Cellulose assembles into helical bundles of uniform handedness in cell walls with abnormal pectin composition.

Plant cells and organs grow into a remarkable diversity of shapes, as directed by cell walls composed primarily of polysaccharides such as cellulose and multiple structurally distinct pectins. The properties of the cell wall that allow for precise control of morphogenesis are distinct from those of the individual polysaccharide components. For example, cellulose, the primary determinant of cell morphology, is a chiral macromolecule that can self-assemble in vitro into larger-scale structures of consistent chirality, and yet most plant cells do not display consistent chirality in their growth. One interesting exception is the Arabidopsis thaliana rhm1 mutant, which has decreased levels of the pectin rhamnogalacturonan-I and causes conical petal epidermal cells to grow with a left-handed helical twist. Here, we show that in rhm1 the cellulose is bundled into large macrofibrils, unlike the evenly distributed microfibrils of the wild type. This cellulose bundling becomes increasingly severe over time, consistent with cellulose being synthesized normally and then self-associating into macrofibrils. We also show that in the wild type, cellulose is oriented transversely, whereas in rhm1 mutants, the cellulose forms right-handed helices that can account for the helical morphology of the petal cells. Our results indicate that when the composition of pectin is altered, cellulose can form cellular-scale chiral structures in vivo, analogous to the helicoids formed in vitro by cellulose nano-crystals. We propose that an important emergent property of the interplay between rhamnogalacturonan-I and cellulose is to permit the assembly of nonbundled cellulose structures, providing plants flexibility to orient cellulose and direct morphogenesis.

Task Challenge and Employee Performance: A Moderated Mediation Model of Resilience and Digitalization.

Lately, organizations are giving attention to enhancing employee resilience due to turbulent economic times caused by lockdowns in the last couple of years. This explanatory research proposes and tests the mediating role of employee resilience to link task challenge and employee performance during the COVID-19 pandemic in government schools of Oman using the broaden-and-build theory. An explanatory research design was used for this empirical study. An Arabic-translated questionnaire version was designed to collect primary data online during the lockdown using simple random sampling. We used the Preacher and Hayes macro to analyze moderated mediation using cross-sectional data and analyzed 441 responses. The findings confirm the mediation roles of employee resilience and moderating roles of digitalization during unusual circumstances. The study has implications for the school administrators, Omani policymakers and schooling staff of the Middle Eastern educational industry.

Optimizing Health Communication Noticeability with Visual Design and Humor: An Applied Approach for COVID-19 Public Messaging.

Official public health communications are typically characterized by generic and staid graphic representation. However, situations requiring sustained public attention (such as the COVID-19 pandemic) would benefit from context-rich, referential messaging which actively incorporates visual design and emotion in its composition. This paper documents the author's application of this philosophy through a graphic poster series framing advisory information within creative, visually-engaging parameters. Summarizing the process of conceptualization, development and reception, it discusses the project's dual objectives - enhancing noticeability through creative visual design, and mitigating pandemic-related negativity with humor. Finally, it situates the project's postulations within the context of Health Communication discourse.

Commemorating exceptional valour: recognising the brave who tamed a pandemic.

COVID-19's ongoing, protracted visitation has prevented due acknowledgement of efforts to control it, particularly those of the medical workers who volunteered to serve on the frontlines, disregarding uncertainty and personal risk, in the early days when the consequences of exposure were unknown. This artwork seeks to memorialise the diligence, selflessness, and professional commitment of the medical community in a time of extraordinary difficulty. Referencing historical patterns of iconography, this graphic imagery proposes a modernised representation of bravery by interweaving narratives of equality, inclusivity, humour and optimism, to recognise the unnamed many who helped make the world a safer place.

Cytokinesis in vertebrate cells initiates by contraction of an equatorial actomyosin network composed of randomly oriented filaments.

The actomyosin ring generates force to ingress the cytokinetic cleavage furrow in animal cells, yet its filament organization and the mechanism of contractility is not well understood. We quantified actin filament order in human cells using fluorescence polarization microscopy and found that cleavage furrow ingression initiates by contraction of an equatorial actin network with randomly oriented filaments. The network subsequently gradually reoriented actin filaments along the cell equator. This strictly depended on myosin II activity, suggesting local network reorganization by mechanical forces. Cortical laser microsurgery revealed that during cytokinesis progression, mechanical tension increased substantially along the direction of the cell equator, while the network contracted laterally along the pole-to-pole axis without a detectable increase in tension. Our data suggest that an asymmetric increase in cortical tension promotes filament reorientation along the cytokinetic cleavage furrow, which might have implications for diverse other biological processes involving actomyosin rings.

Quantification of collagen organization in histopathology samples using liquid crystal based polarization microscopy.

A number of histopathology studies have utilized the label free microscopy method of Second Harmonic Generation (SHG) to investigate collagen organization in disease onset and progression. Here we explored an alternative label free imaging approach, LC-PolScope that is based on liquid crystal based polarized light imaging. We demonstrated that this more accessible technology has the ability to visualize all fibers of interest and has a good to excellent correlation between SHG and LC-PolScope measurements in fibrillar collagen orientation and alignment. This study supports that LC-PolScope is a viable alternative to SHG for label free collagen organization measurements in thin histology sections.

Analysis of Septin Reorganization at Cytokinesis Using Polarized Fluorescence Microscopy.

Septins are conserved filament-forming proteins that act in diverse cellular processes. They closely associate with membranes and, in some systems, components of the cytoskeleton. It is not well understood how filaments assemble into higher-order structures in vivo or how they are remodeled throughout the cell cycle. In the budding yeast S. cerevisiae, septins are found through most of the cell cycle in an hourglass organization at the mother-bud neck until cytokinesis when the collar splits into two rings that disassemble prior to the next cell cycle. Experiments using polarized fluorescence microscopy have suggested that septins are arranged in ordered, paired filaments in the hourglass and undergo a coordinated 90° reorientation during splitting at cytokinesis. This apparent reorganization could be due to two orthogonal populations of filaments disassembling and reassembling or being preferentially retained at cytokinesis. In support of this idea, we report a decrease in septin concentration at the mother-bud neck during cytokinesis consistent with other reports and the timing of the decrease depends on known septin regulators including the Gin4 kinase. We took a candidate-based approach to examine what factors control reorientation during splitting and used polarized fluorescence microscopy to screen mutant yeast strains deficient in septin interacting proteins. Using this method, we have linked known septin regulators to different aspects of the assembly, stability, and reorganization of septin assemblies. The data support that ring splitting requires Gin4 activity and an anillin-like protein Bud4, and normal accumulation of septins at the ring requires phosphorylation of Shs1. We found distinct regulatory requirements for septin organization in the hourglass compared to split rings. We propose that septin subpopulations can vary in their localization and assembly/disassembly behavior in a cell-cycle dependent manner at cytokinesis.

Dissection of molecular assembly dynamics by tracking orientation and position of single molecules in live cells.

Regulation of order, such as orientation and conformation, drives the function of most molecular assemblies in living cells but remains difficult to measure accurately through space and time. We built an instantaneous fluorescence polarization microscope, which simultaneously images position and orientation of fluorophores in living cells with single-molecule sensitivity and a time resolution of 100 ms. We developed image acquisition and analysis methods to track single particles that interact with higher-order assemblies of molecules. We tracked the fluctuations in position and orientation of molecules from the level of an ensemble of fluorophores down to single fluorophores. We tested our system in vitro using fluorescently labeled DNA and F-actin, in which the ensemble orientation of polarized fluorescence is known. We then tracked the orientation of sparsely labeled F-actin network at the leading edge of migrating human keratinocytes, revealing the anisotropic distribution of actin filaments relative to the local retrograde flow of the F-actin network. Additionally, we analyzed the position and orientation of septin-GFP molecules incorporated in septin bundles in growing hyphae of a filamentous fungus. Our data indicate that septin-GFP molecules undergo positional fluctuations within ∼350 nm of the binding site and angular fluctuations within ∼30° of the central orientation of the bundle. By reporting position and orientation of molecules while they form dynamic higher-order structures, our approach can provide insights into how micrometer-scale ordered assemblies emerge from nanoscale molecules in living cells.

Polarized Fluorescence Microscopy to Study Cytoskeleton Assembly and Organization in Live Cells.

The measurement of not only the location but also the organization of molecules in live cells is crucial to understanding diverse biological processes. Polarized light microscopy provides a nondestructive means to evaluate order within subcellular domains. When combined with fluorescence microscopy and GFP-tagged proteins, the approach can reveal organization within specific populations of molecules. This unit describes a protocol for measuring the architectural dynamics of cytoskeletal components using polarized fluorescence microscopy and OpenPolScope open-access software (http://www.openpolscope.org). The protocol describes installation of linear polarizers or a liquid crystal (LC) universal compensator, calibration of the system, polarized fluorescence imaging, and analysis. The use of OpenPolScope software and hardware allows for reliable, user-friendly image acquisition to measure and analyze polarized fluorescence.

MultiFocus Polarization Microscope (MF-PolScope) for 3D polarization imaging of up to 25 focal planes simultaneously.

We have developed an imaging system for 3D time-lapse polarization microscopy of living biological samples. Polarization imaging reveals the position, alignment and orientation of submicroscopic features in label-free as well as fluorescently labeled specimens. Optical anisotropies are calculated from a series of images where the sample is illuminated by light of different polarization states. Due to the number of images necessary to collect both multiple polarization states and multiple focal planes, 3D polarization imaging is most often prohibitively slow. Our MF-PolScope system employs multifocus optics to form an instantaneous 3D image of up to 25 simultaneous focal-planes. We describe this optical system and show examples of 3D multi-focus polarization imaging of biological samples, including a protein assembly study in budding yeast cells.

Polarized light microscopy in reproductive and developmental biology.

The polarized light microscope reveals orientational order in native molecular structures inside living cells, tissues, and whole organisms. It is a powerful tool used to monitor and analyze the early developmental stages of organisms that lend themselves to microscopic observations. In this article, we briefly discuss the components specific to a traditional polarizing microscope and some historically important observations on: chromosome packing in the sperm head, the first zygote division of the sea urchin, and differentiation initiated by the first asymmetric cell division in the sand dollar. We then introduce the LC-PolScope and describe its use for measuring birefringence and polarized fluorescence in living cells and tissues. Applications range from the enucleation of mouse oocytes to analyzing the polarized fluorescence of the water strider acrosome. We end with new results on the birefringence of the developing chick brain, which we analyzed between developmental stages of days 12-20.

pH dependent isotropic to nematic phase transitions in graphene oxide dispersions reveal droplet liquid crystalline phases.

Size fractionation, amplified by the surface charge density of graphene oxide (GO) sheets, broadens the pH dependent isotropic (I) to nematic (N) phase transition in aqueous dispersions of graphene oxide (GO). In this biphasic region, a highly organized droplet nematic phase of uniform size (20 ± 2.8 µm diameter) with an isotropic interior is observed.

Anticoagulation in patients with mechanical valves during pregnancy.

Mechanical valve thrombosis is a life-threatening event, while pregnancy is associated with a hypercoagulable state. Thus, in pregnant women with mechanical valves, adequate anticoagulation becomes even more critical. This prospective study was conducted to establish a uniform anticoagulation regimen for these women. A total of 250 pregnancies in 245 women with mechanical heart valves were evaluated. The patients were divided into 2 groups: group 1 (n = 150) took oral warfarin throughout pregnancy and group 2 (n = 100) received subcutaneous heparin in the 1(st) trimester and oral warfarin for the other trimesters. Both groups received heparin at the time of delivery. There were no coumarin-induced fetal malformations. Minor thromboembolic episodes took place in 5 women in group 1 and 3 in group 2. Valve thrombosis occurred in 1 woman in group 2 and led to 1 maternal death in this series. The incidence of spontaneous abortion was similar between the groups. We conclude that warfarin is safe and convenient to use during pregnancy. The teratogenic effects of warfarin during the 1(st) trimester are overstated, and switching to heparin is not mandatory.