A novel antifungal analog peptide derived from protaetiamycine

Previously, the 9-mer analog peptides, 9Pbw2 and 9Pbw4, were designed based on a defensin-like peptide, protaetiamycine isolated from Protaetia brevitarsis. In this study, antifungal effects of the analog peptides were investigated. The antifungal susceptibility testing exhibited that 9Pbw4 contained more potent antifungal activities than 9Pbw2. A PI influx assay confirmed the effects of the analog peptides and demonstrated that the peptides exerted their activity by a membrane-active mechanism, in an energy-independent manner. As the noteworthy potency of 9Pbw4, the mechanism(s) of 9Pbw4 were further investigated. The membrane studies, using rhodamine-labeled giant unilamellar vesicle (GUV) and fluorescein isothiocyanate (FITC)-dextran loaded liposome, suggested that the membrane-active mechanism of 9Pbw4 could have originated from the poreforming action and the radii of pores was presumed to be anywhere from 1.8 nm to 3.3 nm. These results were confirmed by 3D-flow cytometric contour-plot analysis. The present study suggests a potential of 9Pbw4 as a novel antifungal peptide.     

Overexpression of the downward leaf curling (DLC) gene from melon changes leaf morphology by controlling cell size and shape in Arabidopsis leaves

A plant-specific gene was cloned from melon fruit. This gene was named downward leaf curling (CmDLC) based on the phenotype of transgenic Arabidopsis plants overexpressing the gene. This expression level of this gene was especially upregulated during melon fruit enlargement. Overexpression of CmDLC in Arabidopsis resulted in dwarfism and narrow, epinastically curled leaves. These phenotypes were found to be caused by a reduction in cell number and cell size on the adaxial and abaxial sides of the epidermis, with a greater reduction on the abaxial side of the leaves. These phenotypic characteristics, combined with the more wavy morphology of epidermal cells in overexpression lines, indicate that CmDLC overexpression affects cell elongation and cell morphology. To investigate intracellular protein localization, a CmDLC-GFP fusion protein was made and expressed in onion epidermal cells. This protein was observed to be preferentially localized close to the cell membrane. Thus, we report here a new plant-specific gene that is localized to the cell membrane and that controls leaf cell number, size and morphology.     

Investigating the adaptive model of thermal comfort for naturally ventilated school buildings in Taiwan

Divergence in the acceptability to people in different regions of naturally ventilated thermal environments raises a concern over the extent to which the ASHRAE Standard 55 may be applied as a universal criterion of thermal comfort. In this study, the ASHRAE 55 adaptive model of thermal comfort was investigated for its applicability to a hot and humid climate through a long-term field survey performed in central Taiwan among local students attending 14 elementary and high schools during September to January. Adaptive behaviors, thermal neutrality, and thermal comfort zones are explored. A probit analysis of thermal acceptability responses from students was performed in place of the conventional linear regression of thermal sensation votes against operative temperature to investigate the limits of comfort zones for 90% and 80% acceptability; the corresponding comfort zones were found to occur at 20.1–28.4°C and 17.6–30.0°C, respectively. In comparison with the yearly comfort zones recommended by the adaptive model for naturally ventilated spaces in the ASHRAE Standard 55, those observed in this study differ in the lower limit for 80% acceptability, with the observed level being 1.7°C lower than the ASHRAE-recommended value. These findings can be generalized to the population of school children, thus providing information that can supplement ASHRAE Standard 55 in evaluating the thermal performance of naturally ventilated school buildings, particularly in hot-humid areas such as Taiwan.     

Heat-Resistant Agglutinin 1 Is an Accessory Enteroaggregative Escherichia coli Colonization Factor

Enteroaggregative Escherichia coli (EAEC) is an important cause of acute and persistent diarrhea. The defining stacked brick adherence pattern of Peruvian EAEC isolate 042 has previously been attributed to aggregative adherence fimbriae II (AAF/II), which confer aggregative adherence on laboratory E. coli strains. EAEC strains also show exceptional autoaggregation and biofilm formation, other phenotypes that have hitherto been ascribed to AAF/II. We report that EAEC 042 carries the heat-resistant agglutinin (hra1) gene, also known as hek, which encodes an outer membrane protein. Like AAF/II, the cloned EAEC 042 hra1 gene product is sufficient to confer autoaggregation, biofilm formation, and aggregative adherence on nonadherent and nonpathogenic laboratory E. coli strains. However, an 042 hra1 deletion mutant is not deficient in these phenotypes compared to the wild type. EAEC strain 042 produces a classic honeycomb or stacked brick pattern of adherence to epithelial cells. Unlike wild-type 042, the hra1 mutant typically does not form a tidy stacked brick pattern on HEp-2 cells in culture, which is definitive for EAEC. Moreover, the hra1 mutant is significantly impaired in the Caenorhabditis elegans slow kill colonization model. Our data suggest that the exceptional colonization of strain 042 is due to multiple factors and that Hra1 is an accessory EAEC colonization factor.Enteroaggregative Escherichia coli (EAEC) was originally identified as the etiologic agent of persistent diarrhea in developing countries but is gaining increasing prominence for its role in a wider spectrum of diarrheal syndromes. EAEC strains have been implicated in acute as well as persistent diarrhea among adults and children (reviewed in references 25 and 40). A recent meta-analysis found that EAEC is significantly associated with disease in every group at high risk for diarrhea, including young children, human immunodeficiency virus-positive individuals, and visitors to developing countries (24). In addition to its association with disease in epidemiological studies in developing countries, EAEC has also been identified as a principal cause of diarrheal disease in Germany, the United Kingdom, and the United States (11, 26, 51).Aggregative adherence is the defining characteristic of EAEC (38). EAEC strains adhere to the intestinal epithelium, and to epithelial cells in culture, in a characteristic two-dimensional “stacked brick” fashion. The pattern features bacteria adhering to the eukaryotic surface, other bacteria, and the solid substratum. Four types of fimbriae have so far been documented as conferring aggregative adherence (4, 14, 17, 37). Two noncontiguous plasmid loci containing the complete complement of genes encoding aggregative adherence fimbriae I (AAF/I) or AAF/II are sufficient to confer aggregative adherence on nonadherent E. coli (14, 49). The plasmid bearing type IV pili found in Serbian EAEC outbreak strain C1096 are also sufficient to confer a weak aggregative adherence phenotype on E. coli K-12 (17). AAF additionally play an essential role in production of a superfluous EAEC-associated biofilm, which could account for the association of these strains with persistent diarrhea in epidemiological studies (46).Some categories of diarrheagenic pathogens have a conserved set of adhesins which allow them to overcome flushing across the intestinal epithelium. Typical enteropathogenic E. coli isolates, for example, all possess bundle-forming pili and the outer membrane adhesin intimin, whereas atypical enteropathogenic E. coli isolates possess intimin but not bundle-forming pili (reviewed in reference 10). EAEC strains, by contrast, are considerably heterogeneous. While many EAEC strains carry genes encoding one of the known aggregative adherence fimbriae, some EAEC do not harbor any known AAF even though they do demonstrate aggregative adherence (4, 7, 13, 14). This, and the presence of multiple adhesins in most mucosal colonizers (53), points to the likelihood of other EAEC adhesins. Imuta et al. recently implicated a TolC secreted factor in adherence (27), and Montiero-Neto et al. (33) described a 58-kDa nonstructural adhesin in O111:H12 EAEC. However, the former factor is only a contributor to aggregative adherence and the latter adhesin is not found in other EAEC. Overall, nonstructural EAEC adhesins have received little attention.The outer membrane protein Tia was originally characterized as an invasin and later shown to confer adhesive properties on enterotoxigenic E. coli (ETEC) (20, 21). Fleckenstein et al. (21) observed that a tia gene probe hybridized to DNA from non-ETEC strains, one of which was EAEC strain 042. As the Southern blot data published by Fleckenstein et al. showed bands of different intensities, as well as size, between ETEC strain {"type":"entrez-nucleotide","attrs":{"text":"H10407","term_id":"875229","term_text":"H10407"}}H10407, which carries tia, and EAEC strain 042, we hypothesized that the probe was recognizing a similar, rather than identical, gene (21).We have determined that EAEC strain 042 harbors a gene encoding the heat-resistant agglutinin 1 (hra1), a hemagglutinin originally reported from an O9:H10:K99 porcine ETEC strain. Hra1 has also been reported from uropathogenic E. coli strains and neonatal meningitis E. coli strain RS218, in which context it is otherwise known as Hek (19, 48). (The hek nomenclature was introduced after hra1, to delineate the form of the gene found in invasive human pathogens from that of a porcine isolate [19].) A role for the outer membrane protein Hra1/Hek in adherence by neonatal meningitis E. coli has recently been defined (19).Although hra1/hek has been reported from multiple pathogens, its role in colonization and virulence has only been conclusively studied in the neonatal meningitis E. coli strain RS218 (19). In this paper, we demonstrate that the EAEC hra1 gene is sufficient to confer colonization-associated phenotypes, including aggregative adherence and biofilm formation, on laboratory E. coli strains. Intriguingly, we find that although it confers these phenotypes on K-12 and is expressed in 042, hra1 is not required for in vitro colonization-associated phenotypes demonstrated by 042. The hra1 gene is, however, essential for the formation of a true stacked brick pattern in EAEC and for optimal in vivo colonization in a Caenorhabditis elegans model.     

Expression regulation and function of Pref-1 during adipogenesis of human mesenchymal stem cells (MSCs)

Preadipocyte Factor 1 (Pref-1), also known as Delta-like Protein 1 (DLK-1) is an epidermal growth factor-like domain-containing trans-membrane protein that is involved in adipogenesis and cell fate decision. Its function in adipogenesis is reported inconsistently based on different cellular model systems. Here, by using human mesenchymal stem cells (MSCs), we show that Pref-1 is modulated by both dexamethasone and 3-isobutyl-1methylxanthine (IBMX), two components of the adipogenic induction mixture during the adipogenesis in vitro. IBMX induces the expression of Pref-1 in a time- and dose-dependent manner through cyclic AMP and cyclic GMP independent pathway and attenuates adipocyte differentiation by down-regulating PPARγ (peroxisome proliferator activated receptor gamma) expression. Dexamethasone, on the other hand, is capable of subduing the inhibitory effect of IBMX-induced Pref-1 and initiating the adipogenesis by up-regulating PPARγ expression. Moreover, the treatment of IBMX or dexamethasone alone fails to develop MSCs into mature adipocytes, however, treating cells with both IBMX and dexamethasone leads to a complete adipocyte differentiation as evaluated by lipid-droplet formation. Taken together, our study demonstrates that IBMX accelerates accumulation of lipid in MSCs only under the circumstance that the negative effect of Pref-1 induced by IBMX on the adipogenesis is overcome by dexamethasone.