Kaoru Suzuki

Background: Natural rubber (cis-1,4-polyioprene, NR) is an indispensable industrial raw material obtained from the Pará rubber tree (H. brasiliensis). Natural rubber cannot be replaced by synthetic rubber compounds because of the superior resilience, elasticity, abrasion resistance, efficient heat dispersion, and impact resistance of NR. In NR production, latex is harvested by periodical tapping of the trunk bark. Ethylene enhances and prolongs latex flow and latex regeneration. Ethephon, which is an ethylene-releasing compound, applied to the trunk before tapping usually results in a 1.5- to 2-fold increase in latex yield. However, intense mechanical damage to bark tissues by excessive tapping and/or over-stimulation with ethephon induces severe oxidative stress in laticifer cells, which often causes tapping panel dryness (TPD) syndrome. To enhance NR production without causing TPD, an improved understanding of the molecular mechanism of the ethylene response in the Pará rubber tree is required. Therefore, we investigated gene expression in response to ethephon treatment using Pará rubber tree seedlings as a model system. Results: After ethephon treatment, 3270 genes showed significant differences in expression compared with the mock treatment. Genes associated with carotenoids, flavonoids, and abscisic acid biosynthesis were significantly upregulated by ethephon treatment, which might contribute to an increase in latex flow. Genes associated with secondary cell wall formation were downregulated, which might be because of the reduced sugar supply. Given that sucrose is an important molecule for NR production, a trade-off may arise between NR production and cell wall formation for plant growth and for wound healing at the tapping panel. Conclusions: Dynamic changes in gene expression occur specifically in response to ethephon treatment. Certain genes identified may potentially contribute to latex production or TPD suppression. These data provide valuable information to understand the mechanism of ethylene stimulation, and will contribute to improved management practices and/or molecular breeding to attain higher yields of latex from Pará rubber trees.

Identification of the factors involved in the regulation of senescence and the analysis of their function are important for both a biological understanding of the senescence mechanism and the improvement of agricultural productivity. In this study, we identified an ERF gene termed "ERF gene conferring Postharvest longevity Improvement 1" (EPI1) as a possible regulator of senescence in Arabidopsis. We found that EPI1 possesses transcriptional repression activity and that the transgenic plants overexpressing EPI1 and expressing its chimeric repressor, EPI1-SRDX, commonly suppressed the darkness-induced senescence in their excised aerial parts. These transgenic plants additionally maintained a high level of chlorophyll, even after the methyl jasmonate (MeJA) treatment, which stimulated senescence in the dark. In addition, we found that senescence-induced and -reduced genes are down- and upregulated, respectively, in the MeJA-treated transgenic plants under darkness. Our results suggest that EPI1 functions as a negative regulator of the dark-induced and JA-stimulated senescence.

In this study, we characterized the function of WUSCHEL-RELATED HOMEOBOX 2 (WOX2) using overexpression, CRES-T, and VP16 fusion techniques. Although the function of WOX2 has been described mainly in embryogenesis, it was unclear whether it also plays a role in the post-embryogenic developmental stage. We found that WOX2 has transcriptional repression activity and that either overexpression of WOX2 or expression of its chimeric repressor causes severe growth defects and other morphological phenotypes by impairing plant organ formation and separation. By contrast, VP16-fused WOX2-expressing plants did not display such severe phenotypic defects. In addition, some of them displayed phenotypic defects such as fusion of organs and induction of undifferentiated cells in the boundary regions of organs where GUS staining was clearly observed in the proWOX2:GUS transgenic plants. We suggest that WOX2 is involved in regulation of lateral organ formation and separation during the post-embryogenic development processes.

Biological functions of only some plant transcriptional repressors are known owing to the lack of knockout lines or unclear phenotypes because of redundancy. Here we show that strong viral activation domain VP16 fusion to the transcriptional repressor FLOWERING LOCUS C reversed its function and caused a stronger phenotype than that of the multiple-knockout line of redundant genes, suggesting the potential of this technique to identify transcription factor function that cannot be detected in a single-knockout line. Loss-of-function of transcriptional coactivator Mediator25 did not affect VP16 activity despite their in vivo interaction, suggesting the existence of other key mechanism(s) in plants.

Plant growth and development require proper cell wall organization but little is known about the transcription factors responsible for the regulation of gene expression involved in cell wall organization. Here we show, using Arabidopsis thaliana, that constitutive expression of the chimeric repressor for the MYB87 transcription factor causes suppression of longitudinal elongation, aberrant radial growth, and radially expanded or swollen cells in multiple organs. Microarray analysis revealed that plants expressing the chimeric repressor have altered expression of various cell wall related genes. MYB87 may therefore function as a regulator of genes affecting cell wall organization and remodeling. These findings improve our understanding of cell wall regulation and its roles in plant growth and development and also contribute information that may allow engineering of plant growth and architecture.

Jasmonates (JAs) are plant hormones that regulate the balance between plant growth and responses to biotic and abiotic stresses. Although recent studies have uncovered the mechanisms for JA-induced responses in Arabidopsis thaliana, the mechanisms by which plants attenuate the JA-induced responses remain elusive. Here, we report that a basic helix-loop-helix–type transcription factor, ABA-INDUCIBLE BHLH-TYPE TRANSCRIPTION FACTOR/JA-ASSOCIATED MYC2-LIKE1 (JAM1), acts as a transcriptional repressor and negatively regulates JA signaling. Gain-of-function transgenic plants expressing the chimeric repressor for JAM1 exhibited substantial reduction of JA responses, including JA-induced inhibition of root growth, accumulation of anthocyanin, and male fertility. These plants were also compromised in resistance to attack by the insect herbivore Spodoptera exigua. Conversely, jam1 loss-of-function mutants showed enhanced JA responsiveness, including increased resistance to insect attack. JAM1 and MYC2 competitively bind to the target sequence of MYC2, which likely provides the mechanism for negative regulation of JA signaling and suppression of MYC2 functions by JAM1. These results indicate that JAM1 negatively regulates JA signaling, thereby playing a pivotal role in fine-tuning of JA-mediated stress responses and plant growth.

Ethylene-responsive transcription factor (ERF) family genes, which are involved in regulation of metabolic pathways and/or are useful for metabolic engineering, were investigated in the cultured cells of Arabidopsis thaliana. The pectin content in the gelatinous precipitates after the ethanol precipitation of extracts derived from calli of a transgenic cell line, A17, overexpressing an ERF gene (At1g44830), increased in comparison with the control. Expression of genes involved in pectin biosynthesis was up-regulated in the A17 calli. Overexpression of the ERF gene coordinately activates the pectin biosynthetic pathway genes and increases the content of pectin. These results therefore will be useful as a genetic resource for engineering pectin biosynthesis in plants.