Cellular & Molecular Imaging

Immunofluorescence microscopy images supporting expression of HA-tagged PfMD3 in specific stages of P. falciparum development

Protein synthesis is a core cellular process, necessary throughout the complex lifecycle of Plasmodium parasites, thus specific translation inhibitors would be a valuable class of antimalarial drugs, capable of both treating symptomatic infections in the blood and providing chemoprotection by targeting the initial parasite population in the liver, preventing both human disease and parasite transmission back to the mosquito host. As increasing numbers of antiplasmodial compounds are identified that converge mechanistically at inhibition of cytoplasmic translation, regardless of molecular target or mechanism, it would be useful to gain deeper understanding of how their effectiveness as liver stage translation inhibitors relates to their chemoprotective potential. Here, we probed that relationship using the P. berghei-HepG2 liver stage infection model. Using o-propargyl puromycin-based labeling of the nascent proteome in P. berghei-infected HepG2 monolayers coupled with automated confocal feedback microscopy to generate unbiased, single parasite image sets of P. berghei liver stage translation, we determined translation inhibition EC50s for five compounds, encompassing parasite-specific aminoacyl tRNA synthetase inhibitors, compounds targeting the ribosome in both host and parasite, as well as DDD107498, which targets Plasmodium eEF2, and is a leading antimalarial candidate compound being clinically developed as cabamiquine. Compounds were then tested at equivalent effective concentrations to compare the parasite response to, and recovery from, a brief period of translation inhibition in early schizogony.

A sustainable approach to address climate change and increasing water demand in agriculture is breeding for plant functional traits that conserve water and enhance climate resilience. Stomata regulate plant-water relations and are promising targets for crop improvement. Here, we investigate the variation in stomatal density (SD) in a diverse apple population (Malus domestica Borkh.) consisting of 269 accessions. Genome-wide association studies identified robust associations with SD on chromosomes 2, 9, and 10 (classified as SNPs with p value higher than adjusted Bonferroni threshold of -log10(p) > 8.78 that were consistently identified across datasets). On chromosome 9, a candidate gene that negatively regulates stomatal development, EPIDERMAL PATTERNING FACTOR 1 (EPF1), was identified inside a genomic region of 241 kb determined by six robust associations. On chromosome 10, a positive regulator candidate gene, EPIDERMAL PATTERNING FACTOR LIKE 9 (STOMAGEN), was identified 1680 kb from the robust association. Identification of positive (STOMAGEN) and negative (EPF1) regulators of SD suggest potential antagonistic roles at the population scale in determining SD. On chromosome 2, a gene co-expression analysis identified a gene cluster containing both EPF1 and STOMAGEN together with a novel candidate gene, CYTOCHROME P450 (CYP77A4), that was located 544 kb from the robust association. The percentage of SD phenotypic variance explained by each robust association was between 7% and 10%. These findings provide a foundation for understanding SD variation at the population scale and opportunities to modulate SD by genomics-assisted breeding strategies.

Sequencing the human genome gave new insights into human biology and disease. However, the ultimate goal is to understand the dynamic expression of each of the approximately 20,000 protein-coding genes and the function of each protein. Uhlen et al. now present a map of protein expression across 32 human tissues. They not only measured expression at an RNA level, but also used antibody profiling to precisely localize the corresponding proteins.

Sequencing the human genome gave new insights into human biology and disease. However, the ultimate goal is to understand the dynamic expression of each of the approximately 20,000 protein-coding genes and the function of each protein. Uhlen et al. now present a map of protein expression across 32 human tissues. They not only measured expression at an RNA level, but also used antibody profiling to precisely localize the corresponding proteins.

Indocyanine Green (ICG) has seen widespread use in the operating room as a fluorescent vascular imaging agent. However, its rapid vascular clearance often necessitates redosing, resulting in procedural delays, greater background, and potential adverse physiological effects. Here, we present Chromenylium Green (ChromG), a novel fluorescent tracer exhibiting excellent visualization of vasculature in mice with greater contrast and seven times the vascular half-life relative to ICG. ChromG allows for high-resolution imaging of vasculature at greater depths, enabling whole-body 3D vascular reconstruction with up to 74% connectivity. As a proof of concept of ChromG’s utility in disease contexts, we demonstrate the ability to visualize saphenous artery stenoses above the clinically accepted contrast limit for eight times longer than ICG. With its excellent biocompatibility profile and compatibility with current clinical imaging technologies, ChromG is a promising fluorescent probe for prolonged, high-resolution intraoperative vascular imaging.

Indocyanine Green (ICG) has seen widespread use in the operating room as a fluorescent vascular imaging agent. However, its rapid vascular clearance often necessitates redosing, resulting in procedural delays, greater background, and potential adverse physiological effects. Here, we present Chromenylium Green (ChromG), a novel fluorescent tracer exhibiting excellent visualization of vasculature in mice with greater contrast and seven times the vascular half-life relative to ICG. ChromG allows for high-resolution imaging of vasculature at greater depths, enabling whole-body 3D vascular reconstruction with up to 74% connectivity. As a proof of concept of ChromG’s utility in disease contexts, we demonstrate the ability to visualize saphenous artery stenoses above the clinically accepted contrast limit for eight times longer than ICG. With its excellent biocompatibility profile and compatibility with current clinical imaging technologies, ChromG is a promising fluorescent probe for prolonged, high-resolution intraoperative vascular imaging.

This dataset comprises the raw data associated with Wong et al., EMBO J., 2024. "Regulation of centrosome size by the cell cycle oscillator in Drosophila embryos". This dataset (approximately 900Gb in total) tracks the behavior of individual centrosomes (labelled with different centrosome markers) in early Drosophila embryos during nuclear cycles 11-13 under various conditions. The datasets are organised to align with each individual Figure shown in the paper. The paper abstract is: Mitotic centrosomes assemble when centrioles recruit large amounts of pericentriolar material (PCM) around themselves. In early C. elegans embryos, mitotic centrosome size appears to be set by the limiting amount of a key component. In Drosophila syncytial embryos, thousands of mitotic centrosomes are assembled as the embryo proceeds through 13 rounds of rapid nuclear division, driven by a core cell-cycle oscillator. These divisions slow during nuclear cycles 11-13, and we find that centrosomes respond by reciprocally decreasing their growth rate, but increasing their growth period—so that they grow to a relatively consistent size at each cycle. At the start of each cycle, moderate CCO activity initially promotes centrosome growth, in part by stimulating Polo/PLK1 recruitment to centrosomes. Later in each cycle, high CCO activity inhibits centrosome growth by suppressing the centrosomal recruitment and/or maintenance of centrosome proteins. Thus, in fly embryos, mitotic centrosome size appears to be regulated predominantly by the core cell-cycle oscillator, rather than by the depletion of a limiting component.

This dataset comprises the raw data associated with Wong et al., EMBO J., 2024. "Regulation of centrosome size by the cell cycle oscillator in Drosophila embryos". This dataset (approximately 900Gb in total) tracks the behavior of individual centrosomes (labelled with different centrosome markers) in early Drosophila embryos during nuclear cycles 11-13 under various conditions. The datasets are organised to align with each individual Figure shown in the paper. The paper abstract is: Mitotic centrosomes assemble when centrioles recruit large amounts of pericentriolar material (PCM) around themselves. In early C. elegans embryos, mitotic centrosome size appears to be set by the limiting amount of a key component. In Drosophila syncytial embryos, thousands of mitotic centrosomes are assembled as the embryo proceeds through 13 rounds of rapid nuclear division, driven by a core cell-cycle oscillator. These divisions slow during nuclear cycles 11-13, and we find that centrosomes respond by reciprocally decreasing their growth rate, but increasing their growth period—so that they grow to a relatively consistent size at each cycle. At the start of each cycle, moderate CCO activity initially promotes centrosome growth, in part by stimulating Polo/PLK1 recruitment to centrosomes. Later in each cycle, high CCO activity inhibits centrosome growth by suppressing the centrosomal recruitment and/or maintenance of centrosome proteins. Thus, in fly embryos, mitotic centrosome size appears to be regulated predominantly by the core cell-cycle oscillator, rather than by the depletion of a limiting component.

This dataset comprises the raw data associated with Wong et al., EMBO J., 2024. "Regulation of centrosome size by the cell cycle oscillator in Drosophila embryos". This dataset (approximately 900Gb in total) tracks the behavior of individual centrosomes (labelled with different centrosome markers) in early Drosophila embryos during nuclear cycles 11-13 under various conditions. The datasets are organised to align with each individual Figure shown in the paper. The paper abstract is: Mitotic centrosomes assemble when centrioles recruit large amounts of pericentriolar material (PCM) around themselves. In early C. elegans embryos, mitotic centrosome size appears to be set by the limiting amount of a key component. In Drosophila syncytial embryos, thousands of mitotic centrosomes are assembled as the embryo proceeds through 13 rounds of rapid nuclear division, driven by a core cell-cycle oscillator. These divisions slow during nuclear cycles 11-13, and we find that centrosomes respond by reciprocally decreasing their growth rate, but increasing their growth period—so that they grow to a relatively consistent size at each cycle. At the start of each cycle, moderate CCO activity initially promotes centrosome growth, in part by stimulating Polo/PLK1 recruitment to centrosomes. Later in each cycle, high CCO activity inhibits centrosome growth by suppressing the centrosomal recruitment and/or maintenance of centrosome proteins. Thus, in fly embryos, mitotic centrosome size appears to be regulated predominantly by the core cell-cycle oscillator, rather than by the depletion of a limiting component.