Yif1B Is Involved in the Anterograde Traffic Pathway and the Golgi Architecture

Yif1B is an intracellular membrane‐bound protein belonging to the Yip family, shown previously to control serotonin 5‐HT_1A receptor targeting to dendrites. Because some Yip proteins are involved in the intracellular traffic between the ER and the Golgi, here we investigated the precise localization of Yif1B in HeLa cells. We found that Yif1B is not resident into the Golgi, but rather belongs to the IC compartment. After analyzing the role of Yif1B in protein transport, we showed that the traffic of the VSVG protein marker was accelerated in Yif1B depleted HeLa cells, as well as in hippocampal neurons from Yif1B KO mice. Conversely, Yif1B depletion in HeLa cells did not change the retrograde traffic of ShTx. Interestingly, in long term depletion of Yif1B as in Yif1B KO mice, we observed a disorganized Golgi architecture in CA1 pyramidal hippocampal neurons, which was confirmed by electron microscopy. However, because short term depletion of Yif1B did not change Golgi structure, it is likely that the implication of Yif1B in anterograde traffic does not rely on its role in structural organization of the Golgi apparatus, but rather on its shuttling between the ER, the IC and the Golgi compartments.      

Long‐term impacts of manure amendments on carbon and greenhouse gas dynamics of rangelands

Livestock manure is applied to rangelands as an organic fertilizer to stimulate forage production, but the long‐term impacts of this practice on soil carbon (C) and greenhouse gas (GHG) dynamics are poorly known. We collected soil samples from manured and nonmanured fields on commercial dairies and found that manure amendments increased soil C stocks by 19.0 ± 7.3 Mg C ha^?1 and N stocks by 1.94 ± 0.63 Mg N ha^?1 compared to nonmanured fields (0–20 cm depth). Long‐term historical (1700–present) and future (present–2100) impacts of management on soil C and N dynamics, net primary productivity (NPP), and GHG emissions were modeled with DayCent. Modeled total soil C and N stocks increased with the onset of dairying. Nitrous oxide (N₂O) emissions also increased by ~2 kg N₂O‐N ha^?1 yr^?1. These emissions were proportional to total N additions and offset 75–100% of soil C sequestration. All fields were small net methane (CH₄) sinks, averaging ?4.7 ± 1.2 kg CH₄‐C ha^?1 yr^?1. Overall, manured fields were net GHG sinks between 1954 and 2011 (?0.74 ± 0.73 Mg CO₂ e ha^?1 yr^?1, CO₂e are carbon dioxide equivalents), whereas nonmanured fields varied around zero. Future soil C pools stabilized 40–60 years faster in manured fields than nonmanured fields, at which point manured fields were significantly larger sources than nonmanured fields (1.45 ± 0.52 Mg CO₂e ha^?1 yr^?1 and 0.51 ± 0.60 Mg CO₂e ha^?1 yr^?1, respectively). Modeling also revealed a large background loss of soil C from the passive soil pool associated with the shift from perennial to annual grasses, equivalent to 29.4 ± 1.47 Tg CO₂e in California between 1820 and 2011. Manure applications increased NPP and soil C storage, but plant community changes and GHG emissions decreased, and eventually eliminated, the net climate benefit of this practice.     

Dissecting Arabidopsis Gβ signal transduction on the protein surface

The heterotrimeric G-protein complex provides signal amplification and target specificity. The Arabidopsis (Arabidopsis thaliana) Gβ-subunit of this complex (AGB1) interacts with and modulates the activity of target cytoplasmic proteins. This specificity resides in the structure of the interface between AGB1 and its targets. Important surface residues of AGB1, which were deduced from a comparative evolutionary approach, were mutated to dissect AGB1-dependent physiological functions. Analysis of the capacity of these mutants to complement well-established phenotypes of Gβ-null mutants revealed AGB1 residues critical for specific AGB1-mediated biological processes, including growth architecture, pathogen resistance, stomata-mediated leaf-air gas exchange, and possibly photosynthesis. These findings provide promising new avenues to direct the finely tuned engineering of crop yield and traits.     

Somatic embryogenesis from different tissues of Spanish populations of maritime pine

The somatic embryogenesis (SE) capacity of megagametophytes belonging to Continental and Mediterranean Spanish provenances of maritime pine (Pinus pinaster Aiton) was studied, noting factors (megagametophyte developmental stage and culture medium) that enhanced the induction and establishment of SE lines. In both provenances, initiation and establishment of embryogenic calli was higher on megagametophytes in which the dominant zygotic embryo had begun to develop. In the Mediterranean provenance, however, SE lines were also established from megagametophytes enclosing zygotic embryos with well-developed cotyledons. A modified Litvay medium (mLV) containing 9.9???M 2,4-dichlorophenoxyacetic acid (2,4-D) and 4.4???M 6-benzyladenine (BA) was superior to DCR medium containing 13.6???M 2,4-D and 4.4???M BA for SE induction, but there were no differences between media in terms of the number of SE lines established after 4?months in culture (153 vs. 155 established SE lines, for mLV and DCR media respectively). Of the 26 embryogenic lines tested for maturation, 15 (58?%) produced cotyledonary somatic embryos and 75?% of these gave rise to plants on germination medium. SE-like cultures from adult maritime pine trees were also initiated, but embryogenic lines could not be established. This is the first report on the production of SE in maritime pine of Continental and Mediterranean origin. The micropropagation protocols presented here provide an important tool for the vegetative multiplication of selected families and breeding programs for maritime pines from Spain.     

Separate pathways for O acetylation of polymeric and monomeric sialic acids and identification of sialyl O-acetyl esterase in Escherichia coli K1

O acetylation at carbon positions 7 or 9 of the sialic acid residues in the polysialic acid capsule of Escherichia coli K1 is catalyzed by a phase-variable contingency locus, neuO, carried by the K1-specific prophage, CUS-3. Here we describe a novel method for analyzing polymeric sialic acid O acetylation that involves the release of surface sialic acids by endo-N-acetylneuraminidase digestion, followed by fluorescent labeling and detection of quinoxalinone derivatives by chromatography. The results indicated that NeuO is responsible for the majority of capsule modification that takes place in vivo. However, a minor neuO-independent O acetylation pathway was detected that is dependent on the bifunctional polypeptide encoded by neuD. This pathway involves O acetylation of monomeric sialic acid and is regulated by another bifunctional enzyme, NeuA, which includes N-terminal synthetase and C-terminal sialyl O-esterase domains. A homologue of the NeuA C-terminal domain (Pm1710) in Pasteurella multocida was also shown to be an esterase, suggesting that it functions in the catabolism of acetylated environmental sialic acids. Our combined results indicate a previously unexpected complexity in the synthesis and catabolism of microbial sialic and polysialic acids. These findings are key to understanding the biological functions of modified sialic acids in E. coli K1 and other species and may provide new targets for drug or vaccine development.     

Methods to control ectomycorrhizal colonization: effectiveness of chemical and physical barriers

We conducted greenhouse experiments using Douglas-fir (Pseudotsuga menziesii var. glauca) seedlings where chemical methods (fungicides) were used to prevent ectomycorrhizal colonization of single seedlings or physical methods (mesh barriers) were used to prevent formation of mycorrhizal connections between neighboring seedlings. These methods were chosen for their ease of application in the field. We applied the fungicides, Topas (nonspecific) and Senator (ascomycete specific), separately and in combination at different concentrations and application frequencies to seedlings grown in unsterilized forest soils. Additionally, we assessed the ability of hyphae to penetrate mesh barriers of various pore sizes (0.2, 1, 20, and 500 microm) to form mycorrhizas on roots of neighboring seedlings. Ectomycorrhizal colonization was reduced by approximately 55% with the application of Topas at 0.5 g l(-1). Meshes with pore sizes of 0.2 and 1 microm were effective in preventing the formation of mycorrhizas via hyphal growth across the mesh barriers. Hence, meshes in this range of pore sizes could also be used to prevent the formation of common mycorrhizal networks in the field. Depending on the ecological question of interest, Topas or the employment of mesh with pore sizes <1 microm are suitable for restricting mycorrhization in the field.     

Neurological and physiological disorders in Artemia harboring manipulative cestodes

There are many impressive examples of host manipulation by parasites, but mechanisms underlying these ethological changes, as well as their physiological consequences, are not well characterized. Here, we analyzed part of the cerebral proteome of brine shrimp Artemia infected by manipulative cestodes, using for the first time the ProteinChip Surface-Enhanced Laser Desorption Ionization and Time of Fly Mass Spectrometry (SELDI-TOF MS) system, which has been proposed as an excellent way to analyze the host genome during the host-parasite interaction processes. We found 2 peptides downregulated in individuals infected by the dilepidid, Anomotaenia tringae (4.5 kDa), and by the 2 hymenolepidids, Flamingolepis liguloides and Confluaria podicipina (3.9 kDa), which are potential candidates for involvement with the manipulation process. The identification of 2 head peptides (4.1 and 4.2 kDa) overexpressed in all the categories in brine shrimp living at the surface (both infected individuals and uninfected controls) suggests its association with the different environmental conditions experienced at the water surface. In parallel, brine shrimp infected by C. podicipina showed significant values of triglycerides, potentially augmenting their profitability and attractiveness for the predaceous definitive host (grebes). We discuss our findings in relationship with current ideas on the complexity of parasitically modified organisms.     

Synthesis and evaluation of imidazole–dioxolane compounds as selective heme oxygenase inhibitors: Effect of substituents at the 4-position of the dioxolane ring

Several imidazole–dioxolane compounds were synthesized and evaluated as novel inhibitors of heme oxygenase (HO). These compounds, which include a series of substituted thiophenol and substituted phenol derivatives of (2R,4S)-2-[2-(4-chlorophenyl)ethyl]-2-[(1H-imidazol-1-yl)methyl]-4-[(phenylsulfanyl)methyl]-1,3-dioxolane hydrochloride (3), in addition to smaller functionalized derivatives, continue our structure–activity studies by exploration of the aminothiophenol region (‘northeastern region’) in our original target structure azalanstat (1). In vitro, most of the compounds in this series were found to be highly potent inhibitors of the stress-induced isozyme HO-1 and the constitutive isozyme HO-2, showing only moderate selectivity for HO-1. Nevertheless, a few of the compounds displayed higher selectivity toward HO-1. None of the compounds having a larger appendage in the northeastern region were inhibitors of CYP2E1, whereas a compound having a relatively small fluorine substituent in this region did inhibit CYP2E1; all of the compounds tested exhibited high inhibitory potency against CYP3A1/3A2.