaf allometry, so species with smaller, rounder leaves produce their largest leaves earlier in development. This involves genes that affect both characters together and is exaggerated by additional genes with multiplicative effects on leaf size. A further heteroblasty gene also alters leaf spacing, but none affect other developmental transitions, including flowering.We suggest that differences in heteroblasty have co-evolved with overall leaf shape and size in Antirrhinum because these characters are constrained by common underlying genes. By contrast, heteroblasty is not correlated with other developmental transitions, with the exception of internode length, suggesting independent genetic control and evolution.

Transgenic potato (Solanum tuberosum L.) plants were created with sense and antisense copies of the potato D-enzyme (disproportionating enzyme; EC 2.4.1.25) cDNA linked to patatin and cauli¯ower mosaic virus 35 S promoters, and screened for Denzyme activity in tubers. Transformants with sense constructs mostly had wild type D-enzyme activity but two plants had only about 1% wild-type activity. Transformants with antisense constructs had activity ranging from 90% to about 1% of wild type. Three 35 S antisense plants with very low activity were analysed in detail. Western blot analysis showed that D-enzyme was present in greatly reduced amounts in tubers and in leaves, whereas plastidic starch phosphorylase (EC 2.4.1.1) was unaected. The lack of D-enzyme resulted in slow plant growth but development was otherwise apparently normal. Furthermore, the starch content of tubers was not appreciably altered in amount, proportion of amylose, molecular weight of debranched amylopectin, or branch chain length, despite the lack of D-enzyme. These results do not indicate a direct requirement for D-enzyme in the synthesis and accumulation of storage starch in tubers. The results are discussed in terms of the known reactions catalysed by D-enzyme and possible involvement of D-enzyme in starch metabolism.

INTRODUCTIONThe Antirrhinum species group comprises approximately 20 morphologically diverse members that are able to form fertile hybrids. It includes the cultivated snapdragon Antirrhinum majus, which has been used as a model for biochemical and developmental genetics for more than 75 yr. The research infrastructure for A. majus, together with the interfertility of the species group, allows Antirrhinum to be used to examine the genetic basis for plant diversity.

INTRODUCTIONThis protocol describes general strategies for propagating Antirrhinum (snapdragon) species: self- and cross-pollination, cuttings, and grafting. Antirrhinum majus cultivars and some wild species are self-fertile, but they require self-pollination for high seed yields. Although self-fertile, A. majus shows unilateral incompatibility and can only be crossed to other self-incompatible species as the female parent. All Antirrhinum species can be propagated clonally from cuttings. Antirrhinum also readily forms grafts within and between species.