Project description:In plants, apical meristems allow continuous growth along the body axis. Within the root apical meristem (RAM), a group of slowly dividing quiescent center (QC) cells is thought to limit stem cell activity to directly neighboring cells (Cowels, 1956; van den Berg et al., 1997), thus endowing them with unique properties, distinct from displaced daughters. This binary identity of the stem cells stands in apparent contradiction with the more gradual changes in cell division potential (Bennett and Scheres, 2010) and differentiation (Yamaguchi et al., 2008; 2010; Furuta et al, 2014; Geldner, 2013; Masucci et al., 1996; Dolan and Costa, 2001) that occur as cells move further away from the QC. To address this paradox and to infer molecular organization of the root meristem, we used a whole-genome approach to determine dominant transcriptional patterns along root ontogeny zones. We found that the prevalent patterns are expressed in two opposing gradients. One is characterized by genes associated with development, the other enriched in differentiation genes. We confirmed these transcript gradients, and demonstrate that these translate to gradients in protein accumulation and gradual changes in cellular properties. We also show that gradients are genetically controlled through multiple pathways. Based on these findings, we propose that cells in the Arabidopsis root meristem gradually transition from ‘stemness’ towards differentiation.
Project description:In this study, we sought to determine whether the intensity of the immune response to Plasmodium spp., the parasite causing malaria, depends on time of infection. For this, we considered the known existence of a circadian clock in immune cells which regulates several aspects of the immune response. Experiments were performed using mouse bone marrow-derived macrophages (BMDMs) stimulated ex vivo with red blood cells infected with Plasmodium berghei ANKA (iRBCs) or uninfected RBCs. First, we identified that lysed iRBCs triggered an immune response in macrophages. Then, by stimulating at 4 different circadian time points (16 h, 22 h, 28 h or 34 h post-synchronization of the cells clock), rhythms in reactive oxygen species and cytokines/chemokines were found. Furthermore, the analysis of the macrophage proteome and phosphoproteome revealed global changes according to treatment (iRBC or RBC) and to the circadian time point. In summary, our findings showed that the circadian clock within macrophages determines the magnitude of immune response upon stimulation with iRBCs, along with changes of the cell proteome and phosphoproteome.
Project description:Genetic and molecular approaches have been critical for elucidating the mechanism of the mammalian circadian clock. Here, we demonstrate that the ClockM-NM-^T19 mutant behavioral phenotype is significantly modified by mouse strain genetic background. We map a suppressor of the ClockM-NM-^T19 mutation to a M-bM-^HM-<900 kb interval on mouse chromosome 1 and identify the transcription factor, Usf1, as the responsible gene. A SNP in the promoter of Usf1 causes elevation of its transcript and protein in strains that suppress the Clock mutant phenotype. USF1 competes with the CLOCK:BMAL1 complex for binding to E-box sites in target genes. Saturation binding experiments demonstrate reduced affinity of the CLOCKM-NM-^T19:BMAL1 complex for E-box sites, thereby permitting increased USF1 occupancy on a genome-wide basis. We propose that USF1 is an important modulator of molecular and behavioral circadian rhythms in mammals. DOI:http://dx.doi.org/10.7554/eLife.00426.001. Examination of 3 transcriptional regulators in mouse liver at ZT8
Project description:● Adjustment to energy starvation is crucial to ensure growth and survival. In Arabidopsis thaliana (Arabidopsis), this process relies in part on the phosphorylation of the circadian clock regulator bZIP63 by SnRK1, a key mediator of responses to low energy. ● We investigated the effects of mutations in bZIP63 on plant carbon (C) metabolism and growth. Results from phenotypic, transcriptomic, and metabolomic analysis of bZIP63 mutants prompted us to investigate the starch accumulation pattern and the expression of genes involved in starch degradation and in the circadian oscillator. ● In order to get some clues about the role of transcription factor bZIP type bZIP63 in growth and development, we performed a comparative gene expression analysis of bzip63-2 mutant and wild type Ws, harvested at the end of night (EN = ZT 24), immediately before the onset of the light, to maximize the discovery of genes misregulated in bzip63-2. The resulting gene expression profiles revealed 230 upregulated and 117 downregulated genes in bzip63-2 compared to Ws. ● bZIP63 mutation impairs growth under light-dark cycles, but not under constant light. The reduced growth likely results from the accentuated C depletion towards the end of the night, which is caused by the accelerated starch degradation of bZIP63 mutants. The diel expression pattern of bZIP63 is dictated by both the circadian clock and energy levels, which could determine the changes in the circadian expression of clock and starch metabolic genes observed in bZIP63 mutants. ● We conclude that bZIP63 composes a regulatory interface between the metabolic and circadian control of starch breakdown to optimize C usage and plant growth.
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.