Sterols are required for cell‐fate commitment and maintenance of the stomatal lineage in Arabidopsis

Asymmetric cell division is important for regulating cell proliferation and fate determination during stomatal development in plants. Although genes that control asymmetric division and cell differentiation in stomatal development have been reported, regulators controlling the process from asymmetric division to cell differentiation remain poorly understood. Here, we report a weak allele (fk–J3158) of the Arabidopsis sterol C–14 reductase gene FACKEL (FK) that shows clusters of small cells and stomata in leaf epidermis, a common phenomenon that is often seen in mutants defective in stomatal asymmetric division. Interestingly, the physical asymmetry of these divisions appeared to be intact in fk mutants, but the cell‐fate asymmetry was greatly disturbed, suggesting that the FK pathway links these two crucial events in the process of asymmetric division. Sterol profile analysis revealed that the fk–J3158 mutation blocked downstream sterol production. Further investigation indicated that cyclopropylsterol isomerase1 (cpi1), sterol 14α–demethylase (cyp51A2) and hydra1 (hyd1) mutants, corresponding to enzymes in the same branch of the sterol biosynthetic pathway, displayed defective stomatal development phenotypes, similar to those observed for fk. Fenpropimorph, an inhibitor of the FK sterol C–14 reductase in Arabidopsis, also caused these abnormal small‐cell and stomata phenotypes in wild‐type leaves. Genetic experiments demonstrated that sterol biosynthesis is required for correct stomatal patterning, probably through an additional signaling pathway that has yet to be defined. Detailed analyses of time‐lapse cell division patterns, stomatal precursor cell division markers and DNA ploidy suggest that sterols are required to properly restrict cell proliferation, asymmetric fate specification, cell‐fate commitment and maintenance in the stomatal lineage cells. These events occur after physical asymmetric division of stomatal precursor cells.     

Enzymatic Synthesis of s-Substituted l-Cysteines with Tryptophan Synthase of Escherichia coli

The α₂β₂ complex of tryptophan synthase from Escherichia coli catalyzes β-replacement reactions of l-serine and its derivatives (e.g., β-chloro-l-alanine and O-methyl-Dl-serine) with various alkanethiols. The products from thiobenzyl alcohol and ethanethiol were isolated to demonstrate the enzymatic synthesis of the corresponding S-substituted l-cysteines. Reactivities of various S-substituent donors were examined, and thiols such as thiobenzyl alcohol, 1-propanethiol and 1-butanethiol were found to be much more efficient substituent donors than the physiological substrate, indole. In addition, tryptophan synthase catalyzes β-replacement reactions of l-threonine with thiols to form the corresponding S-substituted β-methylcysteines, which are also produced by β-addition reactions of l-vinylglycine with thiols. These enzymatic reactions facilitate the synthesis of various sulfur-containing amino acids.     

DCC Expression by Neurons Regulates Synaptic Plasticity in the Adult Brain

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Highlights? DCC and netrin-1 are enriched at synapses in the adult mouse forebrain ? DCC is enriched in the PSD and regulates dendritic spine morphology ? LTP induction and memory formation require DCC expression by neurons ? DCC activation of Src is required for NMDAR-dependent LTP in adult CNS     

Endogenous Purification Reveals GREB1 as a Key Estrogen Receptor Regulatory Factor

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Highlights? A proteomic method identifies protein-protein interaction in primary tumors ? GREB1 is the top estrogen-induced ER-interacting protein ? GREB1 is an essential ER cofactor recruited to chromatin ? GREB1 is an independent prognostic marker     

An integrated practical implementation of continuous aqueous two‐phase systems for the recovery of human IgG: From the microdevice to a multistage bench‐scale mixer‐settler device

Aqueous two‐phase systems (ATPS) are a liquid‐liquid extraction technology with clear process benefits; however, its lack of industrial embracement is still a challenge to overcome. Antibodies are a potential product to be recovered by ATPS in a commercial context. The objective of this work is to present a more integral approach of the different isolated strategies that have arisen in order to enable a practical, generic implementation of ATPS, using human immunoglobulin G (IgG) as experimental model. A microfluidic device is used for ATPS parameters preselection for product recovery. ATPS were continuously operated in a mixer‐settler device in one stage, multistage and multistage with recirculation configuration. Single‐stage pure IgG extraction with a polyethylene glycol (PEG) 3350‐phophates ATPS within continuous operation allowed a 65% recovery. Further implementation of a multistage platform promoted a higher particle partitioning reaching a 90% recovery. The processing of IgG from a cell supernatant culture harvest in a multistage system with top phase recirculation resulted in 78% IgG recovery in bottom phase. This work conjugates three not widely spread methodologies for ATPS: microfluidics, continuous and multistage operation.     

Dengue Outbreak in Mombasa City,Kenya, 2013–2014: Entomologic Investigations

Dengue outbreaks were first reported in East Africa in the late 1970s to early 1980s including the 1982 outbreak on the Kenyan coast. In 2011, dengue outbreaks occurred in Mandera in northern Kenya and subsequently in Mombasa city along the Kenyan coast in 2013–2014. Following laboratory confirmation of dengue fever cases, an entomologic investigation was conducted to establish the mosquito species, and densities, causing the outbreak. Affected parts of the city were identified with the help of public health officials. Adult Ae. aegypti mosquitoes were collected using various tools, processed and screened for dengue virus (DENV) by cell culture and RT-PCR. All containers in every accessible house and compound within affected suburbs were inspected for immatures. A total of 2,065 Ae. aegypti adults were collected and 192 houses and 1,676 containers inspected. An overall house index of 22%, container index, 31.0% (indoor = 19; outdoor = 43) and Breteau index, 270.1, were observed, suggesting that the risk of dengue transmission was high. Overall, jerry cans were the most productive containers (18%), followed by drums (17%), buckets (16%), tires (14%) and tanks (10%). However, each site had specific most-productive container-types such as tanks (17%) in Kizingo; Drums in Nyali (30%) and Changamwe (33%), plastic basins (35%) in Nyali-B and plastic buckets (81%) in Ganjoni. We recommend that for effective control of the dengue vector in Mombasa city, all container types would be targeted. Measures would include proper covering of water storage containers and eliminating discarded containers outdoors through a public participatory environmental clean-up exercise. Providing reliable piped water to all households would minimize the need for water storage and reduce aquatic habitats. Isolation of DENV from male Ae. aegypti mosquitoes is a first observation in Kenya and provides further evidence that transovarial transmission may have a role in DENV circulation and/or maintenance in the environment.     

Bioecological Drivers of Rabies Virus Circulation in a Neotropical Bat Community

Introduction

In addition to the commonly accepted importance of the vampire bat in the maintenance and transmission of the rabies virus (RABV) in South America, RABV infection of other species is widely evidenced, challenging their role in the viral cycle.

Methodology / Principles findings

To identify the bioecological drivers of RABV circulation in neotropical bat communities, we conducted a molecular and serological survey on almost 1,000 bats from 30 species, and a 4-year longitudinal survey in two colonies of vampire bats in French Guiana. RABV was molecularly detected in a common vampire and in a frugivorous bat. The sequences corresponded to haematophagous bat-related strains and were close to viruses circulating in the Brazilian Amazon region. Species’ seroprevalence ranged from 0 to 20%, and the risk of seropositivity was higher in bats with a haematophagous diet, living in monospecific colonies and in dense forests. The longitudinal survey showed substantial temporal fluctuations, with individual waves of seroconversions and waning immunity. The high prevalences observed in bat communities, in most habitats and in species that do not share the same microhabitats and bioecological patterns, the temporal variations, and a rather short period of detectable antibodies as observed in recaptured vampires suggest (i) frequent exposure of animals, (ii) an ability of the infected host to control and eliminate the virus, (iii) more relaxed modes of exposure between bats than the commonly assumed infection via direct contact with saliva of infected animals, all of which should be further investigated.

Conclusions / significance

We hypothesize that RABV circulation in French Guiana is mainly maintained in the pristine forest habitats that may provide sufficient food resources to allow vampire bats, the main prevalent species, to survive and RABV to be propagated. However, on the forest edge and in disturbed areas, human activities may induce more insidious effects such as defaunation. One of the ecological consequences is the disappearance of resources for tertiary or secondary consumers. Populations of vampires may then shift to alternative resources such as cattle, domestic animals and humans. Therefore, a good forest status, allowing both a dilution effect in highly rich bat communities and the maintenance of large populations of medium-sized and large mammals used as prey by vampires, should prevent their migration to anthropized areas.     

Down-Regulation of TLR and JAK/STAT Pathway Genes Is Associated with Diffuse Cutaneous Leishmaniasis: A Gene Expression Analysis in NK Cells from Patients Infected with Leishmania mexicana

An important NK-cell inhibition with reduced TNF-α, IFN-γ and TLR2 expression had previously been identified in patients with diffuse cutaneous leishmaniasis (DCL) infected with Leishmania mexicana. In an attempt to pinpoint alterations in the signaling pathways responsible for the NK-cell dysfunction in patients with DCL, this study aimed at identifying differences in the NK-cell response towards Leishmania mexicana lipophosphoglycan (LPG) between patients with localized and diffuse cutaneous leishmaniasis through gene expression profiling. Our results indicate that important genes involved in the innate immune response to Leishmania are down-regulated in NK cells from DCL patients, particularly TLR and JAK/STAT signaling pathways. This down-regulation showed to be independent of LPG stimulation. The study sheds new light for understanding the mechanisms that undermine the correct effector functions of NK cells in patients with diffuse cutaneous leishmaniasis contributing to a better understanding of the pathobiology of leishmaniasis.     

TMEM45A Is Dispensable for Epidermal Morphogenesis,Keratinization and Barrier Formation

TMEM45A gene encodes an initially uncharacterized predicted transmembrane protein. We previously showed that this gene is highly expressed in keratinocytes where its expression correlates with keratinization, suggesting a role in normal epidermal physiology. To test this hypothesis, we generated TMEM45A knockout mice and found that these mice develop without any evident phenotype. The morphology of the epidermis assessed by histology and by labelling differentiation markers in immunofluorescence was not altered. Toluidine blue permeability assay showed that the epidermal barrier develops normally during embryonic development. We also showed that depletion of TMEM45A in human keratinocytes does not alter their potential to form in vitro 3D-reconstructed epidermis. Indeed, epidermis with normal morphogenesis were generated from TMEM45A-silenced keratinocytes. Their expression of differentiation markers quantified by RT-qPCR and evidenced by immunofluorescence labelling as well as their barrier function estimated by Lucifer yellow permeability were similar to the control epidermis. In summary, TMEM45A gene expression is dispensable for epidermal morphogenesis, keratinization and barrier formation. If this protein plays a role in the epidermis, its experimental depletion can possibly be compensated by other proteins in the two experimental models analyzed in this study.