Paralcaligenes ureilyticus gen. nov., sp. nov. isolated from soil of a Korean ginseng field

A bacterial strain, designated GR24-5^T, was isolated from soil cultivated with Korean ginseng. Cells were Gram-negative, strictly aerobic, catalase- and oxidase-positive, non-spore-forming motile rods. Based on the 16S rRNA gene sequence, strain GR24-5^T could be assigned to the family Alcaligenaceae. Strain GR24-5^T showed the highest sequence similarities with Parapusillimonas granuli Ch07^T (97.1%), Pusillimonas noertemannii BN9^T (96.9%), Pigmentiphaga kullae DSM 13608^T (96.5%), and Castellaniella defragrans 54Pin^T (96.3%). Strain GR24-5^T demonstrated a low DNA-DNA relatedness (23%) with P. granuli Ch07^T. The major respiratory quinone is ubiquinone 8 (Q-8) and the major fatty acids are C_16:0, C_17:0 cyclo, and summed feature 1 (C_14:0 3-OH/iso-C_16:1 I/C_12:0 aide). Putrescine, spermidine, and 2-hydroxyputrescine are the major polyamines. The major polar lipids are phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylglycerol, di-phosphatidylglycerol, and an unknown aminophospholipid. Polar lipid patterns of strain GR24-5^T were unique in having a large amount of phosphatidylmethylethanolamine. Based on phylogenetic analysis and physiological and biochemical characteristics, strain GR245^T represents a novel genus and species, for which the name Paralcaligenes ureilyticus gen. nov., sp. nov. is proposed. The type strain of P. aralcaligenes ureilyticus is GR24-5^T (=KACC 13888 =DSM 24591^T).     

Comparison of hair dermal cells and skin fibroblasts in a collagen sponge for use in wound repair

The adult hair follicle has well-defined dermal and epithelial populations that display distinct developmental properties. The follicular dermal cells, namely the dermal papilla and dermal sheath, are derived from the same mesenchymal cells as dermal fibroblasts and therefore, we believed that follicular cells could be useful sources of interfollicular keratinocytes and fibroblast for skin wound repair. In this study, we evaluated the relative effect of various mesenchymal-derived cells on wound healing following skin injury. Human dermal cells, including two different follicular dermal cells and skin fibroblasts were cultured in collagen sponges and compared with respect to wound healing. Results indicated that there was no significant difference in wound contraction and angiogenesis among the cell types. Further, dermal sheath cells exhibited relatively poor results compared with other cells in new collagen synthesis. Finally, basement membrane reformation and new collagen synthesis for the dermal papilla cell grafts was superior to those of the dermal sheath cells or fibroblasts.     

Gene cloning and expression of a 3-ketovalidoxylamine C-N-lyase from <Emphasis Type="Italic">Flavobacterium saccharophilum</Emphasis> IFO 13984

In this study, we report the first functional cloning and heterogeneous expression of 3-ketovalidoxylamine C-N lyase (E.C. 4.3.3.1) from Flavobacterium saccharophilum IFO 13984. This gene is 1,098 bp in length and encodes a peptide of 366 amino acids. The recombinant C-N lyase was successfully overexpressed in E. coli, and its functional activity, degradation of 3-ketovalidoxylamine A, was confirmed by HPLC analysis. The sequence and phylogenetic analysis showed that the C-N lyase has no similarity with other amine lyases (E.C. 4.3.3) but has similarity with the conserved domain present in SusD and RagB. Thus, the C-N lyase may have a similar binding domain for sugar moieties with SusD/RagB. This genetic information may lead to improvements in C-N lyase function for industrial applications.     

A large population genetic study of 15 autosomal short tandem repeat loci for establishment of Korean DNA Profile Database

Genotyping of highly polymorphic short tandem repeat (STR) markers is widely used for the genetic identification of individuals in forensic DNA analyses and in paternity disputes. The National DNA Profile Databank recently established by the DNA Identification Act in Korea contains the computerized STR DNA profiles of individuals convicted of crimes. For the establishment of a large autosomal STR loci population database, 1805 samples were obtained at random from Korean individuals and 15 autosomal STR markers were analyzed using the AmpFlSTR Identifiler PCR Amplification kit. For the 15 autosomal STR markers, no deviations from the Hardy-Weinberg equilibrium were observed. The most informative locus in our data set was the D2S1338 with a discrimination power of 0.9699. The combined matching probability was 1.521 × 10⁻¹⁷. This large STR profile dataset including atypical alleles will be important for the establishment of the Korean DNA database and for forensic applications.     

Synthesis of novel difluoro-cyclopropyl guanine nucleosides and their phosphonate analogues as potent antiviral agents

The synthesis of new rigid guanine analogues with anti-HIV-1 and anti-herpes viral activities is described. The phosphonate of difluorocyclopropane nucleoside analogue 26 exhibits in vitro anti-HIV-1 activity similar to that of PMEA in MT-4 cells. Further, analogue 20 shows moderate anti-HCMV activity in MRC cells.     

Discovery of parallel pathways of kanamycin biosynthesis allows antibiotic manipulation

Kanamycin is one of the most widely used antibiotics, yet its biosynthetic pathway remains unclear. Current proposals suggest that the kanamycin biosynthetic products are linearly related via single enzymatic transformations. To explore this system, we have reconstructed the entire biosynthetic pathway through the heterologous expression of combinations of putative biosynthetic genes from Streptomyces kanamyceticus in the non-aminoglycoside-producing Streptomyces venezuelae. Unexpectedly, we discovered that the biosynthetic pathway contains an early branch point, governed by the substrate promiscuity of a glycosyltransferase, that leads to the formation of two parallel pathways in which early intermediates are further modified. Glycosyltransferase exchange can alter flux through these two parallel pathways, and the addition of other biosynthetic enzymes can be used to synthesize known and new highly active antibiotics. These results complete our understanding of kanamycin biosynthesis and demonstrate the potential of pathway engineering for direct in vivo production of clinically useful antibiotics and more robust aminoglycosides.     

Signaling role of fructose mediated by FINS1/FBP in Arabidopsis thaliana

Sugars are evolutionarily conserved signaling molecules that regulate the growth and development of both unicellular and multicellular organisms. As sugar-producing photosynthetic organisms, plants utilize glucose as one of their major signaling molecules. However, the details of other sugar signaling molecules and their regulatory factors have remained elusive, due to the complexity of the metabolite and hormone interactions that control physiological and developmental programs in plants. We combined information from a gain-of-function cell-based screen and a loss-of-function reverse-genetic analysis to demonstrate that fructose acts as a signaling molecule in Arabidopsis thaliana. Fructose signaling induced seedling developmental arrest and interacted with plant stress hormone signaling in a manner similar to that of glucose. For fructose signaling responses, the plant glucose sensor HEXOKINASE1 (HXK1) was dispensable, while FRUCTOSE INSENSITIVE1 (FINS1), a putative FRUCTOSE-1,6-BISPHOSPHATASE, played a crucial role. Interestingly, FINS1 function in fructose signaling appeared to be independent of its catalytic activity in sugar metabolism. Genetic analysis further indicated that FINS1-dependent fructose signaling may act downstream of the abscisic acid pathway, in spite of the fact that HXK1-dependent glucose signaling works upstream of hormone synthesis. Our findings revealed that multiple layers of controls by fructose, glucose, and abscisic acid finely tune the plant autotrophic transition and modulate early seedling establishment after seed germination.     

Dual roles for Rac2 in neutrophil motility and active retention in zebrafish hematopoietic tissue

Neutrophil homeostasis is essential for host defense. Here we identify dual roles for Rac2 during neutrophil homeostasis using a zebrafish model of primary immune deficiency induced by the human inhibitory Rac2D57N mutation in neutrophils. Noninvasive live imaging of Rac2 morphants or Rac2D57N zebrafish larvae demonstrates an essential role for Rac2 in regulating 3D motility and the polarization of F-actin dynamics and PI(3)K signaling in?vivo. Tracking of photolabeled Rac2-deficient neutrophils from hematopoietic tissue also shows increased mobilization into the circulation, indicating that neutrophil mobilization does not require traditionally defined cell motility. Moreover, excessive neutrophil retention in hematopoietic tissue resulting from a constitutively active CXCR4 mutation in zebrafish warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome is partially rescued by the inhibitory Rac2 mutation. These findings reveal that Rac2 signaling is necessary for both neutrophil 3D motility and CXCR4-mediated neutrophil retention in hematopoietic tissue, thereby limiting neutrophil mobilization, a critical first step in the innate immune response.