Transposon mutagenesis and cloning of the genes encoding the enzymes of fengycin biosynthesis inBacillus subtilis

A total of 20Bacillus subtilis F29-3 mutants defective in fengycin biosynthesis was obtained by Tn917 mutagenesis. Cloning and mapping results showed that the transposon in these mutants was inserted in eleven different locations on the chromosome. We were able to use the chromosomal sequence adjacent to the transposon as a probe to screen for cosmid clones containing the fengycin biosynthesis genes. One of the clones obtained, pFC660, was 46 kb long. Eight transposon insertion sites were mapped within this plasmid. Among the eleven different mutants analyzed, four mutants had Tn917 inserted in regions which encoded peptide sequences similar to part of gramicidin S synthetase, surfactin synthetase, and tyrocidine synthetase. Our results suggest that fengycin is synthesized nonribosomally by the multienzyme thiotemplate mechanism.     

Pancreatic β cell identity is maintained by DNA methylation-mediated repression of Arx

Adult pancreatic β cells can replicate during growth and after injury to maintain glucose homeostasis. Here, we report that β cells deficient in Dnmt1, an enzyme that propagates DNA methylation patterns during cell division, were converted to α cells. We identified the lineage determination gene aristaless-related homeobox (Arx), as methylated and repressed in β cells, and hypomethylated and expressed in α cells and Dnmt1-deficient β cells. We show that the methylated region of the Arx locus in β cells was bound by methyl-binding protein MeCP2, which recruited PRMT6, an enzyme that methylates histone H3R2 resulting in repression of Arx. This suggests that propagation of DNA methylation during cell division also ensures recruitment of enzymatic machinery capable of modifying and transmitting histone marks. Our results reveal that propagation of DNA methylation during cell division is essential for repression of α cell lineage determination genes to maintain pancreatic β cell identity.     

Basidial development, spindle pole body, septal pore, and host-parasite-interaction in Ustilago esculenta

Smut spore-germination, spindle pole bodies, septal pores, and host-parasite interactions have been investigated in Ustilago esculenta . The species is compared with other Ustilago species parasitizing on grasses. The taxonomy of U. esculenta and the Yeniaceae is discussed.     

Optimization of formation and regeneration of protoplasts from biocontrol agents ofTrichoderma species

The optimal conditions necessary for a large yield and a high frequency of regeneration of protoplasts isolated from the biocontrol agentsTrichoderma koningii andT. harzianum were investigated. Protoplast yields were 1.2×10⁸/ml fromT. koningii and 6×10⁷/ml fromT. harzianum when 20-h mycelial culture was treated with a lytic enzyme solution containing Novozym 234 (15 mg/ml), sucrose (0.6 M) and citrate phosphate buffer (0.02 M), pH 5.6 at 31°C. When the protoplasts were grown in the regeneration medium containing yeast extract (1.5%), 1 I of Mandel's salts, pH 5.6, and glucose (0.8 M), a high frequency of regeneration of the protoplast was obseved: 66% forT. koningii and 45% forT. harzianum. Two patterns of regeneration were observed. First, the hyphae arose directly from the regenerated protoplast mother cell. Second, a chain of bud cells developed from the protoplast and subsequently generating hyphae generally protruded from the terminal bud cells.     

Biological Control of Basal Stem Rot of Chrysanthemum By Antagonists*

The basal stem rot of chrysanthemum caused by Rhizoctonia solani is an important chrysanthemum disease in Taiwan, Control of the disease using the antagonists of R.solani, including members of the genera Aspergillus, Gliocladium, Paecilomyces, Penicillhium, Tricboderma and Bacillus, was studied. The antagonists were grown in solid media or culture broths for preparations of rhe inoculums which were u.sed as soil amendments or coating materials for applications on the disease control. Sawdusts, rice husks and sawdust composts were used as supporting material to prepare the solid media. By planting of the chrysanthemums, the solid media growth with antagonists were put in the nursery sand beds, or the chrysanthemum cutting seedlings were coated with antagonists of culture broths; whilst the sand beds were inoculated with R. solani. The results indicated that all antagonists could protect the chrysanthemum from the infection of R. solani, and the effects of the solid media were higher than the culture broths. When the chrysanthemum seedlings were treated with the extracts of the antagonists, the disease could also be depressed. The different control effects on Rhizactonia disease of chrysanthemums resulted from the species of the antagonists and their delivery systems to be used.     

Skp2 is required for incretin hormone-mediated β-cell proliferation

The glucoincretin hormone glucagon-like peptide-1 (GLP-1) and its analog exendin-4 (Ex-4) promote β-cell growth and expansion. Here we report an essential role for Skp2, a substrate recognition component of SCF (Skp, Cullin, F-box) ubiquitin ligase, in promoting glucoincretin-induced β-cell proliferation by regulating the cellular abundance of p27. In vitro, GLP-1 treatment increases Skp2 levels, which accelerates p27 degradation, whereas in vivo, loss of Skp2 prevents glucoincretin-induced β-cell proliferation. Using inhibitors of phosphatidylinositol 3-kinase and Irs2 silencing RNA, we also show that the effects of GLP-1 in facilitating Skp2-dependent p27 degradation are mediated via the Irs2-phosphatidylinositol-3 kinase pathway. Finally, we show that down-regulation of p27 occurs in islets from aged mice and humans, although in these islets, age-dependent accumulation of p16(Ink4a) prevent glucoincretin-induced β-cell proliferation; however, ductal cell proliferation is maintained. Taken together, these data highlight a critical role for Skp2 in glucoincretin-induced β-cell proliferation.     

Brunneocorticium pyriforme, a new corticioid fungal genus and species belonging to the euagarics clade

Evidence derived from molecular studies in recent years has revealed that corticioid fungal genera are present in all major clades of Homobasidiomycetes. Brunneocorticium pyriforme, a corticioid fungus is proposed as a new species and placed in a new genus belonging to the euagarics clade. This fungus has been collected in subtropical-tropical Taiwan and southern Yunnan Province, China. Basidiocarps often occur on bark of living Murraya spp. (Rutaceae). Basidiocarps of B. pyriforme are resupinate with a smooth hymenial surface, a dimitic hyphal system, with nodose-septate generative hyphae and abundant yellowish brown skeletal hyphae, and leptocystidia. It has 2-sterigmate basidia and pear-shaped basidiospores. Phylogenetic analysis based on sequence data derived from LSU rDNA included Brunneocorticium in the euagarics clade of Homobasidiomycetes, allied to the agaricoid genera Marasmiellus, Campanella, etc. The molecular analysis indicated that the Brunneocorticium was independent from other corticioid genera with similar morphological features.     

Cyclin D2 is sufficient to drive β cell self-renewal and regeneration

Diabetes results from an inadequate mass of functional β cells, due to either β cell loss caused by autoimmune destruction (type I diabetes) or β cell failure in response to insulin resistance (type II diabetes). Elucidating the mechanisms that regulate β cell mass may be key to developing new techniques that foster β cell regeneration as a cellular therapy to treat diabetes. While previous studies concluded that cyclin D2 is required for postnatal β cell self-renewal in mice, it is not clear if cyclin D2 is sufficient to drive β cell self-renewal. Using transgenic mice that overexpress cyclin D2 specifically in β cells, we show that cyclin D2 overexpression increases β cell self-renewal post-weaning and results in increased β cell mass. β cells that overexpress cyclin D2 are responsive to glucose stimulation, suggesting they are functionally mature. β cells that overexpress cyclin D2 demonstrate an enhanced regenerative capacity after injury induced by streptozotocin toxicity. To understand if cyclin D2 overexpression is sufficient to drive β cell self-renewal, we generated a novel mouse model where cyclin D2 is only expressed in β cells of cyclin D2^?/? mice. Transgenic overexpression of cyclin D2 in cyclin D2^?/? β cells was sufficient to restore β cell mass, maintain normoglycaemia, and improve regenerative capacity when compared with cyclin D2^?/? littermates. Taken together, our results indicate that cyclin D2 is sufficient to regulate β cell self-renewal and that manipulation of its expression could be used to enhance β cell regeneration.