The challenges and opportunities of open-access microscopy facilities.

Microscopy core facilities are increasingly utilised research resources, but they are generally only available to users within the host institution. Such localised access misses an opportunity to facilitate research across a broader user base. Here, we present the model of an open-access microscopy facility, using the Advanced Imaging Center (AIC) at Howard Hughes Medical Institute Janelia Research Campus as an example.

In vivo visualization of nitrate dynamics using a genetically encoded fluorescent biosensor.

Nitrate (NO) uptake and distribution are critical to plant life. Although the upstream regulation of NO uptake and downstream responses to NO in a variety of cells have been well studied, it is still not possible to directly visualize the spatial and temporal distribution of NO with high resolution at the cellular level. Here, we report a nuclear-localized, genetically encoded fluorescent biosensor, which we named NitraMeter3.

ATP-responsive biomolecular condensates tune bacterial kinase signaling.

Biomolecular condensates formed via liquid-liquid phase separation enable spatial and temporal organization of enzyme activity. Phase separation in many eukaryotic condensates has been shown to be responsive to intracellular adenosine triphosphate (ATP) levels, although the consequences of these mechanisms for enzymes sequestered within the condensates are unknown. Here, we show that ATP depletion promotes phase separation in bacterial condensates composed of intrinsically disordered proteins.

Stress-associated developmental reprogramming in moss protonemata by synthetic activation of the common symbiosis pathway.

Symbioses between angiosperms and rhizobia or arbuscular mycorrhizal fungi are controlled through a conserved signaling pathway. Microbe-derived, chitin-based elicitors activate plant cell surface receptors and trigger nuclear calcium oscillations, which are decoded by a calcium/calmodulin-dependent protein kinase (CCaMK) and its target transcription factor interacting protein of DMI3 (IPD3). Genes encoding CCaMK and IPD3 have been lost in multiple non-mycorrhizal plant lineages yet retained among non-mycorrhizal mosses.

Live imaging of Aiptasia larvae, a model system for coral and anemone bleaching, using a simple microfluidic device.

Coral reefs, and their associated diverse ecosystems, are of enormous ecological importance. In recent years, coral health has been severely impacted by environmental stressors brought on by human activity and climate change, threatening the extinction of several major reef ecosystems. Reef damage is mediated by a process called 'coral bleaching' where corals, sea anemones, and other cnidarians lose their photosynthetic algal symbionts (family Symbiodiniaceae) upon stress induction, resulting in drastically decreased host energy harvest and, ultimately, coral death.

The Eukaryotic CO-Concentrating Organelle Is Liquid-like and Exhibits Dynamic Reorganization.

Approximately 30%-40% of global CO fixation occurs inside a non-membrane-bound organelle called the pyrenoid, which is found within the chloroplasts of most eukaryotic algae. The pyrenoid matrix is densely packed with the CO-fixing enzyme Rubisco and is thought to be a crystalline or amorphous solid. Here, we show that the pyrenoid matrix of the unicellular alga Chlamydomonas reinhardtii is not crystalline but behaves as a liquid that dissolves and condenses during cell division.

The SCAR/WAVE complex polarizes PAN receptors and promotes division asymmetry in maize.

Pre-mitotic establishment of polarity is a key event in the preparation of mother cells for asymmetric cell divisions that produce daughters of distinct fates, and ensures correct cellular patterning of tissues and eventual organ function. Previous work has shown that two receptor-like kinases, PANGLOSS2 (PAN2) and PAN1, and the small GTPase RHO GTPASE OF PLANTS (ROP) promote mother cell polarity and subsequent division asymmetry in developing maize stomata. PAN proteins become polarized prior to asymmetric cell division, however, the mechanism of this polarization is unknown.

Live imaging provides new insights on dynamic F-actin filopodia and differential endocytosis during myoblast fusion in Drosophila.

The process of myogenesis includes the recognition, adhesion, and fusion of committed myoblasts into multinucleate syncytia. In the larval body wall muscles of Drosophila, this elaborate process is initiated by Founder Cells and Fusion-Competent Myoblasts (FCMs), and cell adhesion molecules Kin-of-IrreC (Kirre) and Sticks-and-stones (Sns) on their respective surfaces. The FCMs appear to provide the driving force for fusion, via the assembly of protrusions associated with branched F-actin and the WASp, SCAR and Arp2/3 pathways.

Time-lapse fluorescence imaging of Arabidopsis root growth with rapid manipulation of the root environment using the RootChip.

The root functions as the physical anchor of the plant and is the organ responsible for uptake of water and mineral nutrients such as nitrogen, phosphorus, sulfate and trace elements that plants acquire from the soil. If we want to develop sustainable approaches to producing high crop yield, we need to better understand how the root develops, takes up a wide spectrum of nutrients, and interacts with symbiotic and pathogenic organisms. To accomplish these goals, we need to be able to explore roots in microscopic detail over time periods ranging from minutes to days.

Recent advances in imaging embryonic myoblast fusion in Drosophila.

Myoblast fusion in the Drosophila embryos is a complex process that includes changes in cell movement, morphology and behavior over time. The advent of fluorescent proteins (FPs) has made it possible to track and image live cells, to capture the process of myoblast fusion, and to carry out quantitative analysis of myoblasts in real time. By tagging proteins with FPs, it is also possible to monitor the subcellular events that accompany the fusion process.

Asymmetric Mbc, active Rac1 and F-actin foci in the fusion-competent myoblasts during myoblast fusion in Drosophila.

Myoblast fusion is an intricate process that is initiated by cell recognition and adhesion, and culminates in cell membrane breakdown and formation of multinucleate syncytia. In the Drosophila embryo, this process occurs asymmetrically between founder cells that pattern the musculature and fusion-competent myoblasts (FCMs) that account for the bulk of the myoblasts. The present studies clarify and amplify current models of myoblast fusion in several important ways.

Control of the mitotic cleavage plane by local epithelial topology.

For nearly 150 years, it has been recognized that cell shape strongly influences the orientation of the mitotic cleavage plane (e.g., Hofmeister, 1863).

PAN1: a receptor-like protein that promotes polarization of an asymmetric cell division in maize.

Polarization of cell division is essential for eukaryotic development, but little is known about how this is accomplished in plants. The formation of stomatal complexes in maize involves the polarization of asymmetric subsidiary mother cell (SMC) divisions toward the adjacent guard mother cell (GMC), apparently under the influence of a GMC-derived signal. We found that the maize pan1 gene promotes the premitotic polarization of SMCs and encodes a leucine-rich repeat receptor-like protein that becomes localized in SMCs at sites of GMC contact.

Three Brick genes have distinct functions in a common pathway promoting polarized cell division and cell morphogenesis in the maize leaf epidermis.

We have taken a genetic approach to investigating cytoskeleton-dependent mechanisms governing cell morphogenesis in the maize leaf epidermis. Previously, we showed that the Brick1 (Brk1) gene is required for the formation of epidermal cell lobes as well as for properly polarized divisions of stomatal subsidiary mother cells, and encodes an 8 kDa protein highly conserved in plants and animals. Here, we show that two additional Brick genes, Brk2 and Brk3, are involved in the same aspects of epidermal cell morphogenesis and division.