Expression of candidate marker genes of sugar starvation is upregulated in growth-suppressed parthenocarpic cucumber fruit. Novel gene markers for sugar starvation in growth-suppressed cucumber fruit.

To examine the physiological change in the growth suppression and abortion of parthenocarpic cucumber fruit, the expression of candidate marker genes of sugar starvation in relation to growth activity was examined. Fruits that failed to start exponential growth seemed to eventually abort. Hexose concentration of fruits was low in growth-suppressed fruit and increased in normally growing fruit consistent with the vacuolization. The correlation matrix indicated that the transcript levels of the genes, except CsaV3_6G046050 and CsaV3_5G032930, had a highly significant negative correlation with the relative growth rate in fruit length and had highly significant mutual positive correlations, suggesting that the asparagine synthetase gene, Cucumis sativus putative CCCH-type zinc finger protein CsSEF1, C. sativus BTB/POZ domain-containing protein At1g63850-like, CsaV3_3G000800, CsaV3_3G041280, and CsaV3_7G032930 are good markers of sugar starvation in cucumber fruit. The expression of candidate marker genes together with the hexose analysis strongly suggests that severe sugar starvation is occurring in growth-suppressed fruit.

The expression of a candidate cucumber fruit sugar starvation marker gene CsSEF1 is enhanced in malformed fruit induced by salinity.

The cucumber (Cucumis sativus L.) gene Cucumis sativus Somatic Embryogenesis Zinc Finger 1 (CsSEF1) was suggested to be a good marker gene for sugar starvation in fruit. The expression of this gene in fruits is dramatically upregulated in plants that have suffered either complete defoliation or prolonged darkness. CsSEF1 was initially discovered as a gene that was upregulated during somatic embryogenesis. We examined the difference in fruit parts and the effect of pollination on the upregulation of CsSEF1 induced by defoliation treatment. The results indicated that the upregulation of CsSEF1 in fruit by defoliation is not dependent on the presence of developing embryos. The expression of CsSEF1 was upregulated in malformed fruit induced by salinity in which the development of placenta was arrested. Partial cutting of the distal part of the fruit showed that if placenta tissue remained there was no upregulation of CsSEF1, whereas when placenta tissue did not remain there was a marked upregulation of CsSEF1. These results could be consistently interpreted as showing that placenta tissue induced the transport of photoassimilates to the fruit and that without developing placenta tissue, pericarp tissue suffers from severe sugar starvation. This interpretation, in turn, enforces the view that CsSEF1 is a good marker gene of fruit sugar starvation.

Genetic engineering and metabolite profiling for overproduction of polyhydroxybutyrate in cyanobacteria.

Genetic engineering and metabolite profiling for the overproduction of polyhydroxybutyrate (PHB), which is a carbon material in biodegradable plastics, were examined in the unicellular cyanobacterium Synechocystis sp. PCC 6803. Transconjugants harboring cyanobacterial expression vectors that carried the pha genes for PHB biosynthesis were constructed. The overproduction of PHB by the engineering cells was confirmed through microscopic observations using Nile red, transmission electron microscopy (TEM), or nuclear magnetic resonance (NMR). We successfully recovered PHB from transconjugants prepared from nitrogen-depleted medium without sugar supplementation in which PHB reached approximately 7% (w/w) of the dry cell weight, showing a value of 12-fold higher productivity in the transconjugant than that in the control strain. We also measured the intracellular levels of acetyl-CoA, acetoacetyl-CoA, and 3-hydroxybutyryl-CoA (3HB-CoA), which are intermediate products for PHB. The results obtained indicated that these products were absent or at markedly low levels when cells were subjected to the steady-state growth phase of cultivation under nitrogen depletion for the overproduction of bioplastics. Based on these results, efficient factors were discussed for the overproduction of PHB in recombinant cyanobacteria.

Expression of CsSEF1 gene encoding putative CCCH zinc finger protein is induced by defoliation and prolonged darkness in cucumber fruit.

To find a marker gene for photoassimilate limitation in cucumber fruit, genes induced in young fruit by total defoliation were cloned using the subtraction method. Almost every clone matched perfectly to a member of cucumber unigene ver. 3 of the Cucurbit Genomics Database. From the clones obtained, six genes were selected and the effect of defoliation on their expression was analyzed. In particular, expression of a gene that is highly homologous to the cucumber gene CsSEF1 (CAI30889) encoding putative CCCH zinc finger protein, which is reported to be induced at somatic embryogenesis in suspension culture, was enhanced by the treatment by about 50 times. The sequencing of the full-length cDNA and BLAST search in the Cucurbit Genomics Database indicated that our cloned gene is identical to CsSEF1. In control fruit, the expression of CsSEF1 did not change markedly in terms of development. By contrast, the expression of CsSEF1 was enhanced by prolonged darkness at the transcript level. This increase in the expression of CsSEF1 was temporally correlated with the decline in the fruit respiration rate. In mature leaves under prolonged darkness, enhanced expression was observed in the asparagine synthetase gene, but not in CsSEF1. These results suggest that the asparagine synthetase gene can be a good marker for sugar starvation and that CsSEF1 might be involved in the signal transduction pathway from photoassimilate limitation to growth cessation in cucumber fruit.