The Epstein-Barr virus (EBV) BZLF1 immediate-early gene product differentially affects latent versus productive EBV promoters.

The Epstein-Barr virus (EBV) BZLF1 gene product is thought to mediate the disruption of latent EBV infection. We have examined the regulatory effects of BZLF1 by studying its transactivating effects on seven different EBV promoters. We find that whereas the BZLF1 gene product increases the activity of the two early promoters, BMLF1 and BMRF1, it decreases the activity of three latent promoters (the BamHI-C and BamHI-W Epstein-Barr nuclear antigen promoters and the latent membrane protein promoter). The BZLF1-induced changes in promoter-directed chloramphenicol acetyltransferase activity occur in EBV-negative as well as EBV-positive cell lines and are accompanied by a similar change in chloramphenicol acetyltransferase mRNA. Deletion analysis of the BamHI Z fragment indicates that in a portion of the amino-terminal half of the BZLF1 gene product (amino acids 24 to 86) is not essential for positive transactivating effects but is required for down-regulating effects. Thus, different domains of the same EBV immediate-early gene product can either increase the function of EBV promoters active in productive infection or decrease the function of key promoters active in latent infection.     

Origin heterogeneity of Hb Lepore-Boston gene in Italy

Forty-three hybrid delta-beta-globin genes were characterized by DNA sequence analysis and associated RFLP haplotypes in 40 families from Abruzzo and Campania, which are on the east and west coast of Italy, respectively. All the genes had the delta-globin sequence up to the exon 2 codon 87 and had the beta-globin sequence from IVS-2-8; between these two ends, they had 58 bp in common with the delta- and beta-globin genes. Thus, they were all of the Lepore-Boston type. A chromosomal background heterogeneity was present among the mutant genes. In fact, they were all associated with (+ - - - -) 5' subhaplotype, but 23/31 from Campania were associated with (+ +) 3' subhaplotype, whereas 12/12 genes from Abruzzo and 8/31 from Campania were associated with (+ -). DNA sequencing of homozygous subjects showed that (+ +) 3' subhaplotype was associated, at IVS-2-74, with G, while (+ -) was associated with T; that is they were associated with the beta-globin gene sequence of frameworks 1 and 2, respectively. The molecular characteristics of this heterogeneity, as well as its geographical patterns in the eastern and western regions of Italy, represent strong evidence for the recurrent and multicentric origins of the mutation.     

Calbindin D28k is essentially located in the colonic part of the toad intestine

Summary— The distribution of calbindin D28k in the digestive system and the urinary bladder of the toad was investigated using immunohistochemistry and Western blotting. By analogy with mammals and birds, the protein was expected to be located preferentially in the duodenal part of the intestine. Interestingly, absorptive cells of the duodenum were totally devoid of calbindin D28k while the colon contained high amounts of the calcium-binding protein. This reversed polarity of calbindin D28k content in the toad intestine should obviously correspond to a different scheme of calcium absorption regulation between amphibians and higher vertebrates. Calbindin D28k containing neuroendocrine-like cells were found scattered in the proximal parts of the gut with a similar distribution to what has been described in rat and chick intestine. The oesophagus, the stomach, and the intrinsic nervous sytem of the intestine were negative. No significant amounts of the proteins were found in the urinary bladder, which is known to be a site of Ca²⁺ active transport.     

Evolution of structure and substrate specificity ind-alanine:d-Alanine ligases and related enzymes

Thed-alanine:d-alanine-ligase-related enzymes can have three preferential substrate specificities. Usually, these enzymes synthesized-alanyl-d-alanine. In vancomycin-resistant Gram-positive bacteria, structurally related enzymes synthesized-alanyl-d-lactate or Dalanyl-d-serine. The sequence of internal fragments of eight structurald-alanine:d-alanine ligase genes from enterococci has been determined. Alignment of the deduced amino acid sequences with those of other related enzymes from Gram-negative and Gram-positive bacteria revealed the presence of four distinct sequence patterns in the putative substrate-binding sites, each correlating with specificity to a particular substrate (d-alanine:d-lactate ligases exhibited two patterns). Phylogenetic analysis showed different clusters. The enterococcal subtree was largely superimposable on that derived from 16S rRNA sequences. In lactic acid bacteria, structural divergence due to differences in substrate specificity was observed. Glycopeptide resistance proteins VanA and VanB, the VanC-type ligases, and Dd1A and DdlB from enteric bacteria andHaemophilus influenzae constituted separate clusters. Correspondence to: P. Courvalin     

Regulation of Human (Caco-2) Intestinal Epithelial Cell Differentiation by Extracellular Matrix Proteins

Extracellular matrix regulation of intestinal epithelial differentiation may affect development, differentiation during migration to villus tips, healing, inflammatory bowel disease, and malignant transformation. Cell culture studies of intestinal epithelial biology may also depend on the matrix substrate used. We evaluated matrix effects on differentiation and proliferation in human intestinal Caco-2 epithelial cells, a model for intestinal epithelial differentiation. Proliferation, brush border enzyme specific activity, and spreading were compared in cells cultured on tissue culture plastic with interstitial collagen I and the basement membrane constituents collagen IV and laminin. Each matrix significantly increased alkaline phosphatase, dipeptidyl peptidase, lactase, sucrase-isomaltase, and cell spreading in comparison to plastic. However, the basement membrane proteins collagen IV and laminin further promoted all four brush border enzymes but inhibited spreading compared to collagen I. Proliferation was most rapid on type I collagen and slowest on laminin and tissue culture plastic. Basement membrane matrix proteins may promote intestinal epithelial differentiation and inhibit proliferation compared with interstitial collagen I.     

In Vivo and In Vitro Evidence for the Biosynthesis of Testosterone in the Telencephalon of the Female Frog

Abstract: Neurons and glial cells are capable of synthesizing various steroid hormones, but biosynthesis of testosterone in the CNS has never been reported. The aim of the present study was to demonstrate the synthesis of testosterone in the frog brain. The presence of 17β-hydroxysteroid dehydrogenase (17β-HSD)-like immunoreactivity was detected in a population of glial cells located in the telencephalon. Reversed-phase HPLC analysis of brain tissue extracts combined with radioimmunoassay detection revealed the presence of substantial amounts of testosterone and 5α-dihydrotestosterone (5α-DHT) in the telencephalon where 17β-HSD-positive cells were visualized. In male frogs, castration totally suppressed testosterone and 5α-DHT in the blood and in the rhombencephalon but did not affect the concentration of these two steroids in the telencephalon. Chemical characterization of testosterone in female frog telencephalon extracts was performed by coupling HPLC analysis with gas chromatography-mass spectrometry. Using the pulse-chase technique with [³H]pregnenolone as a precursor, the formation of a series of metabolites was observed, including dehydroepiandrosterone, androstenedione, testosterone, 5α-DHT, and estradiol. These data demonstrate the existence of an active form of 17β-HSD in the frog telencephalon, which is likely involved in testosterone biosynthesis within the brain.     

Trends in plant carbon concentration and plant demand for N throughout this century

 Atmospheric CO₂ concentration has increased by 25% over the preindustrial level. A parallel increase in C concentration and decreases in N concentration and δ¹³C of plants grown throughout this century have been observed in plant specimens stored in herbaria. We tested our previous results in a study of 12 more species collected in the western Mediterranean throughout this century (1920–1930, 1945–1955, and 1985–1990) and tree rings of Quercus pubescens from the same area. These changes were accompanied by apparent increases in condensed tannin concentration. A decreasing trend in δ¹⁵N both in herbarium material and tree rings was also found, indicating that ecosystems might cope with higher plant N demand by decreasing N losses and increasing N fixation and mineralization. These results may contribute to a better understanding of the effects of global change on carbon and nitrogen cycling. Received: 12 November 1995 / Accepted: 17 May 1996     

Dimethylsulfoxide reduction by marine sulfate-reducing bacteria

Abstract Dimethylsulfoxide (DMSO) reduction occurred in five out of nine strains of sulfate-reducing bacteria from marine or saline environments, but not in three freshwater isolates. DMSO reduction supported growth in all positive strains. In Desulfovibrio desulfuricans strain PA2805, DMSO reduction occurred simultaneously with sulfate reduction and was not effectively inhibited by molybdate, a specific inhibitor of sulfate reduction. The growth yield per mol lactate was 26% higher with DMSO than with sulfate as electron acceptor. In extracts of cells of strain PA2805 grown on sulfate, a low level of DMSO-reducing activity was present (0.013 μmol (mg protein)⁻ min⁻); higher levels were found in cells grown on DMSO (0.56 μmol (mg protein)⁻ min⁻). In anoxic marine environments DMSO reduction by sulfate-reducing bacteria may lead to enhanced dimethylsulfide emission rates.     

O-glycosylation of dipeptides using β-galactosidase activity of Achatina achatina digestive juice

Summary Enzymatic O-glycosylation of dipeptide derivatives containing a serine residue in the N or C terminal position and alanine or glycine as the second amino acid was achieved using the transgalactosylation activity of -galactosidase from the Achatina achatina digestive juice. Reactions were performed with lactose as glycosyl donor and the dipeptide ethyl (or methyl) esters N-protected by a benzyloxycarbonyl group (Z) as glycosyl acceptors. Yields of galactosyl-dipeptide derivatives were much higher than those obtained with the E.coli -galactosidase as catalyst.     

Ethylene production and involvement during the first steps of durum wheat (Triticum durum) anther culture

The role of ethylene in anther culture of durum wheat ( Triticum durum Desf. cv. Ardente) was analyzed by testing the effects of 2-chloroethylphosphonic acid (ethrel) silver thiosulfate (Ag⁺), a -aminooxyacetic acid (AOA) and 1-aminocyclopropane-l-carboxylic acid (ACC) on microspore division observed after 21 days of culture and on development of calli estimated at day 45. The use of ethrel and Ag⁺ indicated a positive effect of ethylene on microspore division, whereas the use of AOA, and to a lesser extent ACC, snowed a negative effect. In contrast, the addition of ethrel or Ag⁺ indicated that ethylene inhibits the development of microspore-derived calli. AOA gave contradictory results. Ethylene production by anthers was about 7 pl anther⁻¹h⁻¹ and decreased during culture. ACC content in the anthers was maximal at day 9, whereas malonyl ACC (MACC) increased sharply from day 0 to day 3 and then decreased. The addition of AOA or ACC to the culture medium decreased or increased, respectively, ethylene production of anthers and the ACC and/or MACC content, but at concentrations higher than those that modified the formation of calli. This formation seems to occur in two successive phases: induction and initiation of microspore division, which was promoted by ethylene, followed by callus development, which was inhibited by ethylene.