ABSTRACT: We studied differences in gene expression between Populus P35S::EBB1 lines and control, affecting plant growth and differentiation, and dormancy. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes driven by overexpression of the EBB1 gene. By screening activation tagging population, we identified the EARLY BUD-BREAK1 (EBB1) mutant. We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes an AP2/ERF transcription factor similar to a small B1 gene-subfamily in Arabidopsis encoding strong regulators of meristem function and lateral organ outgrowth. We fully recapitulated the phenotype by overexpression of the gene into the same genotype under a strong constitutive promoter (P35S::EBB1). We also found that EBB1 transcript abundance in wild type plants increases during the period prior to bud-break, and in response to the hormone cytokinin. Our data indicates that EBB1 plays an important role in the process of bud-break. Poplar apex was selected for RNA extraction and hybridization on Affymetrix microarrays. We sought to obtain homogeneous native expression of affected genes in P35S::EBB1 lines and control, in order to increase the resolution of expression profiles inducing the developmental changes in P35S::EBB1. To do that, we selected apex tissue from greenhouse healthy plants.
Project description:We study gene expression Populus amiEBB1 lines affecting dormancy. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes droved by expression of artifical micro RNA (ami) targeting EBB1 gene. By screening activation tagging population we identified the EARLY BUD-BREAK1 (EBB1) mutant. We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes an AP2/ERF transcription factor similar to a small B1 gene-subfamily in Arabidopsis encoding strong regulators of meristem function and lateral organ outgrowth. We fully recapitulated the phenotype by overexpression of the gene into the same genotype under strong constitutive promoter (P35S::EBB1). We also found that EBB1 transcript abundance in wild type plants increases during the period prior to bud-break, and in response to the hormone cytokinin. Down-regualtion of EBB1 influence negatively bud break. Our data indicates that EBB1 plays a important role in the process of bud-break. Poplar apex was selected for RNA extraction and hybridization on Affymetrix microarrays. We used apices tissue from amiEBB1 grown greenhouse healthy plants.
Project description:We study differences in gene expression between Populus P35S::BL (BL-oe) lines and control, affecting plant growth and differentiation, and dormancy. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes droved by overexpression of BL gene. We identified an activation tagging mutant with increased leaf size and correspondingly name it BIG LEAF (BL). We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes a WD40 putative transcription regulator similar to the Arabidopsis thaliana STERILE APETALA (SAP). We recapitulated the phenotype by overexpression of the gene into the same genotype under strong constitutive promoter (P35S::BL, BL-oe). Transgenic up-regulation of the BL gene caused enhanced leaf size, early bud-break, and suppression of secondary growth. BL transcript abundance in wild type plants is in apical tissues, mostly in shoot meristem, leaf primordia and axillary meristem. Our data indicates that BL plays an important role in the process of tree growth. Poplar apex, secondary stem (30th internode), and leaves (at 30 node) was selected for RNA extraction and hybridization on Affymetrix microarrays. We sought to obtain expression of affected genes in P35S::BL lines and control wild type (WT-717), in order to increase the resolution of expression profiles inducing the developmental changes in P35S::BL. To do that, we selected apex, stem and leaf tissue from greenhouse healthy plants.
Project description:We studied differences in gene expression between Populus P35S::EBB1 lines and control, affecting plant growth and differentiation, and dormancy. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes driven by overexpression of the EBB1 gene. By screening activation tagging population, we identified the EARLY BUD-BREAK1 (EBB1) mutant. We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes an AP2/ERF transcription factor similar to a small B1 gene-subfamily in Arabidopsis encoding strong regulators of meristem function and lateral organ outgrowth. We fully recapitulated the phenotype by overexpression of the gene into the same genotype under a strong constitutive promoter (P35S::EBB1). We also found that EBB1 transcript abundance in wild type plants increases during the period prior to bud-break, and in response to the hormone cytokinin. Our data indicates that EBB1 plays an important role in the process of bud-break.
Project description:We study gene expression Populus amiEBB1 lines affecting dormancy. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes droved by expression of artifical micro RNA (ami) targeting EBB1 gene. By screening activation tagging population we identified the EARLY BUD-BREAK1 (EBB1) mutant. We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes an AP2/ERF transcription factor similar to a small B1 gene-subfamily in Arabidopsis encoding strong regulators of meristem function and lateral organ outgrowth. We fully recapitulated the phenotype by overexpression of the gene into the same genotype under strong constitutive promoter (P35S::EBB1). We also found that EBB1 transcript abundance in wild type plants increases during the period prior to bud-break, and in response to the hormone cytokinin. Down-regualtion of EBB1 influence negatively bud break. Our data indicates that EBB1 plays a important role in the process of bud-break.
Project description:We study differences in gene expression between Populus P35S::BL (BL-oe) lines and control, affecting plant growth and differentiation, and dormancy. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes droved by overexpression of BL gene. We identified an activation tagging mutant with increased leaf size and correspondingly name it BIG LEAF (BL). We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes a WD40 putative transcription regulator similar to the Arabidopsis thaliana STERILE APETALA (SAP). We recapitulated the phenotype by overexpression of the gene into the same genotype under strong constitutive promoter (P35S::BL, BL-oe). Transgenic up-regulation of the BL gene caused enhanced leaf size, early bud-break, and suppression of secondary growth. BL transcript abundance in wild type plants is in apical tissues, mostly in shoot meristem, leaf primordia and axillary meristem. Our data indicates that BL plays an important role in the process of tree growth.
Project description:We study the effect of nitrogen limitation on the growth and development of poplar roots. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes driven by low nitrogen in the growth media. We report the effect of nitrogen limitation on the growth and development of poplar roots. Low nitrogen concentration led to increased root elongation followed by lateral root proliferation and finally increased root biomass. These morphological responses correlated with high and specific activation of genes encoding regulators of cell cycle and enzymes involved in cell wall biogenesis, growth and remodeling. Comparative analysis of poplar and Arabidopsis root transcriptomes under nitrogen deficiency indicated many similarities and diversification in the response in the two species. A reconstruction of genetic regulatory network (GRN) analysis revealed a sub-network centered on a PtaNAC1-like transcription factor. Consistent with the GRN predictions, root-specific upregulation of PtaNAC1 in transgenic poplar plants increased root biomass and led to significant changes in the expression of the connected genes specifically under low nitrogen. PtaNAC1 and its regulatory miR164 showed inverse expression profiles during response to LN, suggesting of a micro RNA mediated attenuation of PtaNAC1 transcript abundance in response to nitrogen deprivation. Poplar roots from low nitrogen treated and untreated from in vitro condition was selected for RNA extraction and hybridization on Affymetrix microarrays. Roots were sampled at 6, 12, 24, 48, 96 and 504h after transfer to control and low nitrogen media and RNA was extacted.
Project description:A grape-bud-oriented genomic platform was produced for a large-scale comparative analysis of bud responses to two stimuli of grape-bud dormancy release, hydrogen cyanamide (HC) and heat shock (HS). The results suggested considerable similarity in bud response to the stimuli, both in the repertoire of responding genes and in the temporary nature of the transcriptome reprogramming. Nevertheless, the bud response to HC was slower, more condensed and stronger, as reflected by a higher number of regulated genes and a higher intensity of regulation compared to the response to HS. To facilitate large-scale comparative analysis of early changes in the bud transcriptome by cDNA microarray, HC and HS were applied to canes collected from three vineyards, located in different regions, in three different years. This experimental scheme resulted in two true biological replicates for each treatment, differing in both timing and location, and loop design of technical replicates within time series. Consistent with our previous studies, both application of 5% Dormex (HC) and incubation for 1 h in 50oC water (HS) resulted in increased bud-break rates compared to respective controls. Bud break of HS-treated and HC-treated buds started 10 to 12 days after treatment. Three weeks after treatment, HS-treated buds exhibited 100% bud break while HC-treated buds had reached 80% bud break. The control showed significantly lower levels of bud break during this period. Bud material was collected from control, HC- and HS-treated cuttings at six time points (3, 6, 12, 24, 48 and 96 h) after the treatments and used to prepare total RNA samples.
Project description:Bud dormancy – the repeated phase of rest that punctuates periods of growth in the life cycles of many perennial species. In temperate fruit trees, fulfillment of chilling requirement and subsequent heat requirement enable dormant buds to have uniform blooming in field. However, effects of environmental factors such as chilling underlying dormancy release and bud break has not been fully understood. Histone modification is an important epigenetic regulation system which plays an important role in gene expression in various developmental and adaptive processes. Taking advantage of next-generation sequencing, we generated epigenome and transcriptome profiling at different stages before chilling, after chilling and just before bud break in apple (Malus x domestica). We found H3K27me3 may play dominant role during chilling and the genes involved in lignin and lipid metabolic process showed histone modifications. Interestingly, the higher ratio of genes in chilling-associated network exhibited histone modifications, suggesting crucial epigenetic roles in regulating gene expressions in response to chilling during dormancy. Furthermore, H3K4me3 may play more important role during bud break and the genes related to cell wall modification/organization were strongly modified. Taken together, this study provides important insights into the chromatin-based gene regulation underlying chilling-mediated dormancy release and bud break in apple.
Project description:Bud dormancy – the repeated phase of rest that punctuates periods of growth in the life cycles of many perennial species. In temperate fruit trees, fulfillment of chilling requirement and subsequent heat requirement enable dormant buds to have uniform blooming in field. However, effects of environmental factors such as chilling underlying dormancy release and bud break has not been fully understood. Histone modification is an important epigenetic regulation system which plays an important role in gene expression in various developmental and adaptive processes. Taking advantage of next-generation sequencing, we generated epigenome and transcriptome profiling at different stages before chilling, after chilling and just before bud break in apple (Malus x domestica). We found H3K27me3 may play dominant role during chilling and the genes involved in lignin and lipid metabolic process showed histone modifications. Interestingly, the higher ratio of genes in chilling-associated network exhibited histone modifications, suggesting crucial epigenetic roles in regulating gene expressions in response to chilling during dormancy. Furthermore, H3K4me3 may play more important role during bud break and the genes related to cell wall modification/organization were strongly modified. Taken together, this study provides important insights into the chromatin-based gene regulation underlying chilling-mediated dormancy release and bud break in apple.
Project description:Abscisic acid (ABA) regulates seed and bud dormancy. We show by forward and reverse genetic analysis that the tomato transcription factor SlZFP2 is required for release of bud and seed dormancy through negative regulation of ABA biosynthesis. We also demonstrated that ABA promotes growth and represses flowering in tomato both through transcriptional control on the florigen-encoding gene SINGLE FLOWER TRUSS (SFT) in tomato. To gain further insight on transcriptome changes by overexpresion of HA-SlZFP2, we sequenced two lines of p35S:HA-SlZFP2 in LA1589 background and their nontransgenic siblings on Illumina Hiseq2000 platform.