Cloning of a cryV-type insecticidal protein gene from Bacillus thuringiensis: the cryV-encoded protein is expressed early in stationary phase.

A CryV-type protein (CGCryV) has been isolated from supernatant fluids of Bacillus thuringiensis AB88 cultures. Previous reports have suggested the cryptic nature of the cryV-type genes on the basis of the absence of CryV-type proteins in parasporal crystals. The CryV-type protein reported here is expressed early in stationary phase, and evidence indicates that it is an exported protein. Analysis of the deduced protein sequence from this gene reveals the presence of an N-terminal domain that likely acts as a signal peptide. The CGCryV protein is the first reported case of a delta-endotoxin being a secreted protein, which may influence the biological relevance of these proteins.     

Ethanol production in a continuous fermentation/membrane pervaporation system

The productivity of ethanol fermentation processes, predominantly based on batch operation in the U.S. fuel ethanol industry, could be improved by adoption of continuous processing technology. In this study, a conventional yeast fermentation was coupled to a flat-plate membrane pervaporation unit to recover continuously an enriched ethanol stream from the fermentation broth. The process employed a concentrated dextrose feed stream controlled by the flow rate of permeate from the pervaporation unit via liquid-level control in the fermentor. The pervaporation module contained 0.1 m² commercially available polydimethylsiloxane membrane and consistently produced a permeate of 20%–23% (w/w) ethanol while maintaining a level of 4%–6% ethanol in a stirred-tank fermentor. The system exhibited excellent operational stability. During continuous operation with cell densities of 15–23 g/l, ethanol productivities of 4.9–7.8 gl^–1 h^–1 were achieved utilizing feed streams of 269–619 g/l glucose. Pervaporation flux and ethanol selectivities were 0.31–0.79 lm^–2 h^–1 and 1.8–6.5 respectively.     

Arabidopsis consensus intron sequences

We have analysed 998 Arabidopsis intron sequences in the EMBL database. All Arabidopsis introns to adhere to the :GU...AG: rule with the exception of 1% of introns with :GC at their 5 ends. Virtually all of the introns contained a putative branchpoint sequence (YUNAN) 18 to 60 nt upstream of the 3 splice site. Although a polypyrimidine tract was much less apparent than in vertebrate introns, the most common nucleotide in the region upstream of the 3 splice site was uridine. Consensus sequences for 5 and 3 splice sites and branchpoint sequences for Arabidopsis introns are presented.     

A new major histocompatibility complex class I b gene expressed in the mouse blastocyst and placenta

 Because of the role major histocompatibility complex (MHC) class I b molecules may play during mouse embryonic development, we thought it would be interesting to search for additional MHC class I b molecules that might be expressed in preimplantation embryos, and in particular in the trophoblastic lineage. We therefore screened a mouse preimplantation blastocyst cDNA library for MHC class I sequences. This search led to the identification and characterization of a new MHC class I b gene, blastocyst MHC. Sequences identical to the exons and 3′ untranslated region of this gene have been found in many laboratory mouse strains, as well as in the related mouse species Mus spreciligus. The presence of this gene in mouse strains of different MHC class I haplotypes argues that blastocyst MHC is a unique, newly-described gene rather than a new allele of a previously described mouse MHC class I gene. Blastocyst MHC has the structure of an MHC class I b gene, with the six exons characteristic of T-region genes. It is linked to H2-D. The amino acid sequence encoded by this gene maintains all the features of a functional antigen-presentation domain. The blastocyst MHC gene, like the human class I b gene HLA-G, is expressed at the blastocyst stage and in the placenta, and may be the mouse analog for HLA-G. Received: 31 May 1996 / Revised: 19 August 1996     

Behavior of Colonial Orb-weaving Spiders during a Solar Eclipse

The behavior of colonial orb-weaving spiders (Metepeira incrassata) in tropical Veracruz, Mexico was studied during the total solar eclipse on July 11, 1991. Spiders behaved in a manner typical of daily activity until totality, when many began taking down webs. After solar reappearance, most spiders that had begun taking down webs rebuilt them. There was no significant difference in the overall activity patterns of spiders during totality across a range of colony sizes. Experimental illumination of part of a colony during totality altered web takedown behavior. While spiders in the darkness of totality began to take down webs, those spiders which were artificially illuminated did not. These observations suggest that the primary environmental cue responsible for the daily rhythm of web building behavior in this species is light level.     

A role for the human DNA repair enzyme HAP1 in cellular protection against DNA damaging agents and hypoxic stress.

The HAP1 protein (also known as APE/Ref-1) is a bifunctional human nuclear enzyme required for repair of apurinic/apyrimidinic sites in DNA and reactivation of oxidized proto-oncogene products. To gain insight into the biological roles of HAP1, the effect of expressing antisense HAP1 RNA in HeLa cells was determined. The constructs for antisense RNA expression consisted of either a full-length HAP1 cDNA or a genomic DNA fragment cloned downstream of the CMV promoter in pcDNAneo. Stable HeLa cell transfectants expressing HAP1 antisense RNA were found to express greatly reduced levels of the HAP1 protein compared to equivalent sense orientation and vector-only control transfectants. The antisense HAP1 transfectants exhibited a normal growth rate, cell morphology and plating efficiency, but were hypersensitive to killing by a wide range of DNA damaging agents, including methyl methanesulphonate, hydrogen peroxide, menadione, and paraquat. However, survival after UV irradiation was unchanged. The antisense transfectants were strikingly sensitive to changes in oxygen tension, exhibiting increased killing compared to controls following exposure to both hypoxia (1% oxygen) and hyperoxia (100% oxygen). Consistent with a requirement for HAP1 in protection against hypoxic stress, expression of the HAP1 protein was found to be induced in a time-dependent manner in human cells during growth under 1% oxygen. The possible involvement of a depletion of cellular glutathione being linked to the hypoxic stress-sensitive phenotype of the antisense HAP1 transfectants came from the finding that they also exhibited hypersensitivity to buthionine sulphoximine, an inhibitor of glutathione biosynthesis. We conclude that the HAP1 protein is a key factor in cellular protection against a wide variety of cellular stresses, including DNA damage and a change in oxygen tension.     

Quantitative trait locus analysis of susceptibility to diet-induced atherosclerosis in recombinant inbred mice

Quantitative trait locus (QTL) analysis is a statistical method that can be applied to identify loci making a significant impact on a phenotype. For the phenotype of susceptibility to diet-induced atherosclerosis in the mouse, we have studied four quantitative traits: area of aortic fatty streaks and serum concentrations of high-density lipoprotein-bound cholesterol (HDL-cholesterol), apolipoprotein A-I, and apolipoprotein A-II (apo A-II). QTL analysis revealed a significant locus on chromosome 1 distal impacting serum apo A-II concentration on a high-fat diet and serum HDL-cholesterol concentration on a chow diet. This locus is presumablyApoa-2, the structural gene for apo A-II. QTL analysis of aortic fatty streaks failed to reveal a significant locus.     

EVOLUTIONARY SHIFTS IN THE SPECTRAL PROPERTIES OF SPIDER SILKS

We measured the reflectance properties of unpigmented silks spun by a systematic array of primitive (Deinopoidea) and derived (Araneoidea) aerial, web-spinning spiders, as well as silks spun by Araneomorphae and Mygalomorphae spiders that do not spin aerial webs. Our data show that all of the primitive aerial web spinners produce catching silks with a spectral peak in the ultraviolet (UV), and cladistic analysis suggests that high UV reflection is the primitive character state for silk spectral properties. In contrast, all of the derived aerial web spinners produce silks that are spectrally flat or characterized by reduced reflectance in the UV. Correlated with the evolution of these catching silks is a 37-fold increase in species number and apparent habitat expansion. This suggests that the unique silk proteins spun by the araneoids have been important to their ecological and evolutionary diversity.     

Cloning of the nupC gene of Escherichia coli encoding a nucleoside transport system, and identification of an adjacent Insertion element, IS 186

Escherichia coli is known to contain more than one active transport system for nucleoside uptake. In the present study we report the sequence of a gene encoding a second nucleoside transport system, nupC (in addition to nupG.) An open reading frame (ORF) of 1200bp was identified that codes for a hydrophobic polypeptide of 43 560 Da and an NupC fusion protein was shown to be membrane associated. The native NupC protein is also identified, following over-expression. NupC exhibits short regions of homology to several membrane-associated proteins, including LacY and Cyd. Analysis of the nupC promoter region revealed the presence of at least two putative CRP-binding sites, centred at–40bp and–89bp, which probably flank a CytR-binding site. In addition, an adjacent IS186 element was identified and found to reside within a putative terminator structure, downstream from the nupC ORF. This arrangement is shown to reflect the previously established gene order on the E. coli chromosome.     

Metabolic engineering of plant secondary products

Plants interact with their environment by producing a diverse array of secondary metabolites. Many of these compounds are valued for their medicinal, industrial or agricultural properties. Other secondary products are toxic or otherwise undesirable and can reduce the commercial value of crops. Gene transfer technology offers new opportunities to modify directly plant secondary product synthesis through metabolic engineering. This article reviews some of the strategies which have been used to increase or decrease the synthesis of specific plant metabolites, as well as methods for expanding the biosynthetic capabilities of individual species.