A carrot gene homologous to the ABI3 gene of Arabidopsis was isolated from a carrot somatic embryo cDNA library and designated C-ABI3. The sequence of C-ABI3 was very similar to those of ABI3 of Arabidopsis and VP1 of maize in certain conserved regions. The expression of C-ABI3 was detected specifically in embryogenic cells, somatic embryos and developing seeds. Thus, expression of C-ABI3 was limited to tissues that acquired desiccation tolerance in response to endogenous or exogenous abscisic acid (ABA). Endogenous levels of ABA in seeds increased transiently and then desiccation of seeds started. The expression of C-ABI3 in developing seeds was observed prior to the increase in levels of endogenous ABA that was followed by desiccation of seeds. In transgenic mature leaves in which C-ABI3 was ectopically expressed, expression of ECP31, ECP63 and ECP40 was induced by treatment with ABA, which indicates that the expression of ECP genes was controlled by the pathway(s) that involved C-ABI3 and ABA. This suggests that C-ABI3 has the same function as VP1/ABI3 factor in carrot somatic embryos.

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Three cDNA clones encoding isoforms of carrot glutamine synthetase (GS) were isolated and used as probes for analysis of the patterns of expression of the genes for GS isoforms during somatic embryogenesis and seed development in carrot. Transcripts corresponding to two of the cDNAs, CGS102 and CGS201, accumulated in both somatic embryos and developing seeds in the same manner. Their levels were high at the early stage of embryogenesis but decreased at the late stage. This pattern of expression is similar to the pattern of changes in GS activity observed during somatic embryogenesis. In contrast, expression of the transcript for another GS isoform detected with CGS103 cDNA was observed at the late stage of seed development and in senesced leaves but not in somatic embryos or young leaves. We also analyzed the levels of the transcripts in somatic embryos that had been cultured in media with either ammonium ions or glutamine as the nitrogen source. The amounts of the CGS102 and CGS201 transcripts fell when glutamine was supplied in the medium. These results indicated that GS activity was regulated at the transcriptional level and that the pattern of expression of the genes for GS during somatic embryogenesis reflected that during zygotic embryogenesis. It is possible that somatic embryogenesis and zygotic embryogenesis have common regulatory systems with respect to nitrogen metabolism.

A novel pollen-specific LEA-like protein, LP28, was detected in Lilium longiflorum using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). Immunoblot analysis using antiserum raised against LP28 revealed that the protein was not found in somatic tissues or uninucleate microspores, but accumulated gradually in developing pollen following microspore mitosis. Furthermore, LP28 was abundant in germinated pollen after hydration. The cDNA clone corresponding to LP28 encoded a putative protein of 238 amino acids with a calculated molecular mass of 24·2 kDa and a pI of 4·7. The amino acid sequence is highly hydrophilic except for the N-terminal hydrophobic signal peptide. The sequence has similarities with group 3 LEA (late embryogenesis abundant) proteins. Immunocytochemical analyses demonstrated that LP28 was mainly found in cytoplasmic granules of the vegetative cell until pollen maturation, but after hydration it appeared in the elongating pollen tube wall. LP28 might be a unique pollenspecific protein that is transported to the pollen tube wall after germination. Therefore, it is assumed that LP28 plays a role not only in pollen maturation, but also in the growth of the pollen tube, which penetrates the stylar matrix.

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Many plant species exhibit diurnal flower opening and closing, which is an adaptation influenced by the lifestyle of pollinators and herbivores. However, it remains unclear how these temporal floral movements are modulated. To clarify the role of the circadian clock in flower movement, we examined temporal floral movements in Arabidopsis thaliana. Wild-type (accessions; Col-0, Ler-0, and Ws-4) flowers opened between 0.7 to 1.4 h in a 16 h light period and closed between 7.5 to 8.3 h in a diurnal light period. In the arrhythmic mutants pcl1-1 and prr975, the former flowers closed slowly and imperfectly, and the latter ones never closed. Under continuous light conditions, new flowers emerged and opened within the 23–26 h window in the wild-type, but the flowers in pcl1-1 and prr975 developed straight petals, whose curvatures were severely small. Anti-phasic circadian gene expression of CIRCADIAN CLOCK ASSOCIATED 1 (CCA1), LATE ELONGATED HYPOCOTYLE 1 (LHY1) and TIMING OF CAB EXPRESSION 1 (TOC1) occurred in wild-type flowers, but non-rhythmic expression was observed in pcl1-1 and prr975 mutants. Focusing on excised petals, bioluminescence monitoring revealed rhythmic promoter activities of genes expressed (CCA1 and PHYTOCLOCK 1/LUX ARRHYTHMO, PCL1/LUX) in the morning and evening, respectively. These results suggest that the clock induces flower opening redundantly with unknown light-sensing pathways. In contrast, flower closing is completely dependent on clock control. These findings will lead to further exploration of the molecular mechanisms and evolutionary diversity of timing in flower opening and closing.