Project description:Plants are sessile organisms that have acquired highly plastic developmental strategies to adapt to the environment. Among these processes, the floral transition is essential to ensure reproductive success and is finely regulated by several internal and external genetic networks. The photoperiodic pathway, which controls the plant response to day length, is one of the most important pathways controlling flowering. In Arabidopsis photoperiodic flowering, CONSTANS (CO) is the central gene activating the expression of the florigen FLOWERING LOCUS T (FT) in the leaves at the end of a long day. CO expression is strongly regulated by the circadian clock. However, to date, no evidence has been reported regarding a feedback loop from the photoperiod pathway back to the circadian clock. Using transcriptional networks, we have identified relevant network motifs regulating the interplay between the circadian clock and the photoperiod pathway. Gene expression, chromatin immunoprecipitation experiments and phenotypic analysis allowed us to elucidate the role of CO over the circadian clock. Plants with altered CO expression showed a different internal clock period, measured by daily rhythmic movements in the leaves. We show that CO is able to activate key genes related to the circadian clock, such as CCA1, LHY, PRR5 and GI, at the end of a long day by binding to specific sites on their promoters. Moreover, a significant number of PRR5 repressed target genes are upregulated by CO, and this could explain the phase transition promoted by CO. The CO-PRR5 complex interacts with the bZIP transcription factor HY5 and helps to localize the complex in the promoters of clock genes. Our results indicate that there may be a feedback loop in which CO communicates back to the circadian clock, feeding seasonal information to the circadian system.
Project description:Plants are sessile organisms that have acquired highly plastic developmental strategies to adapt to the environment. Among these processes, the floral transition is essential to ensure reproductive success and is finely regulated by several internal and external genetic networks. The photoperiodic pathway, which controls the plant response to day length, is one of the most important pathways controlling flowering. In Arabidopsis photoperiodic flowering, CONSTANS (CO) is the central gene activating the expression of the florigen FLOWERING LOCUS T (FT) in the leaves at the end of a long day. CO expression is strongly regulated by the circadian clock. However, to date, no evidence has been reported regarding a feedback loop from the photoperiod pathway back to the circadian clock. Using transcriptional networks, we have identified relevant network motifs regulating the interplay between the circadian clock and the photoperiod pathway. Gene expression, chromatin immunoprecipitation experiments and phenotypic analysis allowed us to elucidate the role of CO over the circadian clock. Plants with altered CO expression showed a different internal clock period, measured by daily rhythmic movements in the leaves. We show that CO is able to activate key genes related to the circadian clock, such as CCA1, LHY, PRR5 and GI, at the end of a long day by binding to specific sites on their promoters. Moreover, a significant number of PRR5 repressed target genes are upregulated by CO, and this could explain the phase transition promoted by CO. The CO-PRR5 complex interacts with the bZIP transcription factor HY5 and helps to localize the complex in the promoters of clock genes. Our results indicate that there may be a feedback loop in which CO communicates back to the circadian clock, feeding seasonal information to the circadian system.
Project description:The circadian clock represents a critical regulatory network, which allows plants to anticipate environmental changes as inputs and promote plant survival by regulating various physiological outputs. Here, we examine the function of the clock-regulated transcription factor, CYCLING DOF FACTOR 6 (CDF6), during cold stress in Arabidopsis thaliana. We found that the clock gates CDF6 transcript accumulation in the vasculature during cold stress. CDF6 mis-expression results in an altered flowering phenotype during both ambient and cold stress. A genome-wide transcriptome analysis links CDF6 to genes associated with flowering and seed germination during cold and ambient temperatures, respectively. Analysis of key floral regulators indicates that CDF6 alters flowering during cold stress by repressing photoperiodic flowering components, FLOWERING LOCUS T (FT), CONSTANS (CO), and BROTHER OF FT (BFT). Gene ontology enrichment further suggests that CDF6 regulates circadian and developmental associated genes. These results provide insight into how the clock-controlled CDF6 modulates plant development during moderate cold stress.
Project description:The translation of mRNA into protein is tightly regulated by the light environment as well as by the circadian clock. Although changes in translational efficiency have been well documented at the level of mRNA-ribosome loading, the underlying mechanisms are unclear. The reversible phosphorylation of RIBOSOMAL PROTEIN OF THE SMALL SUBUNIT 6 (RPS6) has been known for forty years, but the biochemical significance of this event remains unclear to this day. Here, we confirm using a clock-deficient strain of Arabidopsis thaliana that RPS6 phosphorylation (RPS6-P) is controlled by the diel light-dark cycle with a peak during the day.
Project description:The epidermis is as a highly regenerative barrier protecting organisms from environmental insults, including ultraviolet radiation, the main cause of skin cancer and skin aging. Here we show that time-restricted feeding (RF) shifts the phase and alters the amplitude of the skin circadian clock and affects the expression of approximately 10% of the skin transcriptome. Furthermore, a strikingly large number of skin-expressed genes are acutely regulated by food intake. While the circadian clock is required for daily rhythms in DNA synthesis in epidermal stem cells, RF-induced shifts in clock phase do not alter the phase of DNA synthesis. However, both the expression of the key DNA repair gene Xpa, and the diurnal sensitivity to UVB-induced DNA damage, are altered by RF. Together our findings indicate an unexpected regulation of skin function by time of feeding and emphasize the important link between circadian rhythm, food intake, and skin health.
Project description:The circadian clock is comprised of proteins that form negative feedback loops, which regulate the timing of global gene expression in a coordinated 24 hour cycle. As a result, the plant circadian clock is responsible for regulating numerous physiological processes central to growth and survival. To date, most plant circadian clock studies have relied on diurnal transcriptome changes to elucidate molecular connections between the circadian clock and observable phenotypes in wild-type plants. Here, we have combined high-throughput RNA-sequencing and mass spectrometry to comparatively characterize the lhycca1, prr7prr9, gi and toc1 circadian clock mutant rosette transcriptome and proteome at the end-of-day and end-of-night.
Project description:Study on differential gene expression and splicing between wildtype and clock mutants. This study is part of a comparative analysis of the role of Protein Methyltransferase 5 in the regulation of transcriptional and post-transcriptional processes simultaneously in Arabidopsis and Drosophila. Circadian rhythms allow organisms to time biological processes to the most appropriate phases of the day/night cycle1. Post-transcriptional regulation is emerging as an important component of circadian networks2-6, but the molecular mechanisms linking the circadian clock to the control of RNA processing are largely unknown. Here we show that Protein Arginine Methyl Transferase 5 (PRMT5), which transfers methyl groups to arginine residues present in histones7 and Sm spliceosomal proteins8,9, links the circadian clock to the control of alternative splicing in plants. Mutations in prmt5impair multiple circadian rhythms in Arabidopsis thaliana and this phenotype is caused, at least in part, by a strong alteration in alternative splicing of the core-clock gene PSEUDO RESPONSE REGULATOR 9 (PRR9). Furthermore, genome wide studies show that PRMT5 contributes to regulate many pre-mRNA splicing events most likely modulating 5´splice site (5´ss) recognition. PRMT5 expression shows daily and circadian oscillations, and this contributes to mediate the circadian regulation of expression and alternative splicing of a subset of genes. Circadian rhythms in locomotor activity are also disrupted in dart5, a mutant affected in the Drosophila melanogaster PRMT5 homolog, and this is associated with alterations in splicing of the core-clock gene period (per) and several clock associated genes. Our results reveal a key role for PRMT5 in the regulation of alternative splicing and indicate that the interplay between the circadian clock and the regulation of alternative splicing by PRMT5 constitutes a common mechanism that helps organisms to synchronize physiological processes with daily changes in environmental conditions.
Project description:Regulators of post-transcriptional events, such as pre-mRNA splicing factors, are involved in controlling the pace of the circadian clock. However, in most cases the underlying mechanisms remain unclear. Previously, we have identified XAP5 CIRCADIAN TIMEKEEPER (XCT) as an Arabidopsis thaliana circadian clock regulator with uncharacterized molecular functions. To investigate a possible role for XCT in RNA splicing, we performed PacBio Isoform Sequencing (Iso-Seq) on wild-type Col-0, reduction-of-function allele xct-1, null allele xct-2, the complemented line xct-2 XCT and prl1-2 (a mutate with known defects in intron retention). We then performed differential splicing analysis on the Iso-Seq data using JunctionSeq R package. Our results indicate that xct mutants have transcriptome-wide pre-mRNA splicing defects, predominantly aberrant 3’ splice site selection. Expression of a genomic copy of XCT fully rescues those splicing defects.
Project description:Circadian pace is modulated by light intensity, known as the Aschoff’s rule, with largely unrevealed mechanisms. Here we report that photoreceptor CRY2 mediates blue light input to circadian clock by directly interacting with clock core component PRR9 in blue light dependent manner. This physical interaction dually blocks the accessibility of PRR9 protein to its co-repressor TPL/TPRs and the resulting kinase PPKs. Notably, phosphorylation of PRR9 by PPKs is critical for its DNA binding and repressive activity, hence to ensure proper circadian speed. Given the labile nature of CRY2 in strong blue light, our findings provide a mechanistic explanation for Aschoff’s rule in plants, i.e., blue light triggers CRY2 turnover in proportional to its intensity, which accordingly releasing PRR9 to fine tune circadian speed. Our findings not only reveal a novel network mediating light input into circadian clock, but also unmask a mechanism by which Arabidopsis circadian clock sensing light intensity.