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Ripening-associated ethylene biosynthesis in tomato fruit is autocatalytically and developmentally regulated.

Yokotani N, Nakano R, Imanishi S, Nagata M, Inaba A, Kubo Y - J. Exp. Bot. (2009)

Bottom Line: These results suggest that ripening-associated ethylene (system 2) in wild-type tomato fruit consists of two parts: a small part regulated by a developmental factor through the ethylene-independent expression of LeACS2 and LeACS4 and a large part regulated by an autocatalytic system due to the ethylene-dependent expression of the same genes.The results further suggest that basal ethylene (system 1) is less likely to be involved in the transition to system 2.Even if the effect of system 1 ethylene is eliminated, fruit can show a small increase in ethylene production due to unknown developmental factors.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Biological Sciences, 7549-1 Yoshikawa, Kibichuo-cho, Okayama, 716-1241 Japan.

ABSTRACT
To investigate the regulatory mechanism(s) of ethylene biosynthesis in fruit, transgenic tomatoes with all known LeEIL genes suppressed were produced by RNA interference engineering. The transgenic tomato exhibited ethylene insensitivity phenotypes such as non-ripening and the lack of the triple response and petiole epinasty of seedlings even in the presence of exogenous ethylene. Transgenic fruit exhibited a low but consistent increase in ethylene production beyond 40 days after anthesis (DAA), with limited LeACS2 and LeACS4 expression. 1-Methylcyclopropene (1-MCP), a potent inhibitor of ethylene perception, failed to inhibit the limited increase in ethylene production and expression of the two 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS) genes in the transgenic fruit. These results suggest that ripening-associated ethylene (system 2) in wild-type tomato fruit consists of two parts: a small part regulated by a developmental factor through the ethylene-independent expression of LeACS2 and LeACS4 and a large part regulated by an autocatalytic system due to the ethylene-dependent expression of the same genes. The results further suggest that basal ethylene (system 1) is less likely to be involved in the transition to system 2. Even if the effect of system 1 ethylene is eliminated, fruit can show a small increase in ethylene production due to unknown developmental factors. This increase would be enough for the stimulation of autocatalytic ethylene production, leading to fruit ripening.

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Response of wild-type and RiEIL (T3 generation) tomatoes to exogenous ethylene. (A and B) Triple response assay in wild-type and RiEIL-18 seedlings (6 d after germination in the dark). Vertical bars are the SD (n=10). (C) Epinasty assay of plants. One-month-old plants were incubated in ethylene for 6 h. Arrows indicate bending points. No symptoms of triple response or epinasty were observed in RiEIL-18 plants even in 100 ppm ethylene.
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fig2: Response of wild-type and RiEIL (T3 generation) tomatoes to exogenous ethylene. (A and B) Triple response assay in wild-type and RiEIL-18 seedlings (6 d after germination in the dark). Vertical bars are the SD (n=10). (C) Epinasty assay of plants. One-month-old plants were incubated in ethylene for 6 h. Arrows indicate bending points. No symptoms of triple response or epinasty were observed in RiEIL-18 plants even in 100 ppm ethylene.

Mentions: In order to obtain transgenic tomato plants with severely suppressed LeEIL genes, an RNA interference construct using LeEIL2 fragments was employed (Fig. 1A). Six regenerated T0 plants expressed various levels of mRNAs of all four known LeEIL genes (Fig. 1B) in leaf tissue, with only RiEIL-10 and -18 reduced to trace levels of all four LeEIL genes and RiEIL-16 having a slightly higher level of LeEIL3. In fruit, mRNAs of the four LeEIL genes in RiEIL-10, -16, and 18 were reduced to trace levels (Fig. 1C). The three transgenic lines (RiEIL-10, -16, and -18) with severely suppressed LeEIL levels exhibited non-ripening phenotypes, as observed in transgenic tomatoes with antisense constructs (Tieman et al., 2001; Fig. 1D). The RiEIL-10 and -18 lines were selected for further experiments because they not only demonstrated a severe suppression of target genes but they had only a single copy of the transgene (data not shown). Only data from RiEIL-18 are presented in the figures as the results from RiEIL-10 were comparable. Wild-type fruit reached the turning stage at ∼40 DAA. On the other hand, the RiEIL-18 fruit showed no significant colour changes except for a faint yellow colour observed beyond 60 DAA. In addition, the petal remained attached to the fruit even at 60 DAA (Fig. 1D). The triple response assay and epinasty assay revealed distinct ethylene-insensitive phenotypes in the T3 generation of the RiEIL-18 line (Fig. 2). In the transgenic line, no reduction of seedling length or hook formation was observed, even in the presence of 100 ppm ethylene. A symptom of epinasty, petiole twisting, did not appear in the transgenic plants even with 100 ppm ethylene, while it was visible in the wild type with only 0.01 ppm ethylene. In the transgenic line, no colour change of the fruit or flower abscission was observed in the presence of 100 ppm ethylene (data not shown).


Ripening-associated ethylene biosynthesis in tomato fruit is autocatalytically and developmentally regulated.

Yokotani N, Nakano R, Imanishi S, Nagata M, Inaba A, Kubo Y - J. Exp. Bot. (2009)

Response of wild-type and RiEIL (T3 generation) tomatoes to exogenous ethylene. (A and B) Triple response assay in wild-type and RiEIL-18 seedlings (6 d after germination in the dark). Vertical bars are the SD (n=10). (C) Epinasty assay of plants. One-month-old plants were incubated in ethylene for 6 h. Arrows indicate bending points. No symptoms of triple response or epinasty were observed in RiEIL-18 plants even in 100 ppm ethylene.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2724697&req=5

fig2: Response of wild-type and RiEIL (T3 generation) tomatoes to exogenous ethylene. (A and B) Triple response assay in wild-type and RiEIL-18 seedlings (6 d after germination in the dark). Vertical bars are the SD (n=10). (C) Epinasty assay of plants. One-month-old plants were incubated in ethylene for 6 h. Arrows indicate bending points. No symptoms of triple response or epinasty were observed in RiEIL-18 plants even in 100 ppm ethylene.
Mentions: In order to obtain transgenic tomato plants with severely suppressed LeEIL genes, an RNA interference construct using LeEIL2 fragments was employed (Fig. 1A). Six regenerated T0 plants expressed various levels of mRNAs of all four known LeEIL genes (Fig. 1B) in leaf tissue, with only RiEIL-10 and -18 reduced to trace levels of all four LeEIL genes and RiEIL-16 having a slightly higher level of LeEIL3. In fruit, mRNAs of the four LeEIL genes in RiEIL-10, -16, and 18 were reduced to trace levels (Fig. 1C). The three transgenic lines (RiEIL-10, -16, and -18) with severely suppressed LeEIL levels exhibited non-ripening phenotypes, as observed in transgenic tomatoes with antisense constructs (Tieman et al., 2001; Fig. 1D). The RiEIL-10 and -18 lines were selected for further experiments because they not only demonstrated a severe suppression of target genes but they had only a single copy of the transgene (data not shown). Only data from RiEIL-18 are presented in the figures as the results from RiEIL-10 were comparable. Wild-type fruit reached the turning stage at ∼40 DAA. On the other hand, the RiEIL-18 fruit showed no significant colour changes except for a faint yellow colour observed beyond 60 DAA. In addition, the petal remained attached to the fruit even at 60 DAA (Fig. 1D). The triple response assay and epinasty assay revealed distinct ethylene-insensitive phenotypes in the T3 generation of the RiEIL-18 line (Fig. 2). In the transgenic line, no reduction of seedling length or hook formation was observed, even in the presence of 100 ppm ethylene. A symptom of epinasty, petiole twisting, did not appear in the transgenic plants even with 100 ppm ethylene, while it was visible in the wild type with only 0.01 ppm ethylene. In the transgenic line, no colour change of the fruit or flower abscission was observed in the presence of 100 ppm ethylene (data not shown).

Bottom Line: These results suggest that ripening-associated ethylene (system 2) in wild-type tomato fruit consists of two parts: a small part regulated by a developmental factor through the ethylene-independent expression of LeACS2 and LeACS4 and a large part regulated by an autocatalytic system due to the ethylene-dependent expression of the same genes.The results further suggest that basal ethylene (system 1) is less likely to be involved in the transition to system 2.Even if the effect of system 1 ethylene is eliminated, fruit can show a small increase in ethylene production due to unknown developmental factors.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Biological Sciences, 7549-1 Yoshikawa, Kibichuo-cho, Okayama, 716-1241 Japan.

ABSTRACT
To investigate the regulatory mechanism(s) of ethylene biosynthesis in fruit, transgenic tomatoes with all known LeEIL genes suppressed were produced by RNA interference engineering. The transgenic tomato exhibited ethylene insensitivity phenotypes such as non-ripening and the lack of the triple response and petiole epinasty of seedlings even in the presence of exogenous ethylene. Transgenic fruit exhibited a low but consistent increase in ethylene production beyond 40 days after anthesis (DAA), with limited LeACS2 and LeACS4 expression. 1-Methylcyclopropene (1-MCP), a potent inhibitor of ethylene perception, failed to inhibit the limited increase in ethylene production and expression of the two 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS) genes in the transgenic fruit. These results suggest that ripening-associated ethylene (system 2) in wild-type tomato fruit consists of two parts: a small part regulated by a developmental factor through the ethylene-independent expression of LeACS2 and LeACS4 and a large part regulated by an autocatalytic system due to the ethylene-dependent expression of the same genes. The results further suggest that basal ethylene (system 1) is less likely to be involved in the transition to system 2. Even if the effect of system 1 ethylene is eliminated, fruit can show a small increase in ethylene production due to unknown developmental factors. This increase would be enough for the stimulation of autocatalytic ethylene production, leading to fruit ripening.

Show MeSH