Neomarica sergipensis sp. nov. (Trimezieae–Iridaceae) from the Brazilian Atlantic Forest

A new species of Neomarica from the Brazilian Atlantic Forest is described and illustrated. Neomarica sergipensis A. Gil & M.C.E. Amaral sp. nov. (Trimezieae, Iridoideae, Iridaceae) grows in restinga forests (forests on the coastal plains dominated by sandy soils) in the state of Sergipe, northeastern Brazil. The new species resembles Neomarica northiana, especially due to the white flowers with the basal third of the tepals yellow and covered by brown maculae, but differs by the inner tepals of with the center of the apical third is tinted violet to purple, yellow stigmatic crests with fimbriate to erose margins, and the endemic occurrence in Sergipe.     

Purification and characterization of the sopB gene product which is responsible for stable maintenance of mini-F plasmid

Summary The mini-F plasmid has the trans-acting sopA, sopB genes and the cis-acting sopC DNA which are essential for plasmid partitioning. In this paper, we report the purification of the sopB gene product from extracts of cells harboring a pBR322 derivative carrying the sopB gene. The purity of the final preparation was more than 95%, as determined by densitometry. The amino acid sequence of the amino-terminal region of the protein for the 17 residues identified was identical to that predicted from the DNA sequence of the sopB gene. Therefore, it was concluded that the protein was the sopB gene product. Using anti-SopB serum, the SopB protein was detected in the cell lysates of F⁺, F, and Hfr strains. The SopB protein bound to the plasmid DNA of a pBR322 derivative carrying the sopC DNA segment, but not to the vector plasmid pBR322.     

Involvement of DnaK protein in mini-F plasmid replication: temperature-sensitive seg mutations are located in the dnaK gene

Summary The seg mutants (seg-1 and seg-2) of Escherichia coli cannot support the replication of the F factor and mini-F plasmids at 42°C. We cloned the wild-type E. coli chromosomal DNA fragment complementing the seg-1 and seg-2 mutations and found that both mutations were complemented by the wild-type dnaK gene coding for a heat shock protein. Transduction with phage P1 indicated that the seg-2 mutation is located at about 0.3 min in the region containing the dnaK gene in the order trpR-thrA-seg-2-leuB, consistent with the locus of the dnaK gene. Cloning and sequencing of the dnaK gene of the seg mutants showed that there was one base substitution within the dnaK gene in each mutant causing an amino acid substitution. These results indicate that the seg gene in which the seg-1 and seg-2 mutations occurred is identical to the dnaK gene. The mini-F plasmid pXX325 did not transform a dnaK null mutant to ampicillin resistance at 30°C in contrast to plasmids pBR322, pACYC184 and pSC101, which did. The active dnaK (seg) gene product is therefore essential for replication of the mini-F plasmid at both 30° and 42°C.     

On the presence of a heme-binding domain homologous to cytochrome b(5) in Neurospora crassa assimilatory nitrate reductase

Assimilatory nitrate reductase has been purified with 55% recovery from a Neurospora crassa nmr-1 nit-6 mutant, using a modification of a published procedure. It possesses one heme per 240 000 g, and subunits of mol. wt. 68 000. Upon digestion with chymotrypsin, a heme-binding domain was isolated by gel filtration; its visible spectrum was highly similar to that of cytochrome b₅. On SDS gels, the fraction showed two heme-containing bands of ˜10 000 and 12 5000 daltons; their amino acid composition was not very different, suggesting that they originated from the same region of the polypeptide chain. After S-carboxymethylation, the mixture of bands was submitted to cyanogen bromide cleavage, and the fragments were separated by h.p.l.c. The two largest fragments yielded an identical sequence upon automated degradation. This sequence (39 residues with some gaps) could be easily aligned with that of cytochrome b₅ starting close to the N terminus. These results are discussed in terms of the possible quaternary structure of N. crassa nitrate reductase, whose heme-binding domain proves to be another member of the family of b₅-like cytochromes.     

odd-skipped genes and lines organize the notum anterior-posterior axis using autonomous and non-autonomous mechanisms

The growth and patterning of Drosophila wing and notum primordia depend on their subdivision into progressively smaller domains by secreted signals that emanate from localized sources termed organizers. While the mechanisms that organize the wing primordium have been studied extensively, those that organize the notum are incompletely understood. The genes odd-skipped (odd), drumstick (drm), sob, and bowl comprise the odd-skipped family of C(2)H(2) zinc finger genes, which has been implicated in notum growth and patterning. Here we show that drm, Bowl, and eyegone (eyg), a gene required for notum patterning, accumulate in nested domains in the anterior notum. Ectopic drm organized the nested expression of these anterior notum genes and downregulated the expression of posterior notum genes. The cell-autonomous induction of Bowl and Eyg required bowl, while the non-autonomous effects were independent of bowl. The homeodomain protein Bar is expressed along the anterior border of the notum adjacent to cells expressing the Notch (N) ligand Delta (Dl). bowl was required to promote Bar and repress Dl expression to pattern the anterior notum in a cell-autonomous manner, while lines acted antagonistically to bowl posterior to the Bowl domain. Our data suggest that the odd-skipped genes act at the anterior notum border to organize the notum anterior-posterior (AP) axis using both autonomous and non-autonomous mechanisms.     

Effect of subinihibitory and inhibitory concentrations of Plectranthus amboinicus (Lour.) Spreng essential oil on Klebsiella pneumoniae

We evaluated the antimicrobial activity and some mechanisms used by subinhibitory and inhibitory concentrations of the essential oil, obtained from leaves of Plectranthus amboinicus, against a standard strain of Klebsiella pneumoniae and 5 multiresistant clinical isolates of the bacteria. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC), the rate of kill and the pH sensitivity of the essential oil were determined by microdilution tests performed in 96-well plates. Subinhibitory and inhibitory concentrations of the essential oil were tested in order to check its action on K. pneumoniae membrane permeability, capsule expression, urease activity and cell morphology. The MIC and MBC of the essential oil were 0.09±0.01%. A complete inhibition of the bacterial growth was observed after 2h of incubation with twice the MIC of the essential oil. A better MIC was found when neutral or alkaline pH broth was used. Alteration in membrane permeability was found by the increase of crystal violet uptake when the bacteria were incubated with twice the MIC levels of the essential oil. The urease activity could be prevented when all the subinhibitory concentrations were tested in comparison to the untreated group (p<0.001). Alteration of the bacterial morphology besides inhibition of the capsule expression was verified by atomic force microscopy, and Anthony's stain method, respectively. Our data allow us to conclude that the essential oil of P. amboinicus can be a good candidate for future research.     

Dehydroepiandrosterone increases beta-cell mass and improves the glucose-induced insulin secretion by pancreatic islets from aged rats

The effect of dehydroepiandrosterone (DHEA) on pancreatic islet function of aged rats, an animal model with impaired glucose-induced insulin secretion, was investigated. The following parameters were examined: morphological analysis of endocrine pancreata by immunohistochemistry; protein levels of insulin receptor, IRS-1, IRS-2, PI 3-kinase, Akt-1, and Akt-2; and static insulin secretion in isolated pancreatic islets. Pancreatic islets from DHEA-treated rats showed an increased beta-cell mass accompanied by increased Akt-1 protein level but reduced IR, IRS-1, and IRS-2 protein levels and enhanced glucose-stimulated insulin secretion. The present results suggest that DHEA may be a promising drug to prevent diabetes during aging.     

Comparative analysis of expression of two p97 homologues in Caenorhabditis elegans

Caenorhabditis elegans possesses two p97/VCP/Cdc48p homologues, named CDC-48.1 (C06A1.1) and CDC-48.2 (C41C4.8), although their expression regulation and functional diversity have not yet been studied. We therefore investigated spatial and temporal expression patterns of two p97 homologues in this study. RT-PCR and Western blot analysis showed that the amount of cdc-48.1 was about twofold of that of cdc-48.2 in adults and that two p97 homologues were induced by ER stress. The amount of cdc-48.1 mRNA did not increase in the cdc-48.2 deletion mutant and vice versa. In situ hybridization showed that two p97 homologues are mainly expressed in germ cells. In vivo expression analysis by using GFP translational fusion constructs revealed that CDC-48.1::GFP was expressed from embryos through to adult worms, while CDC-48.2::GFP was expressed mainly in embryos. These results suggest that the expression of two p97 homologues of C. elegans is differently regulated and independent of each other.     

Identification of a cysteine residue important for the ATPase activity of C. elegans fidgetin homologue

Based on the amino acid alignment, Caenorhabditis elegans F32D1.1 was identified to be a homologue of the mammalian fidgetin. We produced and purified the F32D1.1 protein by using a baculovirus-expression system. F32D1.1 has an ATPase activity, which is sensitive to N-ethylmaleimide. Km and Vmax for the ATPase activity of F32D1.1 were estimated to be 0.44 mM and 225 nmol/mg/min, respectively. When the cysteine at the position of 368 was mutated to alanine, the ATPase activity was greatly decreased; Vmax was decreased to one-sixth, while Km remained similar. These results suggest that the unique position of cysteine 368, located immediately downstream of the Walker A motif, plays an important role in the ATP hydrolysis process of C. elegans F32D1.1 protein.