Copper(II) and zinc(II) complexes of the peptides Ac-HisValHis-NH2 and Ac-HisValGlyAsp-NH2 related to the active site of the enzyme CuZnSOD

Copper(II) and zinc(II) complexes of the peptides Ac-HisValHis-NH2 and Ac-HisValGlyAsp-NH2 related to the active site of the enzyme CuZnSOD were studied by potentiometric and spectroscopic (UV-Vis, CD and EPR) techniques. The results reveal that both ligands have effective metal binding sites, but the tripeptide is a much stronger complexing agent than the tetrapeptide. The formation of a macrochelate via the coordination of the imidazolyl residues is suggested in the copper(II)-Ac-HisValHis-NH2 system in the acidic pH range, while a 4N complex predominates at physiological pH. The interaction of Ac-HisValHis-NH2 with zinc(II) results in the formation of a precipitate indicating polynuclear complex formation. Both copper(II)-Ac-HisValHis-NH2 and copper(II)-HisValHis systems exhibit catalytic activity toward the dismutation of superoxide anion at physiological pH, but the saturated coordination sphere of the metal ions in both systems results in low reactivity as compared to the native enzyme.     

Reaction cycle of the yeast Isw2 chromatin remodeling complex

Members of the ISWI family of chromatin remodeling factors hydrolyze ATP to reposition nucleosomes along DNA. Here we show that the yeast Isw2 complex interacts with DNA in a nucleotide-dependent manner at physiological ionic strength. Isw2 efficiently binds DNA in the absence of nucleotides and in the presence of a nonhydrolyzable ATP analog. Conversely, ADP promotes the dissociation of Isw2 from DNA. In contrast, Isw2 remains bound to mononucleosomes through multiple cycles of ATP hydrolysis. Solution studies show that Isw2 undergoes nucleotide-dependent alterations in conformation not requiring ATP hydrolysis. Our results indicate that during an Isw2 remodeling reaction, hydrolysis of successive ATP molecules coincides with cycles of DNA binding, release, and rebinding involving elements of Isw2 distinct from those interacting with nucleosomes. We propose that progression of the DNA-binding site occurs while nucleosome core contacts are maintained and generates a force dissipated by disruption of histone-DNA interactions.     

Quail as the chimeric counterpart of the chicken: morphology and ontogeny of the bursa of Fabricius

The quail is the chimeric and parabiotic counterpart of the chicken, thus increasing the value of quail in the field of developmental biology. Quail bursa of Fabricius was studied by light microscopy, electron microscopy, and immunocytochemical methods. The basic cellular composition and structural framework are comparable with those of the chicken bursa. One of the major structural differences is the absence of the continuous cortico-medullary arch. In addition to the epithelial reticular cell the bursal secretory dendritic cell is the other medullary-specific bursal cell. The bursal secretory dendritic cell is a highly elongated cell which expresses vimentin intermediate filaments and produces secretory granules. The substance of the granules can be visualized by NIC2 monoclonal antibody, which was produced against guinea fowl bursal secretory dendritic cell. The released granular content appears on the lateral surface of the bursal secretory dendritic cell and is gradually solubilized. Thus, the NIC2-positive substance may occur in membrane-bound and solubilized forms in the isolated environment of the medulla. The bursal secretory dendritic cell establishes membrane contact areas with the B cells; therefore, they may influence B-cell maturation by cell contact and chemical (humoral) product. During embryogenesis bursal secretory dendritic cell precursors enter the epithelium and 1) induce epithelial bud formation, and 2) produce an NIC2-positive substance. Senescent bursal secretory dendritic cells can be phagocytic and migrate into the follicle-associated epithelium. This physiological turnover of the bursal secretory dendritic cell represents a novel pathway of macrophage formation from dendritic cells.     

S100B-Mediated Inhibition of the Phosphorylation of GFAP Is Prevented by TRTK-12

S100B belongs to a family of calcium-binding proteins involved in cell cycle and cytoskeleton regulation. We observed an inhibitory effect of S100B on glial fibrillary acidic protein (GFAP) phosphorylation, when stimulated by cAMP or Ca2+/calmodulin, in a cytoskeletal fraction from primary astrocyte cultures. We found that S100B has no direct effect on CaM KII activity, the major kinase in this cytoskeletal fraction able to phosphorylate GFAP. The inhibition of GFAP phosphorylation is most likely due to the binding of S100B to the phosphorylation sites on this protein and blocking the access of these sites to the protein kinases. This inhibition was dependent on Ca2+. However, Zn2+ could substitute for Ca2+. The inhibitory effect of S100B was prevented by TRTK-12, a peptide that blocks S100B interaction with several target proteins including glial fibrillary acidic protein. These data suggest a role for S100B in the assembly of intermediate filaments in astrocytes.     

Beta 2-agonist treatment enhances uterine oxytocin receptor mRNA expression in pregnant rats

The objective of this study was to disclose an interaction between Beta(2)-adrenergic (Beta(2)-ARs) and oxytocin (OT) receptors (OTRs) in the late-pregnant rat uterus. We investigated the level of uterine OTR mRNA expression after the administration of Beta(2)-AR agonists fenoterol and hexoprenaline to rats from day 18 to 22 of pregnancy, and also tested the effect of fenoterol on uterine explants. Hexoprenaline induced a maximum 24% increase of OTR mRNA. Fenoterol in vivo elicited a maximum 125% increase of OTR mRNA, in vitro produced a maximum fourfold increase in OTR mRNA. In fenoterol-treated rats the maximal contractility increasing effect of OT on isolated uterine rings was significantly higher than in intact term pregnant rats, but the EC50 values were not statistically different. It was concluded that the enhanced expression of OTR mRNA induced by Beta(2)-agonists in the late-pregnant rat uterus may be a possible drawback to effective therapy of preterm uterine contractions with Beta(2)-agonists.     

Adenovirus vectors targeting alphaV integrin or heparan sulfate receptors display different distribution of transgene activity after intramuscular injection

BACKGROUND: Modification of the fiber proteins in replication-deficient adenoviral (Ad) vectors through incorporation of specific receptor-binding motifs may represent a strategy to enhance their tissue targeting capabilities. METHODS: In this study, we compared an unmodified Ad (GV10) with two mutated vectors obtained by insertion of specific target sequences that redirect binding, either toward alpha(V) integrin (RGD) or heparan sulfate (UTV) cellular receptors, for reporter gene expression spatial distribution in the rabbit skeletal muscle. In a first series of experiments, injection volume was kept constant and activity of a lacZ transgene was evaluated 48 h after injection of the Ad vectors at different doses. In separate experiments, the effects of different volumes of injection at a constant dose of Ad vector were monitored. RESULTS: All vectors evaluated showed a significant increase in the number of lacZ-positive muscle segments, with increasing vector dose. However, in muscles treated with the UTV vector, fewer muscle fibers were beta-gal-positive than in GV10 or RGD vector treated animals. In fact, total beta-gal activity increased in a dose-dependent fashion in the GV10- and RGD-treated muscles, but not in the UTV-treated ones. Remarkably, in samples from UTV-treated animals, a volume-dependent enhancement of transgene expression was observed during experiments performed at the same dose and different injection volumes. CONCLUSIONS: The results of the present study demonstrate that altering Ad affinity for cellular receptors modulates the level and distribution of transgene activity, conferring characteristics that may allow for treatment customization.     

The role of repeated transurethral resection in the treatment of bladder tumor

Complete removal of the tumour or deep invasion can be proven by repeated transurethral resection of bladder wall at the previous tumour site. Six weeks after transurethral resection of bladder tumour (TURB), in all but TaG1 cases repeated resection were performed for the evaluation of radicality in 62 patients, 43 males and 19 females, suffering bladder cancer, from October 1998. In the case of positive histology another resection was performed for security reason. In the case of 38 superficial (Tis, Ta, T1) cancers, repeated resection revealed negative, identical or different T stage compared with previous histology in 28, 5 and 5 cases, respectively. In 7 cases repeated resection was applied as second intervention after the incomplete resection of large tumour mass. Indication of repeated resection was insufficient depth of resection and carcinoma in situ in 13 and 4 cases, respectively. Based on our data, we conclude that repeated resection should be performed when tumour-free status is not justified and biopsy according to Bressel was not taken.     

Identification of a new class of recombinant prolamin genes in wheat.

A novel storage protein gene with obvious [corrected] chimeric structure was isolated from an immature kernel-specific cDNA library prepared from the old Hungarian wheat [corrected] variety, Bánkúti 1201. This clone contains gamma-gliadin sequences in the 5' region and LMW-glutenin sequences on the 3' end. A frameshift mutation was also introduced by the putative recombination event. Hence, the amino acid sequence of the C-terminal region was transformed to a completely new polypeptide. Based on this finding, 7 additional recombinant prolamin genes of similar structure were isolated with specific PCR primers. The 8 chimeric clones seem to be derived from 4 individual gamma-gliadin and 3 LMW-glutenin sequences. These genes show remarkable diversity in size, gliadin:glutenin ratio, frameshift mutations, and sulphur content. The putative functional characteristics of the chimeric polypeptides and problems related to the origin of the encoding genes are discussed.     

The stress-induced SCP/HLIP family of small light-harvesting-like proteins (ScpABCDE) protects Photosystem II from photoinhibitory damages in the cyanobacterium <Emphasis Type="Italic">Synechocystis</Emphasis> sp. PCC 6803

Small CAB-like proteins (SCPs) are single-helix light-harvesting-like proteins found in all organisms performing oxygenic photosynthesis. We investigated the effect of growth in moderate salt stress on these stress-induced proteins in the cyanobacterium Synechocystis sp. PCC 6803 depleted of Photosystem I (PSI), which expresses SCPs constitutively, and compared these cells with a PSI-less/ScpABCDE^? mutant. SCPs, by stabilizing chlorophyll-binding proteins and Photosystem II (PSII) assembly, protect PSII from photoinhibitory damages, and in their absence electrons accumulate and will lead to ROS formation. The presence of 0.2 M NaCl in the growth medium increased the respiratory activity and other PSII electron sinks in the PSI-less/ScpABCDE^? strain. We postulate that this salt-induced effect consumes the excess of PSII-generated electrons, reduces the pressure of the electron transport chain, and thereby prevents ¹O₂ production.