Project description:The effects of low and high light intensities on transcriptome of purple non sulfur bacterium R. capsulatus were investigated by comparing expression profiles of dark and low light intensity (2000 lux), and by comparing high light intensity (10,000 lux) with low light intensity (2000 lux).
Project description:This study aim to understand how the long and short day flowering pathways are integrated and the mechanism of photoperiod perception in rice. Trascriptome at different time points under LD and SD conditions reveal that photoperiodism in rice is controlled by the evening complex. Mutants in LUX ARRYTHMO (LUX) and EARLY FLOWERING3 (ELF3) orthologs abolish flowering. We show that light causes a rapid and sustained degradation of ELF3-1, and this response is dependent on phyB. ChIP-seq of ELF3 and LUX reveal that EC controls both LD and SD flowering pathways by directly binding and suppressing the expression of key floral repressors, including PRR7 orthologs and Ghd7.
Project description:Purpose: The goals of this study are to compare RNA-seq profiles of Col-0, acd6-1, Iux-1, and acd6-1lux-1 to identify genes affected by LUX. Methods: Total RNA was extracted from 25-d old Col-0, lux-1, acd6-1, or acd6-1lux-1 plants collected at ZT1 or ZT13. Triplicate biological samples were used for most genotypes at each time point, except acd6-1 and acd6-1lux-1 at ZT13, which had duplicate samples. 0.5 ug RNA per replicate was used to generate cDNA libraries using Illumina TruSeq RNA sample preparation kit (catalog no. RS-122-2001). The samples were multiplexed and sequenced using the Illumina HiSeq sequencing platform in Genomics Resources Core Facility at Weill Cornell Medical College. Sequencing was conducted with a standard run of 51 cycles and single reads. At least 150 million reads per lane were obtained for sequencing. qRT–PCR validation was performed using TaqMan and SYBR Green assays for some selected genes. Results: We found that over 1500 genes are affected by lux-1 based on the RNAseq analysis. GO analysis revealed that LUX-affected genes are enchriched in response to abiotic and biotic stimuli. In particular, genes involved in basal defense, salicyclic acid signaling, and jasmonic acid signaling are affected by lux-1. We also found LUX regulation of the clock genes, including core clock components and those acting in the output pathways. Conclusions: The RNAseq analysis support a role of LUX in regulation of the circadian clock and plant defense.
Project description:To characterize the largely unknown functions of oxidised methylcytosines (oxi-mC; 5hmC/5fC/5caC) in DNA, several sequencing protocols have been recently developed. Quantitative analysis is complicated because DNA methylation modifications need to be deconvoluted from the data which is affected by several experimental parameters, including e.g. imperfect bisulphite conversion, oxidation efficiencies, various chemical labeling and protection steps, and sequencing errors. Here, we present a hierarchical generative model, Lux, for integrative analysis of any combination of BS-seq and âoxi-mCâ-seq (BS-seq/oxBS-seq/TAB-seq/fCAB-seq/CAB-seq/redBS-seq/MAB-seq) data. We show that Lux improves quantification and comparison of methylation levels over existing methods and that Lux can easily process any oxi-mC-seq data sets to quantify all cytosine modifications simultaneously together with their experimental parameters. Application of Lux to targeted data from Tet2 knockdown ESCs and T cells during development demonstrates DNA modification quantification at unprecedented detail, quantifies active demethylation pathway and reveals 5hmC localization in putative regulatory regions. Examine the distribution of C, 5mC, and 5hmC in ES and Tet2 knock-down cells within selected loci. Half of the samples are measured using the traditional bisulphite-seq protocol but the other half is measured using the oxidative bisulphite-seq protocol (oxBS-seq).
Project description:To characterize the largely unknown functions of oxidised methylcytosines (oxi-mC; 5hmC/5fC/5caC) in DNA, several sequencing protocols have been recently developed. Quantitative analysis is complicated because DNA methylation modifications need to be deconvoluted from the data which is affected by several experimental parameters, including e.g. imperfect bisulphite conversion, oxidation efficiencies, various chemical labeling and protection steps, and sequencing errors. Here, we present a hierarchical generative model, Lux, for integrative analysis of any combination of BS-seq and âoxi-mCâ-seq (BS-seq/oxBS-seq/TAB-seq/fCAB-seq/CAB-seq/redBS-seq/MAB-seq) data. We show that Lux improves quantification and comparison of methylation levels over existing methods and that Lux can easily process any oxi-mC-seq data sets to quantify all cytosine modifications simultaneously together with their experimental parameters. Application of Lux to targeted data from Tet2 knockdown ESCs and T cells during development demonstrates DNA modification quantification at unprecedented detail, quantifies active demethylation pathway and reveals 5hmC localization in putative regulatory regions. Examine the distribution of C, 5mC, and 5hmC in DP, CD4 SP, and naïve CD4 T cells within selected loci. Half of the samples are measured using the traditional bisulphite-seq protocol but the other half is measured using the oxidative bisulphite-seq protocol (oxBS-seq).
Project description:This study aim to understand how the long and short day flowering pathways are integrated and the mechanism of photoperiod perception in rice. Trascriptome at different time points under LD and SD conditions reveal that photoperiodism in rice is controlled by the evening complex. Mutants in LUX ARRYTHMO (LUX) and EARLY FLOWERING3 (ELF3) orthologs abolish flowering. We show that light causes a rapid and sustained degradation of ELF3-1, and this response is dependent on phyB. ChIP-seq of ELF3 and LUX reveal that EC controls both LD and SD flowering pathways by directly binding and suppressing the expression of key floral repressors, including PRR7 orthologs and Ghd7.
Project description:Delineating the extracellular interaction network of the cell surface proteotype (the surfaceome) is fundamental to understanding cellular signaling function in health and disease. Here, we developed LUX-MS, an optoproteomic technology that enables the spatiotemporal and proteome-wide study of surfaceome signaling architectures at nanoscale level without the need for genetic manipulation. A tunable, light-triggered singlet oxygen generator (SOG) based mechanism mediates in-situ proximity-tagging for subsequent mass spectrometry-based identification of acute protein interactions in their native cellular context. We applied LUX-MS to the characterization of surfaceome signaling structures engaged by antibodies, small molecule drugs, biologics and intact bacteriophages across organisms and in complex environments. Cell-type resolved dissection of intercellular communication using LUX-MS thereby revealed the molecular architecture of functional immunological signaling synapses in unprecedented detail. Altogether, LUX-MS enables facebooking of the social protein networks within surfaceome signaling architectures and provides an unprecedented molecular framework for the rational design of biomedical intervention strategies.
Project description:To characterize the largely unknown functions of oxidised methylcytosines (oxi-mC; 5hmC/5fC/5caC) in DNA, several sequencing protocols have been recently developed. Quantitative analysis is complicated because DNA methylation modifications need to be deconvoluted from the data which is affected by several experimental parameters, including e.g. imperfect bisulphite conversion, oxidation efficiencies, various chemical labeling and protection steps, and sequencing errors. Here, we present a hierarchical generative model, Lux, for integrative analysis of any combination of BS-seq and “oxi-mC”-seq (BS-seq/oxBS-seq/TAB-seq/fCAB-seq/CAB-seq/redBS-seq/MAB-seq) data. We show that Lux improves quantification and comparison of methylation levels over existing methods and that Lux can easily process any oxi-mC-seq data sets to quantify all cytosine modifications simultaneously together with their experimental parameters. Application of Lux to targeted data from Tet2 knockdown ESCs and T cells during development demonstrates DNA modification quantification at unprecedented detail, quantifies active demethylation pathway and reveals 5hmC localization in putative regulatory regions.
Project description:To characterize the largely unknown functions of oxidised methylcytosines (oxi-mC; 5hmC/5fC/5caC) in DNA, several sequencing protocols have been recently developed. Quantitative analysis is complicated because DNA methylation modifications need to be deconvoluted from the data which is affected by several experimental parameters, including e.g. imperfect bisulphite conversion, oxidation efficiencies, various chemical labeling and protection steps, and sequencing errors. Here, we present a hierarchical generative model, Lux, for integrative analysis of any combination of BS-seq and “oxi-mC”-seq (BS-seq/oxBS-seq/TAB-seq/fCAB-seq/CAB-seq/redBS-seq/MAB-seq) data. We show that Lux improves quantification and comparison of methylation levels over existing methods and that Lux can easily process any oxi-mC-seq data sets to quantify all cytosine modifications simultaneously together with their experimental parameters. Application of Lux to targeted data from Tet2 knockdown ESCs and T cells during development demonstrates DNA modification quantification at unprecedented detail, quantifies active demethylation pathway and reveals 5hmC localization in putative regulatory regions.