On-Probe Sample Pretreatment for Direct Analysis of Lipids in Gram-Positive Bacterial Cells by Matrix-Assisted Laser Desorption Ionization Mass Spectrometry

On-probe sample pretreatment using trifluoroacetic acid as an additional reagent enabled the direct detection of phospholipids in whole bacteria by means of matrix-assisted laser desorption ionization mass spectrometry for not only gram-negative organisms but also gram-positive ones with a thicker peptidoglycan layer.     

Active Transport and Accumulation of Iodide by Newly Isolated Marine Bacteria

Iodide (I⁻)-accumulating bacteria were isolated from marine sediment by an autoradiographic method with radioactive ¹²⁵I⁻. When they were grown in a liquid medium containing 0.1 μM iodide, 79 to 89% of the iodide was removed from the medium, and a corresponding amount of iodide was detected in the cells. Phylogenetic analysis based on 16S rRNA gene sequences indicated that iodide-accumulating bacteria were closely related to Flexibacter aggregans NBRC15975 and Arenibacter troitsensis, members of the family Flavobacteriaceae. When one of the strains, strain C-21, was cultured with 0.1 μM iodide, the maximum iodide content and the maximum concentration factor for iodide were 220 ± 3.6 (mean ± standard deviation) pmol of iodide per mg of dry cells and 5.5 × 10³, respectively. In the presence of much higher concentrations of iodide (1 μM to 1 mM), increased iodide content but decreased concentration factor for iodide were observed. An iodide transport assay was carried out to monitor the uptake and accumulation of iodide in washed cell suspensions of iodide-accumulating bacteria. The uptake of iodide was observed only in the presence of glucose and showed substrate saturation kinetics, with an apparent affinity constant for transport and a maximum velocity of 0.073 μM and 0.55 pmol min⁻¹ mg of dry cells⁻¹, respectively. The other dominant species of iodine in terrestrial and marine environments, iodate (IO₃⁻), was not transported.     

The effects of spatial habitat structure on the evolution of density-dependent growth and reproduction in freshwater snails

We examined the growth and reproductive rates of freshwater snails, Physa acuta, in two habitat types. In the Asabata habitat, snails lived in isolated water pools, which occasionally joined to form a single large pool; in the Kakegawa habitat, they lived in a slow-running water way. Genetic structure assessments using three microsatellite loci supports the idea that a stable panmictic population occupies the Kakegawa habitat. The Asabata habitat, however, is occupied with an alternate mixing population as revealed by microsatellite data. The Asabata population might alternate between localized mating within isolated pools (as revealed by high F _IS and F _IT values) when the water levels are low and panmixia (as revealed by the low F _ST values and AMOVA analysis) when the habitat is flooded. Laboratory experiments, using snails collected from the two habitats, showed that juvenile snails grew faster, laid more eggs, and laid them earlier in the Asabata habitat than in the Kakegawa habitat. Growth rates were lower at high density than at low density in the Kakegawa habitat; the inverse was true in the Asabata habitat. Density-dependent response of individual snail reproduction was higher in the Kakagawa habitat than in the Asabata habitats. The results support the hypothesis that spatial structure affects the evolution of density-dependent growth rates and of timing for reproduction.     

Plant-pollinator interactions in tropical monsoon forests in Southeast Asia

Forests with different flora and vegetation types harbor different assemblages of flower visitors, and plant-pollinator interactions vary among forests. In monsoon-dominated East and Southeast Asia, there is a characteristic gradient in climate along latitude, creating a broad spectrum of forest types with potentially diverse pollinator communities. To detect a geographical pattern of plant-pollinator interactions, we investigated flowering phenology and pollinator assemblages in the least-studied forest type, i.e., tropical monsoon forest, in the Vientiane plain in Laos. Throughout the 5-year study, we observed 171 plant species blooming and detected flower visitors on 145 species. Flowering occurred throughout the year, although the number of flowering plant species peaked at the end of dry season. The dominant canopy trees, including Dipterocarpaceae, bloomed annually, in contrast to the supra-annual general flowering that occurs in Southeast Asian tropical rain forests. Among the 134 native plant species, 68 were pollinated by hymenopterans and others by lepidopterans, beetles, flies, or diverse insects. Among the observed bees, Xylocopa, megachilids, and honeybees mainly contributed to the pollination of canopy trees, whereas long-tongued Amegilla bees pollinated diverse perennials with long corolla tubes. This is the first community-level study of plant-pollinator interactions in an Asian tropical monsoon forest ecosystem.     

Resistance to collagen-induced arthritis in SHPS-1 mutant mice

SHPS-1 is a transmembrane protein that binds the protein tyrosine phosphatases SHP-1 and SHP-2 through its cytoplasmic region and is abundantly expressed on dendritic cells and macrophages. Here we show that mice expressing a mutant form of SHPS-1 fail to develop type-II collagen (CII)-induced arthritis (CIA), a model for rheumatoid arthritis in humans. Histological examinations of the arthritic paws from immunized wild-type mice revealed that cartilage was destroyed in association with marked mononuclear cell infiltration, while only mild cell infiltration was observed in immunized SHPS-1 mutant mice. Consistently, the serum levels of both IgG and IgG2a specific to CII and of IL-1β in immunized SHPS-1 mutant mice were markedly reduced compared with those apparent for wild-type mice. The CII-induced proliferation of, and production of cytokines by, T cells from immunized SHPS-1 mutant mice were reduced compared to wild-type cells. These results suggest that SHPS-1 is essential for development of CIA.     

ERRgamma tethers strongly bisphenol A and 4-alpha-cumylphenol in an induced-fit manner

A receptor-binding assay and X-ray crystal structure analysis demonstrated that the endocrine disruptor bisphenol A (BPA) strongly binds to human estrogen-related receptor γ (ERRγ). BPA is well anchored to the ligand-binding pocket, forming hydrogen bonds with its two phenol-hydroxyl groups. In this study, we found that 4-α-cumylphenol lacking one of its phenol-hydroxyl groups also binds to ERRγ very strongly. The 2.0 Å crystal structure of the 4-α-cumylphenol/ERRγ complex clearly revealed that ERRγ’s Leu345-β-isopropyl plays a role in the tight binding of 4-α-cumylphenol and BPA, rotating in a back-and-forth induced-fit manner.     

Functional dissection of transmembrane domains of human TAP-like (ABCB9)

An ABC transporter, TAP-Like (TAPL), was dissected into its amino-terminal transmembrane domain and the following core domain. When these domains were transiently expressed as tagged proteins with a His6- or Myc-epitope tag, the amino-terminal ones (Met¹-Lys¹⁸²) could not associate with each other, or with the full-length transporter (Met¹-Ala⁷⁶⁶). However, both the core domain (Arg¹⁴¹-Ala⁷⁶⁶) and full-length protein mutually interacted. The amino-terminal domain (Met¹-Arg¹⁴¹) as well as the full-length transporter fused with fluorescent protein GFP was sorted to lysosomal membranes upon their stable expression, as visualized by means of fluorescent microscopy, while the core domain (Arg¹⁴¹-Ala⁷⁶⁶) was broadly distributed in the intra-cellular membranes. These results suggest that the sorting signal for lysosomes is present within the amino-terminal transmembrane domain (Met¹-Arg¹⁴¹) of the TAPL molecule.     

Structural and functional associations of apical junctions with cytoskeleton

Actin dynamics play multiple roles in promoting cell movement, changing cell shapes, and establishing intercellular adhesion. Cell contact events are involved in tissue morphogenesis, immune responses, and cancer cell invasion. In epithelial cells, cell-cell contacts mature to form apical junctions with which the actin cytoskeleton physically associates. Living cell imaging shows, however, that the apical junctional complex is less dynamically regulated than the actin cytoskeleton, indicating that their interaction does not remain stable. Given that several cell adhesion modules are clustered at apical junctions, the sum of weak or transient interactions may create linkages that can be strong yet easily remodeled. Here we describe how subcellular protein interactions are coordinated to induce changes in actin organization and dynamics, in response to the status of apical junctions.     

Mesenchymal stem cells are recruited into wounded skin and contribute to wound repair by transdifferentiation into multiple skin cell type

Mesenchymal stem cells (MSCs) can differentiate not only into mesenchymal lineage cells but also into various other cell lineages. As MSCs can easily be isolated from bone marrow, they can be used in various tissue engineering strategies. In this study, we assessed whether MSCs can differentiate into multiple skin cell types including keratinocytes and contribute to wound repair. First, we found keratin 14-positive cells, presumed to be keratinocytes that transdifferentiated from MSCs in vitro. Next, we assessed whether MSCs can transdifferentiate into multiple skin cell types in vivo. At sites of mouse wounds that had been i.v. injected with MSCs derived from GFP transgenic mice, we detected GFP-positive cells associated with specific markers for keratinocytes, endothelial cells, and pericytes. Because MSCs are predominantly located in bone marrow, we investigated the main MSC recruitment mechanism. MSCs expressed several chemokine receptors; especially CCR7, which is a receptor of SLC/CCL21, that enhanced MSC migration. Finally, MSC-injected mice underwent rapid wound repaired. Furthermore, intradermal injection of SLC/CCL21 increased the migration of MSCs, which resulted in an even greater acceleration of wound repair. Taken together, we have demonstrated that MSCs contribute to wound repair via processes involving MSCs differentiation various cell components of the skin.