|YIN, JING - University Of California|
|CHANG, XIAOXIAO - University Of California|
|REID, MICHAEL - University Of California|
Submitted to: Horticulture Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/7/2015
Publication Date: 12/16/2015
Publication URL: http://doi:10.1038/hortres.2015.59
Citation: Yin, J., Chang, X., Kasuga, T., Bui, M.Q., Reid, M.S., Jiang, C. 2015. A basic helix-loop-helix transcription factor, PhFBH4, regulates flower senescence by modulating ethylene biosynthesis pathway in petunia. Horticulture Research. doi: 10.1038/hortres.2015.59.
Interpretive Summary: Flower senescence is an important coordinated process regulated by internal and environmental changes. Microarray studies of gene expression have been used to generate genome-wide transcriptome profiles of senescing petals in Arabidopsis. The data revealed that hundreds of upregulated and downregulated genes, including various transcription factors (TFs), are involved in flower senescence progress. TFs play critical roles in plant growth and development. The most represented families amongst TFs specifically upregulated during petal senescence in Arabidopsis were AP2-EREBP, homeobox (HB), and AUX-IAA.2 The upregulation of the AP2-EREB TFs establishes the role of ethylene in Arabidopsis. In petunia flowers, expression profiles of the ethylene-responsive element-binding factor (ERF) family genes were studied in detail. Some of ERFs appear to be associated with fruit ripening and with corolla senescence. Among the HB TFs upregulated in Arabidopsis petals was KNAT1, a member of the Class I KNOX family known to modulate cytokinin levels. The expression of genes encoding AUX-IAA proteins in Arabidopsis suggests the role of auxin in petal senescence. Nevertheless, exact roles of these regulatory elements in the process of flower senescence are still largely unknown. Petunia is an ideal model system for studies of flower senescence due to its short life cycle, vast genetic resources, and large number of amenities for biochemical and molecular analysis. We identified a cluster of genes upregulated during development and senescence of petunia flowers, including several transcription factors. In addition, we have successfully employed tobacco rattle virus (TRV)-based virus-induced gene silencing (VIGS) to study the function of senescence-related genes in petunia corollas. By microarray analysis, we previously determined that a homeodomain-leucine-zipper TF, PhHD-ZIP was upregulated during flower senescence. Silencing PhHD-Zip using VIGS resulted in extended flower longevity. Transcript abundances of ethylene biosynthesis related genes and ethylene production were dramatically reduced in the PhHD-Zip-silenced flowers. On the other hand, overexpression of PhHD-Zip in petunia caused early flower senescence. Furthermore, PhHD-Zip transcript levels in petunia flower were increased by hormones (ethylene, ABA) and abiotic stresses (dehydration, NaCl, and cold). The results suggest that PhHD-Zip plays an important role in regulating petunia flower senescence. From the same microarray study, a basic helix-loop-helix (bHLH) TF was also found to be highly expressed in flower petals and upregulated during the flower senescence process, suggesting that it may also play a role in the regulation of flower senescence. Here, we report the functional characterization of a bHLH TF, PhFBH4. Transcriptional level of PhFBH4 is induced by plant hormones and abiotic stress treatments. Silencing of PhFBH4 using virus-induced gene silencing and antisense approach extended flower longevity while transgenic petunia flowers with an over-expression construct showed a reduction in flower lifespan. Abundance of transcripts of senescence-related genes (SAG12, SAG29) was significantly changed in petunia PhFBH4 transgenic flowers. Furthermore, silencing or overexpression of PhFBH4 reduced or increased, respectively, transcript abundances of important ethylene biosynthesis-related genes ACS1 and ACO1, therefore influencing ethylene production. An electrophoretic mobility shift assay (EMSA) showed that PhFBH4 protein physically interacted with the G-box cis-element in the promoter of ACS1, suggesting that ACS1 was a direct target of PhFBH4 protein. In addition, ectopic expression of this gene altered plant development including plant height, internode length, and size of leaves and flowers, accompanying with alterations of transcript abundances in GA biosynthesis-related ge
Technical Abstract: The basic helix-loop-helix (bHLH) transcription factors (TFs) play important roles in regulating multiple biological processes in plants. However, there are few reports about the function of bHLHs in flower senescence. In this study, a bHLH TF, PhFBH4, was found to be dramatically upregulated during flower senescence. Transcription of PhFBH4 is induced by plant hormones and abiotic stress treatments. Silencing of PhFBH4 using virus-induced gene silencing or an antisense approach extended flower longevity, while transgenic petunia flowers with an overexpression construct showed a reduction in flower lifespan. Abundance of transcripts of senescence-related genes (SAG12, SAG29) was significantly changed in petunia PhFBH4 transgenic flowers. Furthermore, silencing or overexpression of PhFBH4 reduced or increased, respectively, transcript abundances of important ethylene biosynthesis-related genes, ACS1 and ACO1, thereby influencing ethylene production. An electrophoretic mobility shift assay showed that the PhFBH4 protein physically interacted with the G-box cis-element in the promoter of ACS1, suggesting that ACS1 was a direct target of the PhFBH4 protein. In addition, ectopic expression of this gene altered plant development including plant height, internode length, and size of leaves and flowers, accompanied by alteration of transcript abundance of the gibberellin biosynthesis-related gene GA2OX3. Our results indicate that PhFBH4 plays an important role in regulating plant growth and development through modulating the ethylene biosynthesis pathway.