Project description:We have investigated the effect of RRP6 depletion on the transcriptome of S2 cells using Affymetrix whole-genome tiling arrays. We have also carried out Illumina ChIP-seq analysis of RRP6 genome occupancy in control S2 cells (GFP-KD) and in cells depleted of SU(VAR)3-9.
Project description:Epigenetic regulation of mutually exclusive transcription within the var gene family is important for infection and pathogenesis of the malaria parasite Plasmodium falciparum. var genes are kept transcriptionally silent via heterochromatic clusters located at the nuclear periphery; however, only a few proteins have been shown to play a direct role in var gene transcriptional regulation. Importantly, the chromatin components that contribute to var gene nuclear organization remain unknown. Here, we adapted a CRISPR-based immunoprecipitation-mass spectrometry approach for de novo identification of factors associated with specific transcriptional regulatory sequences of var genes. Tagged, catalytically inactive Cas9 (“dCas9”) was targeted to var gene promoters or introns, cross-linked, and immunoprecipitated with all DNA, proteins, and RNA associated with the targeted locus. Chromatin immunoprecipitation followed by sequencing demonstrated that genome-wide dCas9 binding was specific and robust. Proteomics analysis of dCas9-immunoprecipitates identified specific proteins for each target region, including known and novel factors such as DNA binding proteins, chromatin remodelers, and structural proteins. We also demonstrate the ability to immunoprecipitate RNA that is closely associated to the targeted locus. Our CRISPR/dCas9 study establishes a new tool for targeted purification of specific genomic loci and advances understanding of virulence gene regulation in the human malaria parasite.
Project description:We have investigated the effect of RRP6 depletion on the transcriptome of S2 cells using Illumina deep RNA sequencing. We have also carried out Illumina ChIP-seq analysis of RRP6 genome occupancy in control S2 cells (GFP-KD) and in cells depleted of SU(VAR)3-9.
Project description:The intent was to study, from transcriptome analysis, shade and drought responses in Solanum tuberosum (potato). We performed Illumina 50 bp single-end RNA-seq in tissues of control and treated var. Spunta wild-type plants. Drought experiments also included two independent AtBBX21-overexpressing (BBX21-OE) potato lines.
Project description:Ananas comosus var. bracteatus has high ornamental value and widespread application because of its chimeric leaves. However, little is known about the molecular mechanism regulating this characteristic. Here, comparative transcriptomic and proteomic analyses of the white parts (Whs) and green parts (Grs) of the chimeric leaves were performed to identify differentially expressed genes (DEGs) and differentially expressed proteins (DEPs). In total, 1,685 DEGs, including 712 up- and 973 down-regulated ones, and 5,428 DEPs, including 1,018 up- and 795 down-regulated ones, were identified between the Whs and Grs. Comparisons with the GO and KEGG annotations revealed that the DEGs were involved mostly in carbon fixation, porphyrin and chlorophyll metabolism and oxidative phosphorylation. The DEPs were mainly involved in ribosomes, photosynthesis, photosynthesis antennas, and porphyrin and chlorophyll metabolism. Combined analysis showed that nine proteins related to chlorophyll biosynthesis, photosynthetic pigments, and photosynthesis were unchanged at mRNA level but suppressed at protein level. These results indicated that the albino phenotype of the Whs was caused by the proteomic-level suppression of key enzymes involved in the chlorophyll biosynthesis pathway and that translational and post-translational regulation may play important roles in both the biosynthesis of photosynthetic pigments and photosynthesis. Biological significance: Leaves of Ananas comosus var. bracteatus serve as the best materials for the study of albino mechanism. Because the chemic trait of A. comosus var. bracteatus is unstable and the molecular mechanism of the albino cells was poorly understood, we performed comparative analyses both at the transcriptome and proteome levels. This work revealed suppressed proteomic-level and translational and post-translational regulation contribute to the albino phenotype formation. Our results provide better information concerning the molecular mechanism within the chimeric leaves of A. comosus var. bracteatus.
Project description:We performed Hi-C assay to address the interaction of central var genes in ruf6-var pairs，subtelomeric upsA-subtype var genes and other HP1-associated genes
Project description:H3K9 methylation is a mark of inactive chromatin. In D.melanogaster, three enzymes (SU(VAR)3-9, EGG, and dG9A) are responsible for creating the H3K9 methyl mark. We have investigated the effects of loss of SU(VAR)3-9 and EGG on gene expression in adult heads using microarray analysis. Keywords: Expression profiling by microarray
Project description:To better understand the molecular bases of resin production, a major source of terpenes for industry, the transcriptome of adult Pinus elliottii var. elliottii (slash pine) trees under field commercial resinosis was obtained.
Project description:Histone modifications represent one of the key factors contributing to proper genome regulation. One of the histone modifications involved in gene silencing is H3K9 methylation, which is found in the chromosomes across different eukaryotes and controlled by SU(VAR)3-9 and its orthologs. Although SU(VAR)3-9 was discovered over two decades ago, little is known about the details of its chromosomal distribution pattern. To fill in this gap, we used DamID-seq approach and obtained high-resolution genome-wide profiles for SU(VAR)3-9 in two somatic and two germline tissues of fruitfly.
Project description:The major virulence factor of Plasmodium falciparum parasites, PfEMP1 is expressed by a multigene family, termed var genes. Here selection linked integration (SLI) was utilized to modify var genes in P. falciparum parasites to select for parasite populations expressing a single var gene. Bulk RNA was isolated from ring stage parasites of these SLI parasite populations and analyzed with next generation sequencing. The proportion of exon 2 transcripts of var genes normalized to transcripts per million was determined per cell line to confirm the predominant expression of the desired var gene.
