Project description:Ovule development is a key process for plant reproduction that ensures correct seed production. Understanding the molecular mechanisms that control ovule formation will also provide new approaches to increase crop yield for breeding. Several molecular factors and plant hormones, including gibberellins, are involved in ovule initiation and development. Gibberellins control ovule development by the destabilization of DELLA proteins, whereas DELLA activity has been proved to act as a positive factor for ovule primordia emergence. But the molecular mechanism by which DELLA act remained unknown. Here we have proved that DELLA proteins control ovule initiation by the formation of a protein complex with the CUC2 transcription factor. The DELLA protein GAI requires CUC2 to promote ovule primordia formation, thus GAI would function by its direct protein-protein interaction with CUC2 in cells of the placenta that determine the boundary regions between ovules during pistil development. Analysis of GAI-CUC2 interaction and colocalization in placenta support this hypothesis. Moreover, molecular analysis of the loci at which GAI protein may act as transcriptional co-regulators in a CUC2-dependent manner identified a subset of target genes that would be regulated by the GAI-CUC2 complex and contribute to regulate ovule primordia emergence.
Project description:The hormones gibberellins (GA) control many aspects of plant growth and development thruough the whole life cycle of the plant. For instance, GA participate in the establishment of the skotomorphogenic developmental program that is triggered when seeds germinate in darkness, i.e. under the soil. Under these conditions seedlings appear etiolated and developmental features usually triggered by light are kept repressed. The GA signaling elements GAI and RGA have a major, partially redundant role in this process. These proteins belong to the DELLA family of transcriptional regulators and are destabilized by GA, acting as negative regulators of the pathway. In order to understand the molecular basis of the regulation of the skotomorphogenesis by GA, we sought to identify early target genes of the activity of GAI in etiolated seedlings. For that purpose we analyzed rapid, global changes in gene expression in response to a transient accummulation of a dominant version of GAI, gai-1, which is resistant to GA-induced destabilization.
Project description:The hormones gibberellins (GA) control many aspects of plant growth and development thruough the whole life cycle of the plant. For instance, GA participate in the establishment of the skotomorphogenic developmental program that is triggered when seeds germinate in darkness, i.e. under the soil. Under these conditions seedlings appear etiolated and developmental features usually triggered by light are kept repressed. The GA signaling elements GAI and RGA have a major, partially redundant role in this process. These proteins belong to the DELLA family of transcriptional regulators and are destabilized by GA, acting as negative regulators of the pathway. In order to understand the molecular basis of the regulation of the skotomorphogenesis by GA, we sought to identify early target genes of the activity of GAI in etiolated seedlings. For that purpose we analyzed rapid, global changes in gene expression in response to a transient accummulation of a dominant version of GAI, gai-1, which is resistant to GA-induced destabilization. Experiments were performed using wild type Col-0 and the transgenic line HS:gai-1 that expresses the dominant version gai-1 under the control of the promoter of the HSP18.2 gene, which is highly and rapidly induced at 37ºC. Three biological repeats were performed, and wild type and transgenic samples were labelled with Cy3 and Cy5, respectively. For each point in the time course (0, 1, 2 and 4h after a 30 minute treatment at 37ºC), a sample from the transgenic line was compared to the corresponding wild type control.
Project description:Macrophages have a pivotal role during viral infections in pigs. By expressing cell surface receptors, macrophages become viral targets and reservoirs. In the case of upper respiratory tract infections, many viruses target the peripheral nasal mucosa. Recent in vivo and in vitro (primary nasal cell cultures) porcine reproductive and respiratory syndrome virus (PRRSV) studies demonstrated that several macrophage subsets exist in the porcine nasal mucosa, and that virus strains with different virulence showed altered tropism for these different macrophage populations. To further investigate these macrophage subsets, total RNA sequencing was performed in parallel on two subsets from the nasal mucosa and lung macrophages. Macrophages were isolated by either fluorescent activated cell sorting (FACS; cf. bulk RNAseq) or laser capture microdissection (LCM; cf. LCM RNAseq) in combination with immunofluorescence staining against two macrophage markers, CD163 and Sialoadhesin (Sn). With both RNAseq methods, nasal macrophages showed a different transcriptomic profile compared to lung macrophages. Differentially expressed genes were identified in the two subsets of nasal macrophages. Gene set enrichment analysis on the three macrophage populations showed that GO terms and KEGG pathways on LCM RNAseq data were more specific to the spatial location of macrophage subsets than bulk RNAseq data. Cell type signature analysis revealed that nasal CD163+Sn- cells resemble squamous epithelial cells (LCM RNAseq) or antigen presenting cells (bulk RNAseq) while nasal CD163+Sn+ cells are more like fibroblasts/stromal cells (LCM RNAseq) or vascular endothelial cell (bulk RNAseq). Our results confirmed that not only macrophages in different tissues but also macrophages in different areas within the same tissue have different transcriptional programs, suggesting their differential roles in their interaction with pathogens and corresponding immune responses.
Project description:RNAseq was used to identify host and EBV viral transcriptome changes in CHAF1B knock-out Akata EBV+ cells. CHAF1B KO Akata EBV+ cells were subjected to RNAseq analysis. The Akata EBV+ cells expressing control sgRNA was used as the control.