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Journal Cover Plant Molecular Biology
  [SJR: 1.915]   [H-I: 137]   [9 followers]  Follow
   Hybrid Journal Hybrid journal (It can contain Open Access articles)
   ISSN (Print) 1573-5028 - ISSN (Online) 0167-4412
   Published by Springer-Verlag Homepage  [2336 journals]
  • UV-mediated Chlamydomonas mutants with enhanced nuclear transgene
           expression by disruption of DNA methylation-dependent and independent
           silencing systems
    • Authors: Sari Dewi Kurniasih; Tomohito Yamasaki; Fantao Kong; Sigeru Okada; Dwiyantari Widyaningrum; Takeshi Ohama
      Pages: 629 - 641
      Abstract: Key message In this investigation, we succeeded to generate Chlamydomonas mutants that bear dramatically enhanced ability for transgene expression. To yield these mutants, we utilized DNA methyltransferase deficient strain. These mutants must be useful as a plant cell factory. Chlamydomonas reinhardtii (hereafter Chlamydomonas) is a green freshwater microalga. It is a promising cell factory for the production of recombinant proteins because it rapidly grows in simple salt-based media. However, expression of transgenes integrated into the nuclear genome of Chlamydomonas is very poor, probably because of severe transcriptional silencing irrespective of the genomic position. In this study, we generated Chlamydomonas mutants by ultraviolet (UV)-mediated mutagenesis of maintenance-type DNA methyltransferase gene (MET1)-null mutants to overcome this disadvantage. We obtained several mutants with an enhanced ability to overexpress various transgenes irrespective of their integrated genomic positions. In addition, transformation efficiencies were significantly elevated. Our findings indicate that in addition to mechanisms involving MET1, transgene expression is regulated by a DNA methylation-independent transgene silencing system in Chlamydomonas. This is in agreement with the fact that DNA methylation occurs rarely in this organism. The generated mutants may be useful for the low-cost production of therapeutic proteins and eukaryotic enzymes based on their rapid growth in simple salt-based media.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0529-9
      Issue No: Vol. 92, No. 6 (2016)
  • SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 2 controls floral organ development
           and plant fertility by activating ASYMMETRIC LEAVES 2 in Arabidopsis
    • Authors: Zhishuo Wang; Ying Wang; Susanne E. Kohalmi; Lisa Amyot; Abdelali Hannoufa
      Pages: 661 - 674
      Abstract: Abstract A network of genes is coordinately expressed to ensure proper development of floral organs and fruits, which are essential for generating new offspring in flowering plants. In Arabidopsis thaliana, microRNA156 (miR156) plays a role in regulating the development of flowers and siliques by targeting members of the SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) gene family. Despite the important roles of the miR156/SPL network, our understanding of its downstream genes that are involved in floral organ and silique growth is still incomplete. Here, we report that the miR156/SPL2 regulatory pathway regulates pollen production, fertility rate, and the elongation of floral organs, including petals, sepals, and siliques in Arabidopsis. Transgenic plants exhibiting both overexpression of miR156 and dominant-negative alleles of SPL2 had reduced ASYMMETRIC LEAVES 2 (AS2) transcript levels in their siliques. Furthermore, their fertility phenotype was similar to that of the AS2 loss-of-function mutant. We also demonstrate that the SPL2 protein binds to the 5′UTR of the AS2 gene in vivo, indicating that AS2 is directly regulated by SPL2. Our results suggest that the miR156/SPL2 pathway affects floral organs, silique development and plant fertility, as well as directly regulates AS2 expression.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0536-x
      Issue No: Vol. 92, No. 6 (2016)
  • Trichome patterning control involves TTG1 interaction with SPL
           transcription factors
    • Authors: Eugenia Ioannidi; Stamatis Rigas; Dikran Tsitsekian; Gerasimos Daras; Anastasios Alatzas; Antonis Makris; Georgia Tanou; Anagnostis Argiriou; Dimitrios Alexandrou; Scott Poethig; Polydefkis Hatzopoulos; Angelos K. Kanellis
      Pages: 675 - 687
      Abstract: Abstract Epidermal cell differentiation is a paramount and conserved process among plants. In Arabidopsis, a ternary complex formed by MYB, bHLH transcription factors and TTG1 modulates unicellular trichome morphogenesis. The formation of multicellular glandular trichomes of the xerophytic shrub Cistus creticus that accumulate labdane-type diterpenes, has attained much attention renowned for its medicinal properties. Here, we show that C. creticus TTG1 (CcTTG1) interacts with the SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPLA/B) proteins, putative homologs of AtSPL4/5 that in turn interact with AtTTG1. These interactions occur between proteins from evolutionarily distant species supporting the conserved function of TTG1-SPL complex. Overexpression of AtSPL4 and AtSPL5 decreased the expression of GLABRA2 (AtGL2), the major regulator of trichome morphogenesis, resulting in trichome reduction on the adaxial surface of cauline leaves, thereby illuminating the significance of TTG1-SPLs interactions in trichome formation control. AtGL2 and AtSPL4 have opposite expression patterns during early stages of leaf development. We postulate an antagonistic effect between SPLs and the heterogeneous MYB-bHLH factors binding to TTG1. Hence, the SPLs potentially rearrange the complex, attenuating its transcriptional activity to control trichome distribution.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0538-8
      Issue No: Vol. 92, No. 6 (2016)
  • ER-localized adenine nucleotide transporter ER-ANT1: an integrator of
           energy and stress signaling in rice
    • Authors: Xiangqian Zhang; Xu Zheng; Shanwen Ke; Haitao Zhu; Fang Liu; Zemin Zhang; Xinxiang Peng; Lin Guo; Ruizhen Zeng; Pei Hou; Ziqiang Liu; Suowei Wu; Meifang Song; Jianping Yang; Guiquan Zhang
      Pages: 701 - 715
      Abstract: Abstract Most environmental perturbations have a direct or indirect deleterious impact on photosynthesis, and, in consequence, the overall energy status of the cell. Despite our increased understanding of convergent energy and stress signals, the connections between photosynthesis, energy and stress signals through putative common nodes are still unclear. Here we identified an endoplasmic reticulum (ER)-localized adenine nucleotide transporter1 (ER-ANT1), whose deficiency causes seedling lethality in air but viable under high CO2, exhibiting the typical photorespiratory phenotype. Metabolic analysis suggested that depletion of ER-ANT1 resulted in circadian rhythm disorders in sucrose synthesis and induced sucrose signaling pathways, indicating that the ER is involved in the regulation of vital energy metabolism in plants. In addition, the defect of ER-ANT1 triggers ER stress and activates the unfolded protein response in plant cells, suggesting ER stress and photorespiration are closely linked. These findings provide an important evidence for a key role of ER-localized ER-ANT1 in convergent energy and stress signals in rice. Our findings support the idea that ATP is a central signal involved in the plant response to a variety of stresses.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0540-1
      Issue No: Vol. 92, No. 6 (2016)
  • Arabidopsis acyl-CoA-binding protein ACBP6 localizes in the phloem and
           affects jasmonate composition
    • Authors: Zi-Wei Ye; Shiu-Cheung Lung; Tai-Hua Hu; Qin-Fang Chen; Yung-Lee Suen; Mingfu Wang; Susanne Hoffmann-Benning; Edward Yeung; Mee-Len Chye
      Pages: 717 - 730
      Abstract: Abstract Arabidopsis thaliana ACYL-COA-BINDING PROTEIN6 (AtACBP6) encodes a cytosolic 10-kDa AtACBP. It confers freezing tolerance in transgenic Arabidopsis, possibly by its interaction with lipids as indicated by the binding of acyl-CoA esters and phosphatidylcholine to recombinant AtACBP6. Herein, transgenic Arabidopsis transformed with an AtACBP6 promoter-driven β-glucuronidase (GUS) construct exhibited strong GUS activity in the vascular tissues. Immunoelectron microscopy using anti-AtACBP6 antibodies showed AtACBP6 localization in the phloem especially in the companion cells and sieve elements. Also, the presence of gold grains in the plasmodesmata indicated its potential role in systemic trafficking. The AtACBP6 protein, but not its mRNA, was found in phloem exudate of wild-type Arabidopsis. Fatty acid profiling using gas chromatography-mass spectrometry revealed an increase in the jasmonic acid (JA) precursor, 12-oxo-cis,cis-10,15-phytodienoic acid (cis-OPDA), and a reduction in JA and/or its derivatives in acbp6 phloem exudates in comparison to the wild type. Quantitative real-time PCR showed down-regulation of COMATOSE (CTS) in acbp6 rosettes suggesting that AtACBP6 affects CTS function. AtACBP6 appeared to affect the content of JA and/or its derivatives in the sieve tubes, which is consistent with its role in pathogen-defense and in its wound-inducibility of AtACBP6pro::GUS. Taken together, our results suggest the involvement of AtACBP6 in JA-biosynthesis in Arabidopsis phloem tissues.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0541-0
      Issue No: Vol. 92, No. 6 (2016)
  • Salinity-mediated transcriptional and post-translational regulation of the
           Arabidopsis aquaporin PIP2;7
    • Authors: Alicia Pou; Linda Jeanguenin; Thomas Milhiet; Henri Batoko; François Chaumont; Charles Hachez
      Pages: 731 - 744
      Abstract: Key message Salt stress triggers a simultaneous transcriptional repression and aquaporin internalization to modify root cell water conductivity. Plasma membrane intrinsic proteins (PIPs) are involved in the adjustment of plant water balance in response to changing environmental conditions. In this study, Arabidopsis wild-type (Col-0) and transgenic lines overexpressing PIP2;7 were used to investigate and compare their response to salt stress. Hydraulic conductivity measurements using a high-pressure flowmeter (HPFM) revealed that overexpression of PIP2;7 induced a sixfold increase in root hydraulic conductivity of four week-old Arabidopsis thaliana plants compared to WT. Exposure to a high salt stress (150 mM NaCl) triggered a rapid repression of overall aquaporin activity in both genotypes. Response to salt stress was also investigated in 8 day-old seedlings. Exposure to salt led to a repression of PIP2;7 promoter activity and a significant decrease in PIP2;7 mRNA abundance within 2 h. Concomitantly, a rapid internalization of fluorescently-tagged PIP2;7 proteins was observed but removal from the cell membrane was not accompanied by further degradation of the protein within 4 h of exposure to salinity stress. These data suggest that PIP transcriptional repression and channel internalization act in concert during salt stress conditions to modulate aquaporin activity, thereby significantly altering the plant hydraulic parameters in the short term.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0542-z
      Issue No: Vol. 92, No. 6 (2016)
  • Over-expression of miR158 causes pollen abortion in Brassica campestris
           ssp. chinensis
    • Authors: Zhiming Ma; Jianxia Jiang; Ziwei Hu; Tianqi Lyu; Yang Yang; Jingjing Jiang; Jiashu Cao
      Abstract: Key message We identified and cloned the two precursors of miR158 and its target gene in Brassica campestris ssp. chinensis, which both had high relative expression in the inflorescences. Further study revealed that over-expression of miR158 caused reduced pollen varbility, which was caused by the degradation of pollen contents from the binucleate microspore stage. These results first suggest the role of miR158 in pollen development of Brassica campestris ssp. chinensis. MicroRNAs (miRNAs) play crucial roles in many important growth and development processes both in plants and animals by regulating the expression of their target genes via mRNA cleavage or translational repression. In this study, miR158, a Brassicaceae specific miRNA, was functionally characterized with regard to its role in pollen development of non-heading Chinese cabbage (Brassica campestris ssp. chinensis). Two family members of miR158 in B. campestris, namely bra-miR158a1 and bra-miR158a2, and their target gene bra027656, which encodes a pentatricopeptide repeat (PPR) containing protein, were identified. Then, qRT-PCR analysis and GUS-reporter system revealed that both bra-miR158 and its target gene had relatively high expression levels in the inflorescences. Further study revealed that over-expression of miR158 caused reduced pollen varbility and pollen germination ratio, and the degradation of pollen contents from the binucleate microspore stage was also found in those deformed pollen grains, which led to pollen shrinking and collapse in later pollen development stage. These results first shed light on the importance of miR158 in pollen development of Brassica campestris ssp. chinensis.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0563-7
  • Multidimensional patterns of metabolic response in abiotic stress-induced
           growth of Arabidopsis thaliana
    • Abstract: Key message Contextualization of specific transcriptional responses of Arabidopsis within the stress–tissue–time perspective provides a simplified representation of the cellular transcriptional response pathways to abiotic stress, while reducing the dimensions in gene-oriented response description. Crops resistant to abiotic stresses are a long-term goal of many research programs, thus understanding the progression of stress responses is of great interest. We reanalyzed the AtGenExpress transcription dataset to go beyond gene-level characterization, and to contextualize the discrete information into (1) a process-level signature of stress-specific, time-specific, and tissue-specific responses and (2) identify patterns of response progression across a time axis. To gain a functional perspective, ∼1000 pathways associated with the differentially-expressed genes were characterized across all experiments. We find that the global response of pathways to stress is multi-dimensional and does not obviously cluster according to stress, time or tissue. The early response to abiotic stress typically involves induction of genes involved in transcription, hormone synthesis and signaling modules; a later response typically involves metabolism of amino acids and secondary metabolites. By linking specific primary and secondary response pathways, we outline possible stress-associated routes of response progression. The contextualization of specific processes within stress–tissue–time perspective provides a simplified representation of cellular response while reducing the dimensions in gene-oriented response description. Such simplified representation allows finding stress-specific markers based on process-combinations pointing whether a stress-specific response was invoked as well as provide a reference point for the conductance of comparative inter-plant study of stress response, bypassing the need in detailed orthologous mapping.
      PubDate: 2016-12-01
      DOI: 10.1007/s11103-016-0539-7
  • Arabidopsis thaliana methionine sulfoxide reductase B8 influences
           stress-induced cell death and effector-triggered immunity
    • Authors: Shweta Roy; Ashis Kumar Nandi
      Abstract: Key message Reactive oxygen species (ROS) oxidize methionine to methionine sulfoxide (MetSO) and thereby inactivate proteins. Methionine sulfoxide reductase (MSR) enzyme converts MetSO back to the reduced form and thereby detoxifies the effect of ROS. Our results show that Arabidopsis thaliana MSR enzyme coding gene MSRB8 is required for effector-triggered immunity and containment of stress-induced cell death in Arabidopsis. Plants activate pattern-triggered immunity (PTI), a basal defense, upon recognition of evolutionary conserved molecular patterns present in the pathogens. Pathogens release effector molecules to suppress PTI. Recognition of certain effector molecules activates a strong defense, known as effector-triggered immunity (ETI). ETI induces high-level accumulation of reactive oxygen species (ROS) and hypersensitive response (HR), a rapid programmed death of infected cells. ROS oxidize methionine to methionine sulfoxide (MetSO), rendering several proteins nonfunctional. The methionine sulfoxide reductase (MSR) enzyme converts MetSO back to the reduced form and thereby detoxifies the effect of ROS. Though a few plant MSR genes are known to provide tolerance against oxidative stress, their role in plant–pathogen interaction is not known. We report here that activation of cell death by avirulent pathogen or UV treatment induces expression of MSRB7 and MSRB8 genes. The T-DNA insertion mutant of MSRB8 exaggerates HR-associated and UV-induced cell death and accumulates a higher level of ROS than wild-type plants. The negative regulatory role of MSRB8 in HR is further supported by amiRNA and overexpression lines. Mutants and overexpression lines of MSRB8 are susceptible and resistant respectively, compared to the wild-type plants, against avirulent strains of Pseudomonas syringae pv. tomato DC3000 (Pst) carrying AvrRpt2, AvrB, or AvrPphB genes. However, the MSRB8 gene does not influence resistance against virulent Pst or P. syringae pv. maculicola (Psm) pathogens. Our results altogether suggest that MSRB8 function is required for ETI and containment of stress-induced cell death in Arabidopsis.
      PubDate: 2016-11-29
      DOI: 10.1007/s11103-016-0550-z
  • Phosphate starvation induced OsPHR4 mediates Pi-signaling and homeostasis
           in rice
    • Authors: Wenyuan Ruan; Meina Guo; Ping Wu; Keke Yi
      Abstract: Key message OsPHR4 mediates the regulation of Pi-starvation signaling and Pi-homeostasis in a PHR1-subfamily dependent manner in rice. Phosphate (Pi) starvation response is a sophisticated process for plant in the natural environment. In this process, PHOSPHATE STARVATION RESPONSE 1 (PHR1) subfamily genes play a central role in regulating Pi-starvation signaling and Pi-homeostasis. Besides the three PHR1 orthologs in Oryza sativa L. (Os) [(Os) PHR1, (Os) PHR2, and (Os) PHR3], which were reported to regulated Pi-starvation signaling and Pi-homeostasis redundantly, a close related PHR1 ortholog [designated as (Os) PHR4] is presented in rice genome with unknown function. In this study, we found that OsPHR4 is a Pi-starvation induced gene and mainly expresses in vascular tissues through all growth and development periods. The expression of OsPHR4 is positively regulated by OsPHR1, OsPHR2 and OsPHR3. The nuclear located OsPHR4 can respectively interact with other three PHR1 subfamily members to regulate downstream Pi-starvation induced genes. Consistent with the positive role of PHR4 in regulating Pi-starvation signaling, the OsPHR4 overexpressors display higher Pi accumulation in the shoot and elevated expression of Pi-starvation induced genes under Pi-sufficient condition. Besides, moderate growth retardation and repression of the Pi-starvation signaling in the OsPHR4 RNA interfering (RNAi) transgenic lines can be observed under Pi-deficient condition. Together, we propose that OsPHR4 mediates the regulation of Pi-starvation signaling and Pi-homeostasis in a PHR1-subfamily dependent manner in rice.
      PubDate: 2016-11-23
      DOI: 10.1007/s11103-016-0564-6
  • Two MYB-related transcription factors play opposite roles in sugar
           signaling in Arabidopsis
    • Authors: Yi-Shih Chen; Yi-Chi Chao; Tzu-Wei Tseng; Chun-Kai Huang; Pei-Ching Lo; Chung-An Lu
      Abstract: Key message Sugar regulation of gene expression has profound effects at all stages of the plant life cycle. Although regulation at the transcriptional level is one of the most prominent mechanisms by which gene expression is regulated, only a few transcription factors have been identified and demonstrated to be involved in the regulation of sugar-regulated gene expression. OsMYBS1, an R1/2-type MYB transcription factor, has been demonstrated to be involved in sugar- and hormone-regulated α-amylase gene expression in rice. Arabidopsis contains two OsMYBS1 homologs. In the present study, we investigate MYBS1 and MYBS2 in sugar signaling in Arabidopsis. Our results indicate that MYBS1 and MYBS2 play opposite roles in regulating glucose and ABA signaling in Arabidopsis during seed germination and early seedling development. MYB proteins have been classified into four subfamilies: R2R3-MYB, R1/2-MYB, 3R-MYB, and 4R-MYB. An R1/2-type MYB transcription factor, OsMYBS1, has been demonstrated to be involved in sugar- and hormone-regulated α-amylase genes expression in rice. In this study, two genes homologous to OsMYBS1, MYBS1 and MYBS2, were investigated in Arabidopsis. Subcellular localization analysis showed that MYBS1 and MYBS2 were localized in the nucleus. Rice embryo transient expression assays indicated that both MYBS1 and MYBS2 could recognize the sugar response element, TA-box, in the promoter and induced promoter activity. mybs1 mutant exhibited hypersensitivity to glucose, whereas mybs2 seedlings were hyposensitive to it. MYBS1 and MYBS2 are involved in the control of glucose-responsive gene expression, as the mybs1 mutant displayed increased expression of a hexokinase gene (HXK1), chlorophyll a/b-binding protein gene (CAB1), ADP-glucose pyrophosphorylase gene (APL3), and chalcone synthase gene (CHS), whereas the mybs2 mutant exhibited decreased expression of these genes. mybs1 also showed an enhanced response to abscisic acid (ABA) in the seed germination and seedling growth stages, while mybs2 showed reduced responses. The ABA biosynthesis inhibitor fluridone rescued the mybs1 glucose-hypersensitive phenotype. Moreover, the mRNA levels of three ABA biosynthesis genes, ABA1, NCED9, and AAO3, and three ABA signaling genes, ABI3, ABI4, and ABI5, were increased upon glucose treatment of mybs1 seedlings, but were decreased in mybs2 seedlings. These results indicate that MYBS1 and MYBS2 play opposite roles in regulating glucose and ABA signaling in Arabidopsis during seed germination and early seedling development.
      PubDate: 2016-11-19
      DOI: 10.1007/s11103-016-0562-8
  • Transgenic citrus expressing synthesized cecropin B genes in the phloem
           exhibits decreased susceptibility to Huanglongbing
    • Authors: Xiuping Zou; Xueyou Jiang; Lanzhen Xu; Tiangang Lei; Aihong Peng; Yongrui He; Lixiao Yao; Shanchun Chen
      Abstract: Key message Expression of synthesized cecropin B genes in the citrus phloem, where Candidatus Liberibacter asiaticus resides, significantly decreased host susceptibility to Huanglongbing. Huanglongbing (HLB), associated with Candidatus Liberibacter asiaticus bacteria, is the most destructive disease of citrus worldwide. All of the commercial sweet orange cultivars lack resistance to this disease. The cationic lytic peptide cecropin B, isolated from the Chinese tasar moth (Antheraea pernyi), has been shown to effectively eliminate bacteria. In this study, we demonstrated that transgenic citrus (Citrus sinensis Osbeck) expressing the cecropin B gene specifically in the phloem had a decreased susceptibility to HLB. Three plant codon-optimized synthetic cecropin B genes, which were designed to secrete the cecropin B peptide into three specific sites, the extracellular space, the cytoplasm, and the endoplasmic reticulum, were constructed. Under the control of the selected phloem-specific promoter GRP1.8, these constructs were transferred into the citrus genome. All of the cecropin B genes were efficiently expressed in the phloem of transgenic plants. Over more than a year of evaluation, the transgenic lines exhibited reduced disease severity. Bacterial populations in transgenic lines were significantly lower than in the controls. Two lines, in which bacterial populations were significantly lower than in others, showed no visible symptoms. Thus, we demonstrated the potential application of the phloem-specific expression of an antimicrobial peptide gene to protect citrus plants from HLB.
      PubDate: 2016-11-19
      DOI: 10.1007/s11103-016-0565-5
  • A bifunctional aminoglycoside acetyltransferase/phosphotransferase
           conferring tobramycin resistance provides an efficient selectable marker
           for plastid transformation
    • Authors: Iman Tabatabaei; Stephanie Ruf; Ralph Bock
      Abstract: Key message A new selectable marker gene for stable transformation of the plastid genome was developed that is similarly efficient as the aadA, and produces no background of spontaneous resistance mutants. More than 25 years after its development for Chlamydomonas and tobacco, the transformation of the chloroplast genome still represents a challenging technology that is available only in a handful of species. The vast majority of chloroplast transformation experiments conducted thus far have relied on a single selectable marker gene, the spectinomycin resistance gene aadA. Although a few alternative markers have been reported, the aadA has remained unrivalled in efficiency and is, therefore, nearly exclusively used. The development of new marker genes for plastid transformation is of crucial importance to all efforts towards extending the species range of the technology as well as to those applications in basic research, biotechnology and synthetic biology that involve the multistep engineering of plastid genomes. Here, we have tested a bifunctional resistance gene for its suitability as a selectable marker for chloroplast transformation. The bacterial enzyme aminoglycoside acetyltransferase(6′)-Ie/aminoglycoside phosphotransferase(2″)-Ia possesses an N-terminal acetyltransferase domain and a C-terminal phosphotransferase domain that can act synergistically and detoxify aminoglycoside antibiotics highly efficiently. We report that, in combination with selection for resistance to the aminoglycoside tobramycin, the aac(6′)-Ie/aph(2″)-Ia gene represents an efficient marker for plastid transformation in that it produces similar numbers of transplastomic lines as the spectinomycin resistance gene aadA. Importantly, no spontaneous antibiotic resistance mutants appear under tobramycin selection.
      PubDate: 2016-11-17
      DOI: 10.1007/s11103-016-0560-x
  • Metabolo-transcriptome profiling of barley reveals induction of chitin
           elicitor receptor kinase gene ( HvCERK1 ) conferring resistance against
           Fusarium graminearum
    • Authors: Shailesh Karre; Arun Kumar; Dhananjay Dhokane; Ajjamada C. Kushalappa
      Abstract: Key message We report plausible disease resistance mechanisms induced by barley resistant genotype CI89831 against Fusarium head blight (FHB) based on metabolo-transcriptomics approach. We identified HvCERK1 as a candidate gene for FHB resistance, which is functional in resistant genotype CI9831 but non-functional in susceptible cultivars H106-371 and Zhedar-2. For the first time, we were able to show a hierarchy of regulatory genes that regulated downstream biosynthetic genes that eventually produced resistance related metabolites that reinforce the cell walls to contain the pathogen progress in plant. The HvCERK1 can be used for replacing in susceptible commercial cultivars, if non-functional, based on genome editing. Fusarium head blight (FHB) management is a great challenge in barley and wheat production worldwide. Though barley genome sequence and advanced omics technologies are available, till date none of the resistance mechanisms has been clearly deciphered. Hence, this study was aimed at identifying candidate gene(s) and elucidating resistance mechanisms induced by barley resistant genotype CI9831 based on integrated metabolomics and transcriptomics approach. Following Fusarium graminearum infection, we identified accumulation of specific set of induced secondary metabolites, belonging to phenylpropanoid, hydroxycinnamic acid (HCAA) and jasmonic acid pathways, and their biosynthetic genes. In association with these, receptor kinases such as chitin elicitor receptor kinase (HvCERK1) and protein kinases such as MAP kinase 3 (HvMPK3) and MAPK substrate 1 (HvMKS1), and transcription factors such as HvERF1/5, HvNAC42, HvWRKY23 and HvWRKY70 were also found upregulated with high fold change. Polymorphism studies across three barley genotypes confirmed the presence of mutations in HvCERK1 gene in two susceptible genotypes, isolating this gene as a potential candidate for FHB resistance. Further, the silencing of functional HvCERK1 gene in the resistant genotype CI9831, followed by gene expression and metabolite analysis revealed its role as an elicitor recognition receptor that triggered downstream regulatory genes, which in turn, regulated downstream metabolic pathway genes to biosynthesize resistance related (RR) metabolites to contain the pathogen to spikelet infection. A putative model on metabolic pathway regulation is proposed.
      PubDate: 2016-11-14
      DOI: 10.1007/s11103-016-0559-3
  • The far-upstream regulatory region of RFL is required for its precise
           spatial-temporal expression for floral development in rice
    • Authors: Sulin Lou; Shuifu Chen; Xiucai Zhao; Letian Chen; Jian Zhang; Hongxiang Fu; Yao-Guang Liu; Yuanling Chen
      Abstract: Key message A rice mutant aberrant floral organ 1 (afo1) was identified, showing increased floral organ number, aberrant floral organ identity and loss of floral meristem determinacy. A disruption of sequence integrity at 6292-bp upstream of RFL by a T-DNA insertion led to varied RFL expression patterns in floral meristem and floret in afo1 and caused the mutant phenotype. The LEAFY (LFY) transcription factor and its homologs affect many aspects of plant development, especially floral development. RICE FLORICAULA/LEAFY (RFL), the rice ortholog of LFY, has complicated expression patterns and different functions in floral development. However, the mechanisms regulating the spatial-temporal expression of RFL remain largely unknown. Here, we describe a rice aberrant floral organ 1 (afo1) mutant that was produced by a T-DNA insertion at 6292-bp upstream of the start codon of RFL. This insertion altered the expression of RFL in floral meristem (FM) and floret. The in situ hybridization result showed that, when florets appear, RFL was expressed almost exclusively at the palea/lemma adaxial base adjacent to lodicules in the wild-type panicle. However, in afo1 florets, RFL mRNA signals were detected in the region between lodicule and stamen, and strong signals persisted in FM. The altered pattern of RFL expression in afo1 resulted in enlarged FMs, more floral organs, aberrant floral organ identity, and loss of FM determinacy. Transformation of rice with an RFL construct driven by the 6292-bp upstream genomic sequence re-built the mutant phenotype similar to afo1. The results suggest that the far-upstream region of RFL may contain potential cis element(s) that are critical to define the precise spatial-temporal expression pattern of RFL for its function in floral development.
      PubDate: 2016-11-12
      DOI: 10.1007/s11103-016-0556-6
  • Kelch-motif containing acyl-CoA binding proteins AtACBP4 and AtACBP5 are
           differentially expressed and function in floral lipid metabolism
    • Authors: Zi-Wei Ye; Jie Xu; Jianxin Shi; Dabing Zhang; Mee-Len Chye
      Abstract: Key message We herein demonstrated two of the Arabidopsis acyl-CoA-binding proteins (ACBPs), AtACBP4 and AtACBP5, both function in floral lipid metabolism and they may possibly play complementary roles in Arabidopsis microspore-to-pollen development. Histological analysis on transgenic Arabidopsis expressing β-glucuronidase driven from the AtACBP4 and AtACBP5 promoters, as well as, qRTPCR analysis revealed that AtACBP4 was expressed at stages 11–14 in the mature pollen, while AtACBP5 was expressed at stages 7–10 in the microspores and tapetal cells. Immunoelectron microscopy using AtACBP4- or AtACBP5-specific antibodies further showed that AtACBP4 and AtACBP5 were localized in the cytoplasm. Chemical analysis of bud wax and cutin using gas chromatographyflame ionization detector and GC-mass spectrometry analyses revealed the accumulation of cuticular waxes and cutin monomers in acbp4, acbp5 and acbp4acbp5 buds in comparison to the wild type (Col-0). Fatty acid profiling demonstrated a decline in stearic acid and an increase in linolenic acid in acbp4 and acbp4acbp5 buds, respectively, over Col-0. Analysis of inflorescences from acbp4 and acbp5 revealed that there was an increase of AtACBP5 expression in acbp4, and an increase of AtACBP4 expression in acbp5. Deletion analysis of the AtACBP4 and AtACBP5 5′-flanking regions indicated the minimal promoter activity for AtACBP4 (−145/+103) and AtACBP5 (−181/+81). Electrophoretic mobility shift assays identified a pollen-specific cis-acting element POLLEN1 (AGAAA) mapped at AtACBP4 (−157/−153) which interacted with nuclear proteins from flower and this was substantiated by DNase I footprinting. In Arabidopsis thaliana, six acyl-CoA-binding proteins (ACBPs), designated as AtACBP1 to AtACBP6, have been identified to function in plant stress and development. AtACBP4 and AtACBP5 represent the two largest proteins in the AtACBP family. Despite having kelch-motifs and sharing a common cytosolic subcellular localization, AtACBP4 and AtACBP5 differ in spatial and temporal expression. Histological analysis on transgenic Arabidopsis expressing β-glucuronidase driven from the respective AtACBP4 and AtACBP5 promoters, as well as, qRT-PCR analysis revealed that AtACBP4 was expressed at stages 11–14 in mature pollen, while AtACBP5 was expressed at stages 7–10 in the microspores and tapetal cells. Immunoelectron microscopy using AtACBP4- or AtACBP5-specific antibodies further showed that AtACBP4 and AtACBP5 were localized in the cytoplasm. Chemical analysis of bud wax and cutin using gas chromatography-flame ionization detector and GC-mass spectrometry analyses revealed the accumulation of cuticular waxes and cutin monomers in acbp4, acbp5 and acbp4acbp5 buds, in comparison to the wild type. Analysis of inflorescences from acbp4 and acbp5 revealed that there was an increase of AtACBP5 expression in acbp4, and an increase of AtACBP4 expression in acbp5. Deletion analysis of the AtACBP4 and AtACBP5 5′-flanking regions indicated the minimal promoter region for AtACBP4 (−145/+103) and AtACBP5 (−181/+81). Electrophoretic mobility shift assays identified a pollen-specific cis-acting element POLLEN1 (AGAAA) within AtACBP4 (−157/−153) which interacted with nuclear proteins from flower and this was substantiated by DNase I footprinting. These results suggest that AtACBP4 and AtACBP5 both function in floral lipidic metabolism and they may play complementary roles in Arabidopsis microspore-to-pollen development.
      PubDate: 2016-11-08
      DOI: 10.1007/s11103-016-0557-5
  • Integrating QTL mapping and transcriptomics identifies candidate genes
           underlying QTLs associated with soybean tolerance to low-phosphorus stress
    • Authors: Dan Zhang; Hengyou Zhang; Shanshan Chu; Hongyan Li; Yingjun Chi; Daniella Triebwasser-Freese; Haiyan Lv; Deyue Yu
      Abstract: Abstract Soybean is a high phosphorus (P) demand species that is sensitive to low-P stress. Although many quantitative trait loci (QTL) for P efficiency have been identified in soybean, but few of these have been cloned and agriculturally applied mainly due to various limitations on identifying suitable P efficiency candidate genes. Here, we combined QTL mapping, transcriptome profiling, and plant transformation to identify candidate genes underlying QTLs associated with low-P tolerance and response mechanisms to low-P stress in soybean. By performing QTL linkage mapping using 152 recombinant inbred lines (RILs) that were derived from a cross between a P-efficient variety, Nannong 94–156, and P-sensitive Bogao, we identified four major QTLs underlying P efficiency. Within these four QTL regions, 34/81 candidate genes in roots/leaves were identified using comparative transcriptome analysis between two transgressive RILs, low-P tolerant genotype B20 and sensitive B18. A total of 22 phosphatase family genes were up-regulated significantly under low-P condition in B20. Overexpression of an acid phosphatase candidate gene, GmACP2, in soybean hairy roots increased P efficiency by 15.43–24.54 % compared with that in controls. Our results suggest that integrating QTL mapping and transcriptome profiling could be useful for rapidly identifying candidate genes underlying complex traits, and phosphatase-encoding genes, such as GmACP2, play important roles involving in low-P stress tolerance in soybean.
      PubDate: 2016-11-04
      DOI: 10.1007/s11103-016-0552-x
  • Novel rice mutants overexpressing the brassinosteroid catabolic gene
    • Authors: Wenjing Qian; Chao Wu; Yaping Fu; Guocheng Hu; Zhengquan He; Wenzhen Liu
      Abstract: Key message Moderate overexpression of CYP734A4 improves grain number per main panicle and seed setting rate. Brassinosteroid (BR) homeostasis and signaling are crucial for plant growth and development. CYP734A genes encode cytochrome P450 monooxygenases that control the level of bioactive BRs by degrading BRs. However, fertile plants overexpressing CYP734As have not been reported in rice. Here, we isolated a novel semi-dominant mutant brd3-D, in which T-DNA was inserted approximately 4 kb upstream of the CYP734A4 gene (GenBank Accession AB488667), causing its overexpression. The mutant is characterized by dwarfism, small grains, and erect leaves and is less sensitive to brassinolide-induced lamina joint inclination and primary root elongation. However, increased grain number per main panicle and improved seed setting rate were also found in heterozygous brd3-D. To our knowledge, these traits have not been reported in other BR deficient mutants. Quantitative real-time PCR analysis indicated that phenotypic severity of the brd3-D mutant is positively correlated with the CYP734A4 transcription level. In accordance with the increased expression of CYP734A4, a lower castasterone (a rice BR) content was detected in the brd3-D mutants. Knockout of brd3-D by using the CRISPR/Cas9 system rescued the mutation. In addition, transgenic plants overexpressing CYP734A4 with the 35S enhancer mimicked the brd3-D phenotypes, confirming that moderate overexpression of the CYP734A4 gene can improve grain number per main panicle and the seed setting rate in rice. Further studies showed that overexpression of CYP734A4 influences the expressions of multiple genes involved in the BR pathway, and the expression of CYP734A4 is induced by exogenous brassinolide, confirming the negative regulatory role of CYP734A4 in the BR pathway. CYP734A4 might provide a useful gene resource for developing new high-yielding rice varieties.
      PubDate: 2016-11-04
      DOI: 10.1007/s11103-016-0558-4
  • The protein phosphatase 2A catalytic subunit StPP2Ac2b acts as a positive
           regulator of tuberization induction in Solanum tuberosum L.
    • Authors: María Noelia Muñiz García; María Catalina Muro; Luciana Carla Mazzocchi; Silvia Marina País; Margarita Stritzler; Mariana Schlesinger; Daniela Andrea Capiati
      Abstract: Key message This study provides the first genetic evidence for the role of PP2A in tuberization, demonstrating that the catalytic subunit StPP2Ac2b positively modulates tuber induction, and that its function is related to the regulation of gibberellic acid metabolism. The results contribute to a better understanding of the molecular mechanism controlling tuberization induction, which remains largely unknown. The serine/threonine protein phosphatases type 2A (PP2A) are implicated in several physiological processes in plants, playing important roles in hormone responses. In cultivated potato (Solanum tuberosum), six PP2A catalytic subunits (StPP2Ac) were identified. The PP2Ac of the subfamily I (StPP2Ac1, 2a and 2b) were suggested to be involved in the tuberization signaling in leaves, where the environmental and hormonal signals are perceived and integrated. The aim of this study was to investigate the role of PP2A in the tuberization induction in stolons. We selected one of the catalytic subunits of the subfamily I, StPP2Ac2b, to develop transgenic plants overexpressing this gene (StPP2Ac2b-OE). Stolons from StPP2Ac2b-OE plants show higher tuber induction rates in vitro, as compared to wild type stolons, with no differences in the number of tubers obtained at the end of the process. This effect is accompanied by higher expression levels of the gibberellic acid (GA) catabolic enzyme StGA2ox1. GA up-regulates StPP2Ac2b expression in stolons, possibly as part of the feedback system by which the hormone regulates its own level. Sucrose, a tuber-promoting factor in vitro, increases StPP2Ac2b expression. We conclude that StPP2Ac2b acts in stolons as a positive regulator tuber induction, integrating different tuberization-related signals mainly though the modulation of GA metabolism.
      PubDate: 2016-11-03
      DOI: 10.1007/s11103-016-0555-7
  • A Chinese cabbage ( Brassica campetris subsp. Chinensis) τ-type
           glutathione- S -transferase stimulates Arabidopsis development and primes
           against abiotic and biotic stress
    • Authors: Chih-Wei Kao; Madhunita Bakshi; Irena Sherameti; Sheqin Dong; Michael Reichelt; Ralf Oelmüller; Kai-Wun Yeh
      Abstract: Abstract The beneficial root-colonizing fungus Piriformospora indica stimulates root development of Chinese cabbage (Brassica campestris subsp. Chinensis) and this is accompanied by the up-regulation of a τ-class glutathione (GSH)-S-transferase gene (BcGSTU) (Lee et al. 2011) in the roots. BcGSTU expression is further promoted by osmotic (salt and PEG) and heat stress. Ectopic expression of BcGSTU in Arabidopsis under the control of the 35S promoter results in the promotion of root and shoot growth as well as better performance of the plants under abiotic (150 mM NaCl, PEG, 42 °C) and biotic (Alternaria brassicae infection) stresses. Higher levels of glutathione, auxin and stress-related (salicylic and jasmonic acid) phytohormones as well as changes in the gene expression profile result in better performance of the BcGSTU expressors upon exposure to stress. Simultaneously the plants are primed against upcoming stresses. We propose that BcGSTU is a target of P. indica in Chinese cabbage roots because the enzyme participates in balancing growth and stress responses, depending on the equilibrium of the symbiotic interaction. A comparable function of BcGST in transgenic Arabidopsis makes the enzyme a valuable tool for agricultural applications.
      PubDate: 2016-10-31
      DOI: 10.1007/s11103-016-0531-2
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