A Recombinant Saccharomyces cerevisiae Strain Overproducing Mannoproteins Stabilizes Wine against Protein Haze

Stabilization against protein haze was one of the first positive properties attributed to yeast mannoproteins in winemaking. In previous work we demonstrated that deletion of KNR4 leads to increased mannoprotein release in laboratory Saccharomyces cerevisiae strains. We have now constructed strains with KNR4 deleted in two different industrial wine yeast backgrounds. This required replacement of two and three alleles of KNR4 for the EC1118 and T73-4 backgrounds, respectively, and the use of three different selection markers for yeast genetic transformation. The actual effect of the genetic modification was dependent on both the genetic background and the culture conditions. The fermentation performance of T73-4 derivatives was clearly impaired, and these derivatives did not contribute to the protein stability of the wine, even though they showed increased mannoprotein release in vitro. In contrast, the EC1118 derivative with both alleles of KNR4 deleted released increased amounts of mannoproteins both in vitro and during wine fermentation assays, and the resulting wines were consistently less susceptible to protein haze. The fermentation performance of this strain was slightly impaired, but only with must with a very high sugar content. These results pave the way for the development of new commercial strains with the potential to improve several mannoprotein-related quality and technological parameters of wine.     

New and little known cladocerans (Crustacea: Anomopoda) from southeastern Mexico

Mexico represents a transitional area in which two biogeographic regions meet. A survey of the cladocerans collected in true tropical waterbodies from the country revealed a diverse fauna. The environments examined included temporary pools in the forest, rivers, permanent lagoons and karstic sinkholes. New records or little known taxa include: Diaphanosoma brevireme, D. fluviatile, Pseudosida ramosa, Simocephalus mixtus, Onchobunops tuberculatus,Guernella raphaelis, Chydorus nitidilus, Notoalonacf. globulosa, Alonella brasiliensis and Leydigiopsis curvirostris. In contrast to high altitude systems, with clear nearctic affinity, the lowlands of southeastern Mexico showed a striking resemblance with the South and Central American cladoceran fauna. Apparently, orographic barriers control the distribution of the cladoceran taxa in this region.     

Systematic revision of the genus Petromica Topsent (Demospongiae: Halichondrida), with a new species from the southwestern Atlantic

The status, scope and classification of the sublithistid demosponge genus Petromica Topsent are revised through morphological analysis of museum specimens of all seven species (including proposed synonyms and varieties), two of which were collected and observed in situ along the Brazilian coast (P. ciocalyptoides (Van Soest & Zea) and P. citrina sp. n.). The synonymy of Petromica and Monanthus Kirkpatrick with priority to the former is justified due to the consistent presence of monocrepid rhizoclone desmas and oxeas in an halichondroid arrangement, and to the lack of co-variance in other morphological characters among the species studied (presence and shape of papillae, surface texture, ectosomal skeleton and desma shape). The proposed synonymy of P. grimaldii Topsent and P. massalis Dendy is refuted due to differences in habit and spicule shape between the two species. Three forms described as varieties of Monanthus plumosus Kirkpatrick are raised to species level: P. plumosa (Kirkpatrick), P. tubulata (Kirkpatrick) and P. digitata (Burton). Phylogenetic analysis indicates that two possibly monophyletic clades may be recognized within Petromica, although with low bootstrap support (35–59%): (P. ciocalyptoides, P. citrina) and (P. grimaldii, P. massalis) (P. plumosa) (P. tubulata) (P. digitata). The classification of Petromica within the Halichondriidae (order Halichondrida) is supported by the confused reticulation of long oxeote spicules with ascending spicule tracts, present in all species of the genus.     

Proteolytic activation of respiratory syncytial virus fusion protein. Cleavage at two furin consensus sequences.

The F (fusion) protein of the respiratory syncytial viruses is synthesized as an inactive precursor F(0) that is proteolytically processed at the multibasic sequence KKRKRR(136) into the subunits F(1) and F(2) by the cellular protease furin. This maturation process is essential for the F protein to gain fusion competence. We observed that proteolytic cleavage additionally occurs at another basic motif, RARR(109), that also meets the requirements for furin recognition. Cleavage at both sites leads to the removal from the polypeptide chain of a glycosylated peptide of 27 amino acids. When the sequence RARR(109) was changed to NANR(109) or to RANN(109) by site-directed mutagenesis, cleavage by furin was completely prevented. Although the mutants were still processed at position Arg(136), they did not show any syncytia formation. Proteolytic cleavage of the modified motifs was achieved by treatment of transfected cells with trypsin converting the F mutants into their fusogenic forms. Our findings indicate that both furin consensus sequences have to be cleaved in order to activate the fusion protein.     

Genome organization in dicots. II. Arabidopsis as a ’bridging species’ to resolve genome evolution events among legumes

Analysis of molecular linkage groups within the soybean (Glycine max L. Merr.) genome reveals many homologous regions, reflecting the ancient polyploidy of soybean. The fragmented arrangement of the duplicated regions suggests that extensive rearrangements, as well as additional duplications, have occurred since the initial polyploidization event. In this study we used comparisons between homoeologous regions in soybean, and the homologous regions in the related diploids Phaseolus vulgaris and Vigna radiata, to elucidate the evolutionary history of the three legume genomes. Our results show that there is not only conservation of large regions of the genomes but that these conserved linkage blocks are also represented twice in the soybean genome. To gain a better understanding of the process of genome evolution in dicots, molecular comparisons have been extended to another well-studied species, Arabidopsis thaliana. Interestingly, the conserved regions we identified in the legume species are also relatively conserved in Arabidopsis. Our results suggest that there is conservation of blocks of DNA between species as distantly related as legumes and brassicas, representing 90 million years of divergence. We also present evidence for an additional, presumably earlier, genome duplication in soybean. These duplicated regions were only recognized by using Arabidopsis as a ’bridging’ species in the genome comparisons. Received: 10 October 2000 / Accepted: 13 January 2001     

Semifree-ranging Tufted Capuchins (Cebus apella) Spontaneously Use Tools to Crack Open Nuts

Naturalistic studies on tool use by nonhuman primates have focused almost exclusively on Old World monkeys or hominoids. We studied the cracking of Syagrus nuts with the aid of stones by a group of semifree-ranging capuchins living in a reforested area (Tietê Ecological Park, São Paulo, Brazil). Our data are from direct observation and from mapping nut-cracking site utilization. All adults, subadults and juveniles (plus one infant) crack nuts, but individual differences in frequency and proficiency are marked. Juveniles do most of the nut-cracking, but adults are, on average, more efficient; the frequency of inept stone manipulation decreases with age. About 10% of the nut-cracking episodes were watched by other individuals—mostly infants and juveniles, suggesting a role for observational learning, even if restricted to stimulus enhancement.     

Complementation of Bacteriophage Induction and Recombination Defects in <Emphasis Type="Italic">Escherichia coli</Emphasis> RecA<Superscript>−</Superscript> Mutants by Expression of the Cloned T4 Bacteriophage <Emphasis Type="Italic">uvsX</Emphasis> Gene

Previous workers reported that the T4 bacteriophage UvsX protein could promote neither RecA-LexA-mediated DNA repair nor induction of lysogenized bacteriophage, only recombination. Reexamination of these phenotypes demonstrated that, in contrast to these prior studies, when this gene was cloned into a medium but not a low-copy-number vector, it stimulated both a high frequency of spontaneous induction and mitomycin C-stimulated bacteriophage induction in a strain containing a recA13 mutation, but not a recA1 defect. The gene when cloned into a low- or medium- copy-number vector also promoted a low frequency of recombination of two duplicated genes in Escherichia coli in a strain with a complete recA gene deletion. These results suggest that a narrow concentration range of T4 UvsX protein is required to promote both high-frequency spontaneous and mitomycin C-stimulated bacteriophage induction in a recA13 gene mutant, but it facilitates recombination of duplicated genes at only a very low frequency in E. coli RecA⁻ mutants with a complete recA deletion. These results also suggest that the different UvsX phenotypes are affected differentially by the concentration of UvsX protein present. Received: 11 February 2002 / Accepted: 12 April 2002     

Local enrichment with homopolymeric (dA/dT) DNA in genomes of some lower dipterans and Drosophila melanogaster

An investigation into the chromosomal localization of homopolymeric dA/dT was carried out with species of the genera Rhynchosciara, Chironomus, Drosophila and several other taxa. In situ hybridisation probing mitotic and polytene chromosomes with RNA homopolymers was performed, followed by immunological detection of the DNA/RNA hybrid. Use of this method allowed us to assess specific regions of some dipteran genomes, where the signal was generally, but not always, located in heterochromatic regions. Human and Drosophila chromosome regions known to contain dA/dT runs of up to 153 bp were devoid of consistent labelling. The stability of the rA/dT hybrid formed in situ was in agreement with the T(m) for long rA/dT hybrid complexes, suggesting that the method used in this work is able to identify unusually long homopolymeric dA/dT tracts.