A major feature of embryogenesis is the specification of stem cell systems, but in contrast to the situation in most animals, plant stem cells remain quiescent until the postembryonic phase of development. Here, we dissect how light and metabolic signals are integrated to overcome stem cell dormancy at the shoot apical meristem. We show on the one hand that light is able to activate expression of the stem cell inducer WUSCHEL independently of photosynthesis and that this likely involves inter-regional cytokinin signaling. Metabolic signals, on the other hand, are transduced to the meristem through activation of the TARGET OF RAPAMYCIN (TOR) kinase. Surprisingly, TOR is also required for light signal dependent stem cell activation. Thus, the TOR kinase acts as a central integrator of light and metabolic signals and a key regulator of stem cell activation at the shoot apex.DOI: http://dx.doi.org/10.7554/eLife.17023.001
The glucocorticoid receptor (GR) binds as a homodimer to genomic response elements, which have particular sequence and shape characteristics. Here we show that the nucleotides directly flanking the core-binding site, differ depending on the strength of GR-dependent activation of nearby genes. Our study indicates that these flanking nucleotides change the three-dimensional structure of the DNA-binding site, the DNA-binding domain of GR and the quaternary structure of the dimeric complex. Functional studies in a defined genomic context show that sequence-induced changes in GR activity cannot be explained by differences in GR occupancy. Rather, mutating the dimerization interface mitigates DNA-induced changes in both activity and structure, arguing for a role of DNA-induced structural changes in modulating GR activity. Together, our study shows that DNA sequence identity of genomic binding sites modulates GR activity downstream of binding, which may play a role in achieving regulatory specificity towards individual target genes.
Summary. Cross-linkage of the CD95 (FAS/APO-1) antigen is responsible for the induction of programmed cell death or apoptosis in a variety of normal and malignant cells of the haemopoietic system. In order to evaluate predominant expression of the CD95 gene in a cell lineage-specific manner, we have determined the CD95 expression patterns in cell lines of myeloid, T-, pre-B-or B-cell origin as well as those established from Hodgkin's disease (HD). Our results reveal constitutive transcriptional activation of the CD95 gene in all cell lines derived from the lymphoid and myeloid lineages. Despite the ubiquitous expression of CD95 transcripts in haemopoietic cells, the corresponding protein was undetectable in 2/5 cell lines derived from Burkitt lymphomas and 6/16 leukaemia cell lines of the megakaryocytic or monocytic lineage. In an effort to identify apoptosis-resistant cell lines resulting from mutations in the death-signalling domain of CD95 or from defects in the apoptotic pathway or in survival programmes, we applied a CD95-mediated apoptosis assay. However, 21/38 CD95-expressing cell lines were sensitive upon induction with an anti-CD95 antibody whereas the remaining cell lines (predominantly of myeloid derivation) were resistant to antibody-induced cell death. Resistance to CD95-mediated apoptosis was not due to mutations within the CD95 open reading frame as confirmed by a combined reverse transcription PCR sequencing method. Five myeloid out of 13 tumour lines with the apoptosis-resistance phenotype analysed showed programmed cell death, when protein synthesis was blocked by treatment with cycloheximide prior to CD95-mediated induction. These data suggest an active cellular mechanism for the maintenance of an apoptosis-resistant phenotype. Elucidating the steps in such an active process of resistance to apoptosis might be expected to provide new approaches for therapeutic intervention in certain tumours.
The binding of transcription factors to short recognition sequences plays a pivotal role in controlling the expression of genes. The sequence and shape characteristics of binding sites influence DNA binding specificity and have also been implicated in modulating the activity of transcription factors downstream of binding. To quantitatively assess the transcriptional activity of tens of thousands of designed synthetic sites in parallel, we developed a synthetic version of STARR-seq (synSTARR-seq). We used the approach to systematically analyze how variations in the recognition sequence of the glucocorticoid receptor (GR) affect transcriptional regulation. Our approach resulted in the identification of a novel highly active functional GR binding sequence and revealed that sequence variation both within and flanking GR’s core binding site can modulate GR activity without apparent changes in DNA binding affinity. Notably, we found that the sequence composition of variants with similar activity profiles was highly diverse. In contrast, groups of variants with similar activity profiles showed specific DNA shape characteristics indicating that DNA shape may be a better predictor of activity than DNA sequence. Finally, using single cell experiments with individual enhancer variants, we obtained clues indicating that the architecture of the response element can independently tune expression mean and cell-to cell variability in gene expression (noise). Together, our studies establish synSTARR as a powerful method to systematically study how DNA sequence and shape modulate transcriptional output and noise.
The glucocorticoid (GR) and androgen (AR) receptors execute unique functions in vivo, yet have nearly identical DNA binding specificities. To identify mechanisms that facilitate functional diversification among these transcription factor paralogs, we studied them in an equivalent cellular context. Analysis of chromatin and sequence suggest that divergent binding, and corresponding gene regulation, are driven by different abilities of AR and GR to interact with relatively inaccessible chromatin. Divergent genomic binding patterns can also be the result of subtle differences in DNA binding preference between AR and GR. Furthermore, the sequence composition of large regions (>10 kb) surrounding selectively occupied binding sites differs significantly, indicating a role for the sequence environment in guiding AR and GR to distinct binding sites. The comparison of binding sites that are shared shows that the specificity paradox can also be resolved by differences in the events that occur downstream of receptor binding. Specifically, shared binding sites display receptor-specific enhancer activity, cofactor recruitment and changes in histone modifications. Genomic deletion of shared binding sites demonstrates their contribution to directing receptor-specific gene regulation. Together, these data suggest that differences in genomic occupancy as well as divergence in the events that occur downstream of receptor binding direct functional diversification among transcription factor paralogs.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.