The transcription factor PjERF1 enhances the biosynthesis of triterpenoid saponins in Panax japonicus

Chen Qin,Yu Yilin,Zhang Xiang,Zhao Ren,Zhang Jinyu,Liu Diqiu,Cui Xiuming,Ge Feng 한국식물생명공학회 2021 Plant biotechnology reports Vol.15 No.5

The ERF-type transcription factors (TFs) play vital roles in plant secondary metabolism. ERF TFs simultaneously regulate the expression levels of key enzyme genes involved in the biosynthesis of secondary metabolites due to its “multi-point control” function. In this study, one gene of ERF TFs from Panax japonicus (PjERF1) was cloned. The open reading frame of PjERF1 was 801 bp and encoded 266 amino acids. Phylogenetic analysis showed that PjERF1 belonged to ERF subfamily with a typical conserved domain. Subcellular localization found that PjERF1 protein might be located in eukaryotic cell nucleus. Yeast one-hybrid assay demonstrated that PjERF1 could bind to the promoters of PjβAS, PjCAS, and PjSE specifically and regulate the expression levels of such key enzyme genes involved in the triterpene saponins biosynthesis. Therefore, in the PjERF1 overexpression cell lines, the expression levels of some key enzyme genes involved in the triterpenoid saponins biosynthesis were significantly increased compared with those in non-transgenic cell line. As a result of it, the biosynthesis of chikusetsusaponin IV and IVa, and other ginsenosides (Rd, Rb1, Re, and R0) were also promoted in the PjERF1 over- expression cell lines. This study indicated that PjERF1 could regulate the biosynthesis of saponins in P. japonicus through controlling the expression levels of key enzyme genes related to the biosynthesis of triterpenoid saponins.

Analysis of the Cold-Responsive Transcriptome in the Mature Pollen of Arabidopsis

Zou Changsong,Yu Diqiu 한국식물학회 2010 Journal of Plant Biology Vol.53 No.6

Mature pollen is very sensitive to cold stress in chilling-sensitive plants. To understand the genetic regulation of tolerance to cold stress, we analyzed the transcript expression profile in mature pollen of Arabidopsis using Affymetrix GeneChips containing ∼24,000 genes. Expression of 2,127 genes was cold-regulated, of which 697 genes were upregulated and 1,430 genes were downregulated. Further analysis showed that a large number of signal transduction components were significantly affected by cold treatment, indicating extensive changes in the gene regulatory networks of mature pollen. Many coldresponsive genes encode transcription factors, suggesting a multitude of transcriptional cascades. A number of genes important for the biosynthesis or signaling of plant hormones, such as abscisic acid, auxin, and jasmonate,were regulated by cold stress, which is of potential importance in coordinating cold tolerance with pollen growth and development. In addition, 159 mature pollenspecific genes that might be involved in pollen viability were also cold-regulated. Expression of the cold-responsive transcripts identified by microarray analysis was confirmed by quantitative real-time PCR. Our study provides an overall picture of the cold-responsive transcriptome in Arabidopsis pollen and is valuable for understanding gene regulation in response to cold stress and the molecular mechanisms of cold tolerance in mature pollen.

WRKY22 Transcription Factor Mediates Dark-Induced Leaf Senescence in Arabidopsis

Xiang Zhou,Diqiu Yu,Yanjuan Jiang 한국분자세포생물학회 2011 Molecules and cells Vol.31 No.4

Arabidopsis WRKY proteins are plant-specific transcrip-tion factors, encoded by a large gene family, which contain the highly conserved amino acid sequence WRKYGQK and the zinc-finger-like motifs, Cys_2His_2 or Cys_2HisCys. They can recognize and bind the TTGAC(C/T) W-box cis-elements found in the promoters of target genes, and are involved in the regulation of gene expression during pathogen defense, wounding, trichome development, and senescence. Here we investigated the physiological function of the Arabidopsis WRKY22 transcription factor during dark-induced senescence. WRKY22 transcription was suppressed by light and promoted by darkness. In addi-tion, AtWRKY22 expression was markedly induced by H_2O_2. These results indicated that AtWRKY22 was involved in signal pathways in response to abiotic stress. Dark-treated AtWRKY22 over-expression and knockout lines showed accelerated and delayed senescence phenotypes, respectively, and senescence-associated genes exhibited increased and decreased expression levels. Mutual regulation existed between AtWRKY22 and AtWRKY6, AtWR-KY53, and AtWRKY70, respectively. Moreover, AtWRKY22 could influence their relative expression levels by feedback regulation or by other, as yet unknown mechanisms in response to dark. These results prove that AtWRKY22 participates in the dark-induced senescence signal transduction pathway.

Identification of an Arabidopsis Nodulin-Related Protein in Heat Stress

Qiantang Fu,Shujia Li,Diqiu Yu 한국분자세포생물학회 2010 Molecules and cells Vol.29 No.1

We identified a Nodulin-related protein 1 (NRP1) encoded by At2g03440, which was previously reported to be RPS2 interacting protein in yeast-two-hybrid assay. Northern blotting showed that AtNRP1 expression was suppressed by heat stress (42°C) and induced by low temperature (4°C) treatment. Strong GUS staining was observed in the sites of meristematic tissues of pAtNRP1:: GUS transgenic plants, such as shoot apex and root tips, young leaf veins, stamens and stigmas of flowers, and abscission layers of young siliques. To study AtNRP1 biological functions, we have characterized both loss-of-function T-DNA insertion and transgenic overexpression plants for AtNRP1 in Arabidopsis. The T-DNA insertion mutants displayed no obvious difference as compared to wild-type Arabidopsis under heat stress, but the significant enhanced suscepti-bility to heat stress was revealed in two independent AtNRP1-overexpressing transgenic lines. Further study found that the decreased thermtolerance in AtNRP1-overexpressing lines accompanied significantly decreased accumulation of ABA after heat treatment, which was probably due to AtNRP1 playing a role in negative-feedback regulation of the ABA synthesis pathway. These results support the viewpoint that the application of ABA inhibits nodulation and nodulin-related gene expression and threaten adverse ambient temperature can impact the nodulin-related gene expression.

Identification of an Arabidopsis Nodulin-Related Protein in Heat Stress

Fu, Qiantang,Li, Shujia,Yu, Diqiu Korean Society for Molecular and Cellular Biology 2010 Molecules and cells Vol.29 No.1

We identified a Nodulin-related protein 1 (NRP1) encoded by At2g03440, which was previously reported to be RPS2 interacting protein in yeast-two-hybrid assay. Northern blotting showed that AtNRP1 expression was suppressed by heat stress (42$^{\circ}C$) and induced by low temperature (4$^{\circ}C$) treatment. Strong GUS staining was observed in the sites of meristematic tissues of pAtNRP1:: GUS transgenic plants, such as shoot apex and root tips, young leaf veins, stamens and stigmas of flowers, and abscission layers of young siliques. To study AtNRP1 biological functions, we have characterized both loss-of-function T-DNA insertion and transgenic overexpression plants for AtNRP1 in Arabidopsis. The T-DNA insertion mutants displayed no obvious difference as compared to wild-type Arabidopsis under heat stress, but the significant enhanced susceptibility to heat stress was revealed in two independent AtNRP1-overexpressing transgenic lines. Further study found that the decreased thermtolerance in AtNRP1-overexpressing lines accompanied significantly decreased accumulation of ABA after heat treatment, which was probably due to AtNRP1 playing a role in negative-feedback regulation of the ABA synthesis pathway. These results support the viewpoint that the application of ABA inhibits nodulation and nodulin-related gene expression and threaten adverse ambient temperature can impact the nodulin-related gene expression.

Bacillus megaterium Strain XTBG34 Promotes Plant Growth by Producing 2-Pentylfuran

Changsong Zou,Zhifang Li,Diqiu Yu 한국미생물학회 2010 The journal of microbiology Vol.48 No.4

Several chemical changes in soil are associated with plant growth-promoting rhizobacteria. An endosporeforming bacterium, strain XTBG34, was isolated from a Xishuangbanna Tropical Botanical Garden soil sample and identified as Bacillus megaterium. The strain’s volatiles had remarkable plant growth promotion activity in Arabidopsis thaliana plants; after 15 days treatment, the fresh weight of plants inoculated with XTBG34 was almost 2-fold compared with those inoculated with DH5α. Head space volatile compounds produced by XTBG34, trapped with headspace solid phase microextraction and identified by gas chromatography–mass spectrometry, included aldehydes, alkanes, ketones and aroma components. Of the 11 compounds assayed for plant growth promotion activity in divided Petri plates, only 2-pentylfuran increased plant growth. We have therefore identified a new plant growth promotion volatile of B. megaterium XTBG34, which deserves further study in the mechanisms of interaction between plant growth-promoting rhizobacteria and plants.

Arabidopsis MiR396 Mediates the Development of Leaves and Flowers in Transgenic Tobacco

Fengxi Yang,Gang Liang,Dongmei Liu,Diqiu Yu 한국식물학회 2009 Journal of Plant Biology Vol.52 No.5

MicroRNAs (miRNAs) are single-stranded, noncoding small RNAs that usually function as posttranscriptional negative regulators by base pairing to target genes. They are pivotal to plant development. MiR396 is conserved among plant species and is predicted to target GRF (growth-regulating factor) genes in Arabidopsis. Here, overexpression of ath-miR396 in tobacco reduced the levels of three NtGRF-like genes containing an miR396 match site. Furthermore, its elevated expression resulted in a small, narrow leaf phenotype similar to that found with the Arabidopsis grf1grf2grf3 triple mutant. We also demonstrated that 35S:MIR396a transgenic plants were defective in the four whorls of floral organs. These results provide a link between the miR396- mediated regulatory pathway of NtGRF-like gene expression and the developmental processes for leaves and flowers in tobacco.

Functional Characterization of Arabidopsis thaliana WRKY39 in Heat Stress

Shujia Li,Xiang Zhou,Ligang Chen,Weidong Huang,Diqiu Yu 한국분자세포생물학회 2010 Molecules and cells Vol.29 No.5

Arabidopsis thaliana WRKY39, a transcription factor that is induced by heat stress, is a member of the group II WRKY proteins and responds to both abiotic and biotic stress. Heat-treated seeds and plants of WRKY39 knock-down mutants had increased susceptibility to heat stress, show-ing reduced germination, decreased survival, and elevated electrolyte leakage compared with wild-type plants. In con-trast, WRKY39 over-expressing plants exhibited enhanced thermotolerance compared with wild-type plants. RT-PCR and qRT-PCR analysis of wrky39 mutants and WRKY39 over-expressing plants identified putative genes regulated by WRKY39. Consistent with a role for WRKY39 in heat tolerance, the expression levels of salicylic acid (SA)-regulated PR1 and SA-related MBF1c genes were down-regulated in wrky39 mutants. In contrast, over-expression of WRKY39 increased the expression of PR1 and MBF1c. The WRKY39 transcript was induced in response to treat-ment with SA or methyljasmonate. Analysis of heat stress-induced WRKY39 in defense signaling mutants, including coi1, ein2, and sid2, further indicated that WRKY39 was positively co-regulated by the SA and jasmonate (JA) sig-naling pathways. Together, these findings reveal that heat stress-induced WRKY39 positively regulates the coopera-tion between the SA- and JA-activated signaling pathways that mediate responses to heat stress.

Functional Characterization of Arabidopsis thaliana WRKY39 in Heat Stress

Li, Shujia,Zhou, Xiang,Chen, Ligang,Huang, Weidong,Yu, Diqiu Korean Society for Molecular and Cellular Biology 2010 Molecules and cells Vol.29 No.5

Arabidopsis thaliana WRKY39, a transcription factor that is induced by heat stress, is a member of the group II WRKY proteins and responds to both abiotic and biotic stress. Heat-treated seeds and plants of WRKY39 knock-down mutants had increased susceptibility to heat stress, showing reduced germination, decreased survival, and elevated electrolyte leakage compared with wild-type plants. In contrast, WRKY39 over-expressing plants exhibited enhanced thermotolerance compared with wild-type plants. RT-PCR and qRT-PCR analysis of wrky39 mutants and WRKY39 over-expressing plants identified putative genes regulated by WRKY39. Consistent with a role for WRKY39 in heat tolerance, the expression levels of salicylic acid (SA)-regulated PR1 and SA-related MBF1c genes were down-regulated in wrky39 mutants. In contrast, over-expression of WRKY39 increased the expression of PR1 and MBF1c. The WRKY39 transcript was induced in response to treatment with SA or methyljasmonate. Analysis of heat stress-induced WRKY39 in defense signaling mutants, including coi1, ein2, and sid2, further indicated that WRKY39 was positively co-regulated by the SA and jasmonate (JA) signaling pathways. Together, these findings reveal that heat stress-induced WRKY39 positively regulates the cooperation between the SA- and JA-activated signaling pathways that mediate responses to heat stress.