Project description:LHY and CCA1 encode single MYB transcription factors, involved in circadian clock. However, direct target genes of LHY and CCA1 in a genomic scale were largely unknown. To reveal bound genes by CCA1, chimeric protein CCA1-FLAG was expressed under CCA1 promoter in cca1 lhy (CCA1pro:CCA1-FLAG/ cca1 lhy). ChIP was performed using anti-FLAG antibody (F3165; SIGMA), which was bound to Dynabeads Protein G (100-03D; Life Technologies), and ChIP DNA were analyzed by IonPGM or Illumina GAII. Chromatin immunoprecipitation was performed for CCA1-FLAG-expressing Arabidopsis. ChIP DNA was analyzed 2 types of deep sequencers (Illumina GAII and IonPGM).
Project description:LHY and CCA1 encode single MYB transcription factors, involved in circadian clock. However, direct target genes of LHY and CCA1 in a genomic scale were largely unknown. To reveal bound genes by CCA1, chimeric protein CCA1-FLAG was expressed under CCA1 promoter in cca1 lhy (CCA1pro:CCA1-FLAG/ cca1 lhy). ChIP was performed using anti-FLAG antibody (F3165; SIGMA), which was bound to Dynabeads Protein G (100-03D; Life Technologies), and ChIP DNA were analyzed by IonPGM or Illumina GAII.
Project description:The plant circadian clock exerts a critical role in the regulation of multiple biological processes including responses to biotic and abiotic stresses. It is estimated that the clock regulates up to 80% of the transcriptome in Arabidopsis, thus understanding the molecular mechanisms that control this rhythmic transcriptome requires identification of the targets of each clock component. The Arabidopsis core clock is partially comprised of a transcriptional regulatory loop between the MYB domain containing transcription factors CIRCADIAN CLOCK ASSOCIATED1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY), and TIMING OF CAB EXPRESSION1 (TOC1). As a key component of the clock, CCA1 is able to initiate and set the phase of clock-controlled rhythms. CCA1 regulates the transcription of several genes by directly binding to the evening element (EE) motif primarily found in the promoters of evening expressed genes. Using a genome-wide approach we have identified direct targets of CCA1 in plants grown in constant (LL) and driven conditions (LD). These CCA1 targets are enriched for a myriad of biological processes and stress responses. While many of these target genes are evening phased and contain the EE in their promoter regions, a significant subset is morning phased and lack an EE. Furthermore, several CCA1 targets do not cycle in either LL or LD or both. Expression analysis in CCA1 overexpressing plants confirms CCA1 regulation of analyzed targets. Our results emphasize an expanded role for the circadian clock in regulation of key pathways in Arabidopsis, and provide a comprehensive and solid resource for future functional studies. ChIP-Seq of CCA1-GFP plants under control of the CCA1 promoter in continuous light and diel conditions
Project description:The plant circadian clock exerts a critical role in the regulation of multiple biological processes including responses to biotic and abiotic stresses. It is estimated that the clock regulates up to 80% of the transcriptome in Arabidopsis, thus understanding the molecular mechanisms that control this rhythmic transcriptome requires identification of the targets of each clock component. The Arabidopsis core clock is partially comprised of a transcriptional regulatory loop between the MYB domain containing transcription factors CIRCADIAN CLOCK ASSOCIATED1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY), and TIMING OF CAB EXPRESSION1 (TOC1). As a key component of the clock, CCA1 is able to initiate and set the phase of clock-controlled rhythms. CCA1 regulates the transcription of several genes by directly binding to the evening element (EE) motif primarily found in the promoters of evening expressed genes. Using a genome-wide approach we have identified direct targets of CCA1 in plants grown in constant (LL) and driven conditions (LD). These CCA1 targets are enriched for a myriad of biological processes and stress responses. While many of these target genes are evening phased and contain the EE in their promoter regions, a significant subset is morning phased and lack an EE. Furthermore, several CCA1 targets do not cycle in either LL or LD or both. Expression analysis in CCA1 overexpressing plants confirms CCA1 regulation of analyzed targets. Our results emphasize an expanded role for the circadian clock in regulation of key pathways in Arabidopsis, and provide a comprehensive and solid resource for future functional studies.
Project description:Genomic integrity requires faithful chromosome duplication. Origins of replication are the genomic sites where DNA replication initiates in every cell cycle. There are multiple origins scattered throughout the eukaryotic genome whose genome-wide identification has been a hard challenge, especially in multicellular organisms. Thus, very little is known on the distinctive features of origins in terms of DNA sequence and chromatin context at a genomic scale. As part of a project for profiling replication origins in Arabidopsis thaliana, we have performed ChIP-chip analysis of the binding of ORC1 and CDC6, two proteins involved in initiation of DNA replication. Here, we provide the data of the ORC1-bound and CDC6-bound genomic sites using as control genomic DNA.
Project description:FLP and MYB88 are two paralogous MYB proteins, regulating the symmetric division of guard mother cell during Arabidopsis stomatal development. To understand their molecular functions, we performed genome-wide identification of FLP/MYB88 binding targets using ChIP-chip with FLP/MYB88 antibody. By comparing ChIP-chip between wild-type and flp-1 myb88 lines, a total genes were identified as putative direct targets for FLP/MYB88.