Dynamic changes in primexine during the tetrad stage of pollen development

Formation of pollen wall exine is preceded by the development of several transient layers of extracellular materials deposited on the surface of developing pollen grains. One such layer is primexine (PE), a thin, ephemeral structure that is present only for a short period of time and is difficult to visualize and study. Recent genetic studies suggested that PE is a key factor in the formation of exine, making it critical to understand its composition and the dynamics of its formation. In this study, we used high-pressure frozen/freeze-substituted samples of developing Arabidopsis (Arabidopsis thaliana) pollen for a detailed transmission electron microscopy analysis of the PE ultrastructure throughout the tetrad stage of pollen development. We also analyzed anthers from wild-type Arabidopsis and three mutants defective in PE formation by immunofluorescence, carefully tracing several carbohydrate epitopes in PE and nearby anther tissues during the tetrad and the early free-microspore stages. Our analyses revealed likely sites where these carbohydrates are produced and showed that the distribution of these carbohydrates in PE changes significantly during the tetrad stage. We also identified tools for staging tetrads and demonstrate that components of PE undergo changes resembling phase separation. Our results indicate that PE behaves like a much more dynamic structure than has been previously appreciated and clearly show that Arabidopsis PE creates a scaffolding pattern for formation of reticulate exine.

Delivery of Foreign Materials into Adherent Cells by Gold Nanoparticle-Mediated Photoporation

Delivering extracellular materials into adherent cells presents several challenges. A homemade photoporation platform, mediated by gold nanoparticles (AuNPs), was constructed to find a suitable method for finding all adherent cells in this process with high delivery efficiency. The thermal dynamics of AuNPs could be monitored. Based on this system, 60 nm AuNPs were selected to be attached to cells for optimal photoporation. After irradiating the cells covered with AuNPs using a nanosecond pulse laser, fluorescein isothiocyanate-dextran in the medium were delivered into optoporated adherent HeLa (human cervical cell lines) cells. The delivery efficiency and cell viability of this process were evaluated using a fluorescence microscope and flow cytometry. The experimental results showed that targeting cells using antibodies, laser irradiation from the top of the cell culture well, and reducing the cell medium are important for improving the delivery efficiency. The optimal loading efficiency for adherent HeLa cells was 53.4%.

A gradient of force in the endocytic coat is revealed by new coiled-coil force sensors

Clathrin mediated endocytosis (CME) is an evolutionarily conserved process responsible for the entry of extracellular materials, membrane proteins and lipids into the cell. During CME, a cargo enriched patch of membrane is deformed into an endocytic vesicle through membrane bending, elongation and scission, with the choreographed action of 60+ endocytic proteins. The plasma membrane is deformed by the forces produced by the actin cytoskeleton and transmitted to the membrane by a multi-protein coat. However, the actual forces required for endocytosis remain unknown. Here we present a new series of in vivo force sensors to measure the forces on the fission yeast HIP1R homologue End4p, a protein that links the endocytic membrane to the actin cytoskeleton. These new force sensors are based on calibrated coiled-coils that phase separate when they are under force. The measured forces on End4p are between 11 and 20 pN near the actin meshwork, between 10 and 11 pN near the clathrin lattice, and between 8 and 10 pN near the plasma membrane. Our results predict the participation of additional proteins to relay forces in different layers of the endocytic machinery during CME, and our approach points to a novel direction for in vivo force measurement.

The Dual Role of Macropinocytosis in Cancers: Promoting Growth and Inducing Methuosis to Participate in Anticancer Therapies as Targets

Macropinocytosis is an important mechanism of internalizing extracellular materials and dissolved molecules in eukaryotic cells. Macropinocytosis has a dual effect on cancer cells. On the one hand, cells expressing RAS genes (such as K-RAS, H-RAS) under the stress of nutrient deficiency can spontaneously produce constitutive macropinocytosis to promote the growth of cancer cells by internalization of extracellular nutrients (like proteins), receptors, and extracellular vesicles(EVs). On the other hand, abnormal expression of RAS genes and drug treatment (such as MOMIPP) can induce a novel cell death associated with hyperactivated macropinocytosis: methuosis. Based on the dual effect, there is immense potential for designing anticancer therapies that target macropinocytosis in cancer cells. In view of the fact that there has been little review of the dual effect of macropinocytosis in cancer cells, herein, we systematically review the general process of macropinocytosis, its specific manifestation in cancer cells, and its application in cancer treatment, including anticancer drug delivery and destruction of macropinocytosis. This review aims to serve as a reference for studying macropinocytosis in cancers and designing macropinocytosis-targeting anticancer drugs in the future.

A corset function of exoskeletal ECM promotes body elongation in Drosophila

Body elongation is a general feature of development. Postembryonically, the body needs to be framed and protected by extracellular materials, such as the skeleton, the skin and the shell, which have greater strength than cells. Thus, body elongation after embryogenesis must be reconciled with those rigid extracellular materials. Here we show that the exoskeleton (cuticle) coating the Drosophila larval body has a mechanical property to expand less efficiently along the body circumference than along the anteroposterior axis. This “corset” property of the cuticle directs a change in body shape during body growth from a relatively round shape to an elongated one. Furthermore, the corset property depends on the functions of Cuticular protein 11 A and Tubby, protein components of a sub-surface layer of the larval cuticle. Thus, constructing a stretchable cuticle and supplying it with components that confer circumferential stiffness is the fly’s strategy for executing postembryonic body elongation.

Phage Therapy Against Biofilm of Multidrug-Resistant Klebsiella Pneumoniae Isolated from Zakho Hospital Samples

Klebsiella pneumoniae causes infection in human, especially in immunocompromised patients. About 80% of nosocomial infection caused by K. pneumoniae is due to multidrug-resistant strain. The emergence of antibiotic-resistant bacterial strains necessitates the exploration of alternative antibacterial therapies, which led to studying the ability of viruses that infect the bacteria (known as bacteriophage) to treat infection with K. pneumoniae. Bacterial biofilm which are crucial in the pathogenesis of much clinically important infection and are difficult to eradicate because they exist resistant to many antimicrobial treatment. Biofilm formation by K. pneumoniae is responsible for the catheter associated infection such as urinary tract infection and respiratory tract infection due to the colonization of the polymeric surface by forming multilayered cell cluster embedded in extracellular materials. In this study K. pneumoniae isolated from the hospital environment and characterized it and form the biofilm of that organism by microplate quantitative assay. Similarly bacteriophage specific for K. pneumoniae isolated from river water. The aim of work is the use of bacteriophage as a possible alternative for the treatment of bacterial infection of K. pneumoniae. We showed that biofilm is reduced by isolated phages by the comparative account of colony-forming unit versus plaque-forming unit. The result of this study, therefore, suggests that the timing of starting the phage therapy after initiation of infection significantly contributes toward the success of the treatment.

Characteristics of Extracellular Polymeric Substances (EPS) and Enzymes during the Aerobic/anoxic Digestion of Sewage Sludge after Ultrasonic Pretreatment

In this paper, ultrasonic pretreatment was used to improve the aerobic/anoxic digestion of sewage sludge. The extracellular polymeric substances (EPS) and enzymes in the digestion were analysed to understand the mechanisms responsible for this improvement. The results showed that the ultrasonic pretreatment enhanced enzymatic activities and accelerated the solubilisation of the EPS. In addition, ultrasonic pretreatment could not only break flocs and release extracellular materials but also destroy cells and release intercellular materials. With ultrasonic pretreatment, the sludge reduction for volatile suspend solids (VSS) in aerobic/anoxic digestion was 41.97±0.66%, compared with 22.81±1.40% for the control treatment, after an aerobic/anoxic digestion time of 10 d. Fourier transform infrared (FT-IR) spectroscopy results showed that the polymeric compounds, proteins and polysaccharides were easily hydrolysed and biodegraded in ultrasonic-treated sludge.