Cell type- and transcription-independent spatial proximity between enhancers and promoters.

Cell type-specific enhancers are critically important for lineage specification. The mechanisms that determine cell-type specificity of enhancer activity, however, are not fully understood. Most current models for how enhancers function invoke physical proximity between enhancer elements and their target genes. Here, we use an imaging-based approach to examine the spatial relationship of cell type-specific enhancers and their target genes with single-cell resolution. Using high-throughput microscopy, we measure the spatial distance from target promoters to their cell type-specific active and inactive enhancers in individual pancreatic cells derived from distinct lineages. We find increased proximity of all promoter-enhancer pairs relative to non-enhancer pairs separated by similar genomic distances. Strikingly, spatial proximity between enhancers and target genes was unrelated to tissue-specific enhancer activity. Furthermore, promoter-enhancer proximity did not correlate with the expression status of target genes. Our results suggest that promoter-enhancer pairs exist in a distinctive chromatin environment but that genome folding is not a universal driver of cell-type specificity in enhancer function.

Advances in cutaneous squamous cell carcinoma.

Human malignancies arise predominantly in tissues of epithelial origin, where the stepwise transformation from healthy epithelium to premalignant dysplasia to invasive neoplasia involves sequential dysregulation of biological networks that govern essential functions of epithelial homeostasis. Cutaneous squamous cell carcinoma (cSCC) is a prototype epithelial malignancy, often with a high tumour mutational burden. A plethora of risk genes, dominated by UV-induced sun damage, drive disease progression in conjunction with stromal interactions and local immunomodulation, enabling continuous tumour growth. Recent studies have identified subpopulations of SCC cells that specifically interact with the tumour microenvironment. These advances, along with increased knowledge of the impact of germline genetics and somatic mutations on cSCC development, have led to a greater appreciation of the complexity of skin cancer pathogenesis and have enabled progress in neoadjuvant immunotherapy, which has improved pathological complete response rates. Although measures for the prevention and therapeutic management of cSCC are associated with clinical benefit, the prognosis remains poor for advanced disease. Elucidating how the genetic mechanisms that drive cSCC interact with the tumour microenvironment is a current focus in efforts to understand, prevent and treat cSCC.

Identification of Novel Regulators of Plant Transpiration by Large-Scale Thermal Imaging Screening in Helianthus Annuus.

Plant adaptation to biotic and abiotic stresses is governed by a variety of factors, among which the regulation of stomatal aperture in response to water deficit or pathogens plays a crucial role. Identifying small molecules that regulate stomatal movement can therefore contribute to understanding the physiological basis by which plants adapt to their environment. Large-scale screening approaches that have been used to identify regulators of stomatal movement have potential limitations: some rely heavily on the abscisic acid (ABA) hormone signaling pathway, therefore excluding ABA-independent mechanisms, while others rely on the observation of indirect, long-term physiological effects such as plant growth and development. The screening method presented here allows the large-scale treatment of plants with a library of chemicals coupled with a direct quantification of their transpiration by thermal imaging. Since evaporation of water through transpiration results in leaf surface cooling, thermal imaging provides a non-invasive approach to investigate changes in stomatal conductance over time. In this protocol, Helianthus annuus seedlings are grown hydroponically and then treated by root feeding, in which the primary root is cut and dipped into the chemical being tested. Thermal imaging followed by statistical analysis of cotyledonary temperature changes over time allows for the identification of bioactive molecules modulating stomatal aperture. Our proof-of-concept experiments demonstrate that a chemical can be carried from the cut root to the cotyledon of the sunflower seedling within 10 minutes. In addition, when plants are treated with ABA as a positive control, an increase in leaf surface temperature can be detected within minutes. Our method thus allows the efficient and rapid identification of novel molecules regulating stomatal aperture.

ABA-glucose ester hydrolyzing enzyme ATBG1 and PHYB antagonistically regulate stomatal development.

The integration of conflicting signals in response to environmental constraints is essential to efficient plant growth and development. The light-dependent and the stress hormone abscisic acid (ABA)-dependent signaling pathways play opposite roles in many aspects of plant development. While these pathways have been extensively studied, the complex nature of their molecular dialogue is still obscure. When mobilized by the Arabidopsis thaliana ß-glucosidase 1 (AtBG1), the glucose ester-conjugated inactive form of ABA has proven to be a source of the active hormone that is essential for the adaptation of the plant to water deficit, as evidenced by the impaired stomatal closure of atbg1 mutants in response to water stress. In a suppressor screen designed to identify the molecular components of AtBG1-associated physiological and developmental mechanisms, we identified the mutation variant of AtBG1 traits (vat1), a new mutant allele of the red light/far-red light photoreceptor PHYTOCHROME B (PHYB). Our study reveals that atbg1 plants harbor increased stomatal density in addition to impaired stomatal closure. We also provide evidence that the vat1/phyb mutation can restore the apparent transpiration of the atbg1 mutant by decreasing stomatal aperture and restoring a stomatal density similar to wild-type plants. Expression of key regulators of stomatal development showed a crosstalk between AtBG1-mediated ABA signaling and PHYB-mediated stomatal development. We conclude that the AtBG1-dependent regulation of ABA homeostasis and the PHYB-mediated light signaling pathways act antagonistically in the control of stomatal development.