bluuebunny/crossref_metadata_2025 huggingface dataset:
Genetic and physical mapping of homoeologous recombination points involving wheat chromosome 2B and rye chromosome 2R
Authors: Lukaszewski A J , Rybka K , Korzun V , Malyshev S V , Lapinski B , Whitkus R
Published: April 2004

Wide hybrids have been used in generating genetic maps of many plant species. In this study, genetic and physical mapping was performed on ph1b-induced recombinants of rye chromosome 2R in wheat (Triticum aestivum L.). All recombinants were single breakpoint translocations. Recombination 2RS–2BS was absent from the terminal and the pericentric regions and was distributed randomly along an intercalary segment covering approximately 65% of the arm's length. Such a distribution probably resulted from structural differences at the telomeres of 2RS and wheat 2BS arm that disrupted telomeric initiation of pairing. Recombination 2RL–2BL was confined to the terminal 25% of the arm's length. A genetic map of homoeologous recombination 2R–2B was generated using relative recombination frequencies and aligned with maps of chromosomes 2B and 2R based on homologous recombination. The alignment of the short arms showed a shift of homoeologous recombination toward the centromere. On the long arms, the distribution of homoeologous recombination was the same as that of homologous recombination in the distal halves of the maps, but the absence of multiple crossovers in homoeologous recombination eliminated the proximal half of the map. The results confirm that homoeologous recombination in wheat is based on single exchanges per arm, indicate that the distribution of these single homoeologous exchanges is similar to the distribution of the first (distal) crossovers in homologues, and suggest that successive crossovers in an arm generate specific portions of genetic maps. A difference in the distribution of recombination between the short and long arms indicates that the distal crossover localization in wheat is not dictated by a restricted distribution of DNA sequences capable of recombination but by the pattern of pairing initiation, and that can be affected by structural differences. Restriction of homoeologous recombination to single crossovers in the distal part of the genetic map complicates chromosome engineering efforts targeting genes in the proximal map regions.Key words: homoeologous recombination, genetic mapping, RFLP, RAPD, wheat, rye.
Sequence assessment of comigrating AFLPTM bands in Echinacea — implications for comparative biological studies
Authors: Mechanda Subbaiah M , Baum Bernard R , Johnson Douglas A , Arnason John T
Published: April 2004

The extent of sequence identity among clones derived from monomorphic and polymorphic AFLPTM polymorphism bands was quantified. A total of 79 fragments from a monomorphic band of 273 bp and 48 fragments from a polymorphic band of 159 bp, isolated from individuals belonging to different populations, varieties, and species of Echinacea, were cloned and sequenced. The monomorphic fragments exhibited above 90% sequence identity among clones within samples. Sequence identity within variety ranged from 82.78% to 94.87% and within species from 75.82% to 98.9% and was 57.97% in the genus. The polymorphic fragments exhibited much less sequence identity. In some instances, even two clones from the same fragment were different in their size and sequence. Within sample, clone sequence identity ranged from 100% to 51.57%, within variety from 33.33% to 100% in one variety, and from 23.66% to 45% within species and was as low as 1.25% within the genus. In addition, sequences of the same size were aligned to verify the nature of their sequence dissimilarity/similarity. Within each size group, identical sequences were found across species and varieties. In general, comigrating bands cannot be considered homologous. Thus, the use of AFLPTM band data for comparative studies is appropriate only if the results emanating from such analyses are considered as approximations and are interpreted as phenotypic but not genotypic.Key words: AFLP markers, false homologies.
Construction of a high-density linkage map of Italian ryegrass (Lolium multiflorumLam.) using restriction fragment length polymorphism, amplified fragment length polymorphism, and telomeric repeat associated sequence markers
Authors: Inoue Maiko , Gao Zhensheng , Hirata Mariko , Fujimori Masahiro , Cai Hongwei
Published: April 2004

To construct a high-density molecular linkage map of Italian ryegrass (Lolium multiflorum Lam.), we used a two-way pseudo-testcross F1population consisting of 82 individuals to analyze three types of markers: restriction fragment length polymorphism markers, which we detected by using genomic probes from Italian ryegrass as well as heterologous anchor probes from other species belonging to the Poaceae family, amplified fragment length polymorphism markers, which we detected by using PstI/MseI primer combinations, and telomeric repeat associated sequence markers. Of the restriction fragment length polymorphism probes that we generated from a PstI genomic library, 74% (239 of 323) of randomly selected probes detected hybridization patterns consistent with single-copy or low-copy genetic locus status in the screening. The 385 (mostly restriction fragment length polymorphism) markers that we selected from the 1226 original markers were grouped into seven linkage groups. The maps cover 1244.4 cM, with an average of 3.7 cM between markers. This information will prove useful for gene targeting, quantitative trait loci mapping, and marker-assisted selection in Italian ryegrass.Key words: linkage map, molecular markers, synteny, Italian ryegrass, Lolium multiflorum Lam.
Cloning and comparative analysis of the bovine, porcine, and equine sex chromosome genesZFXandZFY
Authors: Poloumienko Arkadi
Published: April 2004

A growing body of evidence suggests the involvement of sex chromosome genes in mammalian development. We report the cloning and characterization of the complete coding regions of the bovine Y chromosome ZFY and X chromosome ZFX genes, and partial coding regions of porcine and equine ZFX and ZFY genes. Bovine ZFY and ZFX are highly similar to each other and to ZFX and ZFY from other species. While bovine and human ZFY proteins are both 801 amino acids long, bovine ZFX is 5 amino acids shorter than human ZFX. Like in humans, both bovine ZFY and ZFX contain 13 zinc finger motifs and belong to the Krueppel family of C2H2-type zinc finger proteins. The internal exon–intron organization of the bovine, porcine and equine ZFX and ZFY genes has been determined and compared. Within this region, the exon lengths and the positions of the splice sites are conserved, further suggesting a high evolutionary conservation of the ZFX and ZFY genes. Additionally, new alternatively spliced forms of human ZFX have been identified.Key words: sex chromosome genes, zinc finger proteins, ZFX, ZFY, alternative splicing.
Nuclear and chloroplast DNA differentiation in Andean potatoes
Authors: Sukhotu Thitaporn , Kamijima Osamu , Hosaka Kazuyoshi
Published: April 2004

Over 3500 accessions of Andean landraces have been known in potato, classified into 7 cultivated species ranging from 2x to 5x (Hawkes 1990). Chloroplast DNA (ctDNA), distinguished into T, W, C, S, and A types, showed extensive overlaps in their frequencies among cultivated species and between cultivated and putative ancestral wild species. In this study, 76 accessions of cultivated and 19 accessions of wild species were evaluated for ctDNA types and examined by ctDNA high-resolution markers (ctDNA microsatellites and H3 marker) and nuclear DNA restriction fragment length polymorphisms (RFLPs). ctDNA high-resolution markers identified 25 different ctDNA haplotypes. The S- and A-type ctDNAs were discriminated as unique haplotypes from 12 haplotypes having C-type ctDNA and T-type ctDNA from 10 haplotypes having W-type ctDNA. Differences among ctDNA types were strongly correlated with those of ctDNA high-resolution markers (r = 0.822). Differentiation between W-type ctDNA and C-, S-, and A-type ctDNAs was supported by nDNA RFLPs in most species except for those of recent or immediate hybrid origin. However, differentiation among C-, S-, and A-type ctDNAs was not clearly supported by nDNA RFLPs, suggesting that frequent genetic exchange occurred among them and (or) they shared the same gene pool owing to common ancestry.Key words: potato, chloroplast DNA, microsatellite markers, nuclear DNA RFLPs.
Comparative physical mapping reveals features of microsynteny betweenGlycine max,Medicago truncatula, andArabidopsis thaliana
Authors: Yan H H , Mudge J , Kim D-J , Shoemaker R C , Cook D R , Young N D
Published: April 2004

To gain insight into genomic relationships between soybean (Glycine max) and Medicago truncatula, eight groups of bacterial artificial chromosome (BAC) contigs, together spanning 2.60 million base pairs (Mb) in G. max and 1.56 Mb in M. truncatula, were compared through high-resolution physical mapping combined with sequence and hybridization analysis of low-copy BAC ends. Cross-hybridization among G. max and M. truncatula contigs uncovered microsynteny in six of the contig groups and extensive microsynteny in three. Between G. max homoeologous (within genome duplicate) contigs, 85% of coding and 75% of noncoding sequences were conserved at the level of cross-hybridization. By contrast, only 29% of sequences were conserved between G. max and M. truncatula, and some kilobase-scale rearrangements were also observed. Detailed restriction maps were constructed for 11 contigs from the three highly microsyntenic groups, and these maps suggested that sequence order was highly conserved between G. max duplicates and generally conserved between G. max and M. truncatula. One instance of homoeologous BAC contigs in M. truncatula was also observed and examined in detail. A sequence similarity search against the Arabidopsis thaliana genome sequence identified up to three microsyntenic regions in A. thaliana for each of two of the legume BAC contig groups. Together, these results confirm previous predictions of one recent genome-wide duplication in G. max and suggest that M. truncatula also experienced ancient large-scale genome duplications.Key words: Glycine max, Medicago truncatula, Arabidopsis thaliana, conserved microsynteny, genome duplication.
Sequence characterized amplified region markers tightly linked to the mating factors ofLentinula edodes
Authors: Tanaka Ayako , Miyazaki Kazuhiro , Murakami Haruki , Shiraishi Susumu
Published: April 2004

Detecting the mating types in shiitake, Lentinula edodes (Berk.) Pegler, is important for making progress in the breeding of this mushroom and determining the compatibility of the pair to cross. Shiitake is a tetrapolar fungus with two unlinking mating factors, A factor and B factor. We screened molecular markers linked to the mating factors using the randomly amplified polymorphic DNA (RAPD) method to develop the mating type identification procedure. Using 147 oligonucleotide primers, a total of 6 linkage markers for the shiitake mating factors, 4 markers for the A factor and 2 markers for the B factor, were discovered with a logarithm of the odds threshold of 3.0 for linkage. Two RAPDs that perfectly segregated with each mating factor among 72 basidiospore strains were detected. Both of these RAPDs were cloned and sequenced to convert them to the sequence characterized amplified region (SCAR) markers. Four primers, two sets of primers, were designed according to the internal sequences of two RAPDs tightly linking to the A factor or B factor. Consequently, we determined the polymerase chain reaction condition for multiplex analyses of these SCAR markers.Key words: Lentinula edodes, SCAR, diagnostic, mating type.
The relation between pairing preference and chiasma frequency in tetrasomics of rye
Authors: Benavente E , Sybenga J
Published: April 2004

The association pattern of marked tetrasomes of Secale chromosome 1R at meiotic first metaphase was analyzed. Two of the four chromosomes were identical with terminal C-bands at both arms; the other two were also identical but lacked C-bands and were homologous or homeologous with the first two. Four different types of heterozygotes for 1R were studied: (i) autotetraploid hybrids between genetic variants within Secale cereale subsp. cereale, (ii) tetraploid hybrids between subspecies of Secale cereale, (iii) tetraploid hybrids between species of Secale, and (iv) autotetrasomes of S. cereale in a wheat background. Earlier observations that heterozygous associations (banded with unbanded) had consistently higher chiasma frequencies than homozygous associations were extended and confirmed. To analyze this phenomenon more closely, the possible relations between this correlation and several other meiotic phenomena were studied. For this analysis, three genetically different autotetraploid hybrids within S. cereale were selected that differed with respect to the relation between pairing type and chiasma frequency. Special attention was given to different patterns of interference and other meiotic phenomena in the two chromosome arms of chromosome 1R. No relations between such phenomena and the relation between pairing type and chiasma frequency could be established. A hypothesis is formulated assuming that long-distance homologue attraction is concentrated in a limited number of sites and that in different genotypes, different patterns of active sites are present. Moderately weak attraction sites can pair with strong homologous sites under favorable genetic conditions, but two weak sites cannot. Then, heterozygotes have more effective pairing initiation and consequently chiasma formation than homozygotes. Under less favorable conditions, only strong sites are effective, and then, homozygotes pair better, but the chiasma frequency is lower. A model of the forces involved in homologue attraction is presented.Key words: autopolyploids, preferential pairing, chiasma frequency, homologue attraction, Secale.
Caveolins, caveolae, and lipid rafts in cellular transport, signaling, and disease
Authors: Quest Andrew F.G , Leyton Lisette , Párraga Mario
Published: April 2004

Caveolae were initially described some 50 years ago. For many decades, they remained predominantly of interest to structural biologists. The identification of a molecular marker for these domains, caveolin, combined with the possibility to isolate such cholesterol- and sphingolipid-rich regions as detergent-insoluble membrane complexes paved the way to more rigorous characterization of composition, regulation, and function. Experiments with knock-out mice for the caveolin genes clearly demonstrate the importance of caveolin-1 and -3 in formation of caveolae. Nonetheless, detergent-insoluble domains are also found in cells lacking caveolin expression and are referred to here as lipid rafts. Caveolae and lipid rafts were shown to represent membrane compartments enriched in a large number of signaling molecules whose structural integrity is essential for many signaling processes. Caveolin-1 is an essential structural component of cell surface caveolae, important for regulating trafficking and mobility of these vesicles. In addition, caveolin-1 is found at many other intracellular locations. Variations in subcellular localization are paralleled by a plethora of ascribed functions for this protein. Here, more recent data addressing the role of caveolin-1 in cellular signaling and the development of diseases like cancer will be preferentially discussed.Key words: caveolae, rafts, membrane microdomains, caveolins, signal transduction, disease, cancer.
Low concentration of guanidine hydrochloride induces the formation of an aggregation-prone state in α-urease
Authors: McDuff F -O , Doucet A , Beauregard M
Published: April 2004

Canavalia ensiformis (jack bean) α-urease is a hexameric protein characterized by a complex denaturation mechanism. In previous papers, we have shown that a hydrophobic 8-anilino-1-naphthalenesulfonic acid (ANSA) binding conformer could be populated in a moderate concentration of denaturant. This state was obtained under conditions that had no detectable impact on its tertiary structure, as indicated by fluorescence measurements. In the present study, we further characterized this ANSA-binding state in an attempt to understand urease behavior. Evidence presented here shows that the presence of ANSA was not required for the generation of the conformer and that its affinity for ANSA came from an increase in hydrophobicity leading to aggregation. Circular dichroism investigation of urease revealed that it had periodical secondary structure content similar to Klebsiella aerogenes urease (secondary structures calculated on the basis of crystallographic data). The impact of 0.9 M guanidine hydrochloride (GuHCl) on soluble urease secondary structures was minimal but is compatible with a slight increase in beta-sheet structures. Such modification may indicates that aggregation involves amyloid-like fibril formation. Electron microscopy analysis of urease in the absence of GuHCl revealed the presence of urease hexamers (round shape 13 nm in diameter). These particles disappeared in the presence of moderate denaturant concentration owing to the formation of aggregates and fibril-like structures. The fibrils obtained in 1.5 M GuHCl had an average diameter of 6.5 nm, suggesting that urease hexamers dissociated into smaller oligomeric forms when forming such fibrils.Key words: protein structure, protein folding, denaturation, aggregation, multimeric proteins, protein fibrils, hydrophobicity, molten globule state.
Phospholipid biosynthesis in mammalian cells
Authors: Vance Jean E , Vance Dennis E
Published: April 2004

Identification of the genes and gene products involved in the biosynthesis of phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine has lagged behind that in many other fields because of difficulties encountered in purifying the respective proteins. Nevertheless, most of these genes have now been identified. In this review article, we have highlighted important new findings on the individual enzymes and the corresponding genes of phosphatidylcholine synthesis via its two major biosynthetic pathways: the CDP-choline pathway and the methylation pathway. We also review recent studies on phosphatidylethanolamine biosynthesis by two pathways: the CDP-ethanolamine pathway, which is active in the endoplasmic reticulum, and the phosphatidylserine decarboxylase pathway, which operates in mitochondria. Finally, the two base-exchange enzymes, phosphatidylserine synthase-1 and phosphatidylserine synthase-2, that synthesize phosphatidylserine in mammalian cells are also discussed.Key words: phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidic acid, biosynthesis.
Genetic analysis of intracellular aminoglycerophospholipid traffic
Authors: Voelker Dennis R
Published: April 2004

Inter- and intramembrane phospholipid transport processes are central features of membrane biogenesis and homeostasis. Relatively recent successes in the molecular genetic analysis of aminoglycerophospholipid transport processes in both yeast and mammalian cells are now providing important new information defining specific protein and lipid components that participate in these reactions. Studies focused on phosphatidylserine (PtdSer) transport to the mitochondria reveal that the process is regulated by ubiquitination. In addition, a specific mutation disrupts PtdSer transport between mitochondrial membranes. Analysis of PtdSer transport from the endoplasmic reticulum to the locus of PtdSer decarboxylase 2 demonstrates the requirement for a phosphatidylinositol-4-kinase, a phosphatidylinositol-binding protein, and the C2 domain of the decarboxylase. Examination of NBD-phosphatidylcholine transport demonstrates the involvement of the prevacuolar compartment and a requirement for multiple genes involved in regulating vacuolar protein sorting for transport of the lipid to the vacuole. In intramembrane transport, multiple genes are now identified including those encoding multidrug resistant protein family members, DNF family members, ATP binding cassette transporters, and pleiotropic drug resistance family members. The scramblase family constitutes a collection of putative transmembrane transporters that function in an ATP-independent manner. The genetic analysis of lipid traffic is uncovering new molecules involved in all aspects of the regulation and execution of the transport steps and also providing essential tools to critically test the involvement of numerous candidate molecules.Key words: lipid transport, lipid sorting, membrane biogenesis, organelles, flippase.
Diabetes, lipids, and adipocyte secretagogues
Authors: Faraj May , Lu Hui Ling , Cianflone Katherine
Published: April 2004

That obesity is associated with insulin resistance and type II diabetes mellitus is well accepted. Overloading of white adipose tissue beyond its storage capacity leads to lipid disorders in non-adipose tissues, namely skeletal and cardiac muscles, pancreas, and liver, effects that are often mediated through increased non-esterified fatty acid fluxes. This in turn leads to a tissue-specific disordered insulin response and increased lipid deposition and lipotoxicity, coupled to abnormal plasma metabolic and (or) lipoprotein profiles. Thus, the importance of functional adipocytes is crucial, as highlighted by the disorders seen in both "too much" (obesity) and "too little" (lipodystrophy) white adipose tissue. However, beyond its capacity for fat storage, white adipose tissue is now well recognised as an endocrine tissue producing multiple hormones whose plasma levels are altered in obese, insulin-resistant, and diabetic subjects. The consequence of these hormonal alterations with respect to both glucose and lipid metabolism in insulin target tissues is just beginning to be understood. The present review will focus on a number of these hormones: acylation-stimulating protein, leptin, adiponectin, tumour necrosis factor α, interleukin-6, and resistin, defining their changes induced in obesity and diabetes mellitus and highlighting their functional properties that may protect or worsen lipid metabolism.Key words: C3adesarg, fatty acid trapping, lipolysis, lipogenesis.
Phospholipase D
Authors: McDermott Mark , Wakelam Michael J.O , Morris Andrew J
Published: April 2004

Phospholipase D catalyses the hydrolysis of the phosphodiester bond of glycerophospholipids to generate phosphatidic acid and a free headgroup. Phospholipase D activities have been detected in simple to complex organisms from viruses and bacteria to yeast, plants, and mammals. Although enzymes with broader selectivity are found in some of the lower organisms, the plant, yeast, and mammalian enzymes are selective for phosphatidylcholine. The two mammalian phospholipase D isoforms are regulated by protein kinases and GTP binding proteins of the ADP-ribosylation and Rho families. Mammalian and yeast phospholipases D are also potently stimulated by phosphatidylinositol 4,5-bisphosphate. This review discusses the identification, characterization, structure, and regulation of phospholipase D. Genetic and pharmacological approaches implicate phospholipase D in a diverse range of cellular processes that include receptor signaling, control of intracellular membrane transport, and reorganization of the actin cytoskeleton. Most ideas about phospholipase D function consider that the phosphatidic acid product is an intracellular lipid messenger. Candidate targets for phospholipase-D-generated phosphatidic acid include phosphatidylinositol 4-phosphate 5-kinases and the raf protein kinase. Phosphatidic acid can also be converted to two other lipid mediators, diacylglycerol and lyso phosphatidic acid. Coordinated activation of these phospholipase-D-dependent pathways likely accounts for the pleitropic roles for these enzymes in many aspects of cell regulation.Key words: phospholipase D, phosphatidic acid, GTP-binding proteins, membrane transport, cytoskeletal regulation.
Regulation of arachidonic acid availability for eicosanoid production
Authors: Leslie Christina C
Published: April 2004

Mammalian cells have developed specific pathways for the incorporation, remodeling, and release of arachidonic acid. Acyltransferase and transacylase pathways function to regulate the levels of esterified arachidonic acid in specific phospholipid pools. There are several distinct, differentially regulated phospholipases A2in cells that mediate agonist-induced release of arachidonic acid. These pathways are important in controlling cellular levels of free arachidonic acid. Both arachidonic acid and its oxygenated metabolites are potent bioactive mediators that regulate a myriad of physiological and pathophysiological processes.Key words: phospholipase A2, arachidonic acid, eicosanoid, phospholipid.
Yeast sphingolipid metabolism: clues and connections
Authors: Sims Kellie J , Spassieva Stefka D , Voit Eberhard O , Obeid Lina M
Published: April 2004

This review of sphingolipid metabolism in the budding yeast Saccharomyces cerevisiae contains information on the enzymes and the genes that encode them, as well as connections to other metabolic pathways. Particular attention is given to yeast homologs, domains, and motifs in the sequence, cellular localization of enzymes, and possible protein–protein interactions. Also included are genetic interactions of special interest that provide clues to the cellular biological roles of particular sphingolipid metabolic pathways and specific sphingolipids.Key words : yeast, sphingolipid metabolism, subcellular localization, protein–protein interactions, stress response, aging.
The MARCKS family of phospholipid binding proteins: regulation of phospholipase D and other cellular components
Authors: Sundaram Meenakshi , Cook Harold W , Byers David M
Published: April 2004

Myristoylated alanine-rich C kinase substrate (MARCKS) and MARCKS-related protein (MRP) are essential proteins that are implicated in coordination of membrane-cytoskeletal signalling events, such as cell adhesion, migration, secretion, and phagocytosis in a variety of cell types. The most prominent structural feature of MARCKS and MRP is a central basic effector domain (ED) that binds F-actin, Ca2+-calmodulin, and acidic phospholipids; phosphorylation of key serine residues within the ED by protein kinase C (PKC) prevents the above interactions. While the precise roles of MARCKS and MRP have not been established, recent attention has focussed on the high affinity of the MARCKS ED for phosphatidylinositol 4,5-bisphosphate (PIP2), and a model has emerged in which calmodulin- or PKC-mediated regulation of these proteins at specific membrane sites could in turn control spatial availability of PIP2. The present review summarizes recent progress in this area and discusses how the above model might explain a role for MARCKS and MRP in activation of phospholipase D and other PIP2-dependent cellular processes.Key words: MARCKS, MRP, protein kinase C, PIP2, phospholipase D.
Oxysterol binding proteins: in more than one place at one time?
Authors: Olkkonen Vesa M , Levine Timothy P
Published: April 2004

Oxysterols are potent signalling lipids that directly bind liver X receptors (LXRs) and a subset of oxysterol binding protein (OSBP) related proteins (ORPs). It is relatively well established that the oxysterol-regulated function of LXRs is to control the expression of genes involved in reverse cholesterol transport, catabolism of cholesterol, and lipogenesis. In contrast, the mechanisms by which oxysterols and ORPs affect cellular lipid metabolism have remained poorly understood. In this review, we summarize the information available on function of the ORPs and compare the two families of proteins binding oxysterol to demonstrate the different responses that similar lipids can elicit within cells. The other focus is on the membrane targeting determinants and the protein interaction partners of ORPs, which provide interesting clues to the mode(s) of ORP action. Specifically, we suggest a model in which a general property of ORPs is to function at membrane contact sites, specialized zones of communication between two different organelles.Key words: endoplasmic reticulum, lipid transport, LXR, membrane contact sites, ORP, OSBP, Osh, sterol metabolism.
Biological functions of phosphatidylinositol transfer proteins
Authors: Routt Sheri M , Bankaitis Vytas A
Published: April 2004

Phosphatidylinositol/phosphatidylcholine transfer proteins (PITPs) are ubiquitous and highly conserved proteins that are believed to regulate lipid-mediated signaling events. Their ubiquity and conservation notwithstanding, PITPs remain remarkably uninvestigated. Little is known about the coupling of specific PITPs to explicit cellular functions or the mechanisms by which PITPs interface with apppropriate cellular functions. The available information indicates a role for these proteins in regulating the interface between lipid metabolism and membrane trafficking in yeast, signaling in plant development, the trafficking of specialized luminal cargo in mammalian enterocytes, and neurological function in mammals. Herein, we review recent advances in PITP biology and discuss as yet unresolved issues in this field.Key words: phosphatidylinositol transfer protein, secretion, lipid signaling, phosphoinositide.
Acid and neutral sphingomyelinases: roles and mechanisms of regulation
Authors: Marchesini Norma , Hannun Yusuf A
Published: April 2004

Ceramide, an emerging bioactive lipid and second messenger, is mainly generated by hydrolysis of sphingomyelin through the action of sphingomyelinases. At least two sphingomyelinases, neutral and acid sphingo myelinases, are activated in response to many extracellular stimuli. Despite extensive studies, the precise cellular function of each of these sphingomyelinases in sphingomyelin turnover and in the regulation of ceramide-mediated responses is not well understood. Therefore, it is essential to elucidate the factors and mechanisms that control the activation of acid and neutral sphingomyelinases to understand their the roles in cell regulation. This review will focus on the molecular mechanisms that regulate these enzymes in vivo and in vitro, especially the roles of oxidants (glu ta thi one, peroxide, nitric oxide), proteins (saposin, caveolin 1, caspases), and lipids (diacylglycerol, arachidonic acid, and ceramide).Key words: sphingomyelinase, ceramide, apoptosis, Niemann-Pick disease, FAN (factor associated with N-SMase activation).
Anoxia-induced transcriptional upregulation ofsarp-19: cloning and characterization of a novel EF-hand containing gene expressed in hepatopancreas ofLittorina littorea
Authors: Larade Kevin , Storey Kenneth B
Published: April 2004

Many marine molluscs have well-developed biochemical adaptations that allow them to live without oxygen for long periods of time, but very little is currently known about the molecular biology underlying these processes. Differential screening of a cDNA library derived from the hepatopancreas of the marine snail Littorina littorea revealed a novel anoxia-induced gene, sarp-19 (snail anoxia-responsive protein, 19 kDa). Examination of the sarp-19 transcript revealed an open reading frame that encoded a protein of 168 amino acids containing an N-terminal signal sequence and two putative EF-hand domains. Expression analysis of transcript levels established that sarp-19 accumulated over a time course of anoxia exposure, reaching a maximum 5.6-fold increase after 96 h compared with aerobic controls. However, transcript levels were reduced by 50% within 1 h when aerobic conditions were reestablished. Nuclear runoff assays confirmed transcriptional upregulation of sarp-19 during anoxia exposure, and organ explant experiments showed that the gene was also responsive to anoxia exposure in vitro. sarp-19 transcripts were also elevated in response to freezing, suggesting that the protein may have a role in the physiological responses of this intertidal snail to both aerial exposure and winter freezing. Hepatopancreas explants treated with a calcium ionophore showed increased levels of the sarp-19 transcript, suggesting a possible feedback mechanism regulated by levels of intracellular calcium. Expression was also responsive to tissue incubation with cyclic GMP and phorbol 12-myristate 13-acetate but was not affected by cyclic AMP, implicating involvement of protein kinases G and C but not protein kinase A in the expression of sarp-19. The SARP-19 protein may play a role in calcium-activated signaling during anoxia exposure in L. littorea.Key words: anoxia, mollusc, gastropod, calcium, EF hand.
ProbingDictyosteliumseverin structure and function by cross linking to actin
Authors: Summerscales Joanna , Dawson John F
Published: April 2004

DS151 is the first 151 amino acids of the Dictyostelium discoidium protein severin, which shares high sequence similarity with segment 1 of the actin-severing protein gelsolin. DS151 is able to mediate the depolymerization of F-actin in a calcium-dependent fashion, much like segment 1 of gelsolin. A structural model of DS151 was obtained by comparative modeling studies with segment 1 of gelsolin. This model was tested by studies of chemical cross linking between DS151 and bound actin, suggesting that Cys residues on DS151 are cross linked with Lys residues of actin. The model suggests that Cys125 of DS151 cross links with either Lys326 or Lys328 of actin. Mutagenesis of DS151 demonstrates that Cys125 of DS151 dominates the cross linking, whereas Cys25 of DS151 makes a minor con tribution through a longer-range cross link with Cys374 of actin, which likely involves flexibility of both proteins in that region.Key words: actin-binding proteins, homology modeling, chemical cross linking, actin-severing proteins.
A study of the relationships of interactions between Asp-201, Na+or K+, and galactosyl C6 hydroxyl and their effects on binding and reactivity of β-galactosidase
Authors: Xu Julia , McRae Mary A.A , Harron Scott , Rob Beatrice , Huber Reuben E
Published: April 2004

The interactions between Na+(and K+) and Asp-201 of β-galactosidase were studied. Analysis of the changes in Kmand Vmaxshowed that the Kdfor Na+of wild type β-galactosidase (0.36 ± 0.09 mM) was about 10× lower than for K+(3.9 ± 0.6 mM). The difference is probably because of the size and other physical properties of the ions and the binding pocket. Decreases of Kmas functions of Na+and K+for oNPG and pNPG and decreases of the Kiof both shallow and deep mode inhibitors were similar, whereas the Kmand Kiof substrates and inhibitors without C6 hydroxyls remained constant. Thus, Na+and K+are important for binding galactosyl moieties via the C6 hydroxyl throughout catalysis. Na+and K+had lesser effects on the Vmax. The Vmaxof pNPF and pNPA (substrates that lack a C6 hydroxyl) did not change upon addition of Na+or K+, showing that the catalytic effects are also mediated via the C6 hydroxyl. Arrhenius plots indicated that Na+, but not K+, caused k3(degalactosylation) to increase. Na+also caused the k2(galactosylation) with oNPG, but not with pNPG, to increase. In contrast, K+caused the k2values with both oNPG and pNPG to increase. Na+and K+mainly altered the entropies of activation of k2and k3with only small effects on the enthalpies of activation. This strongly suggests that only the positioning of the substrate, transition states, and covalent intermediate are altered by Na+and K+. Further evidence that positioning is important was that substitution of Asp-201 with a Glu caused the Kmand Kivalues to increase significantly. In addition, the Kdvalues for Na+or K+were 5 to 8 fold higher. The negative charge of Asp-201 was shown to be vital for Na+and K+binding. Large amounts of Na+or K+had no effect on the very large Kmand Kivalues of D201N-β-galactosidase and the Vmaxvalues changed minimally and in a linear rather than hyperbolic way. D201F-β-galactosidase, with a very bulky hydrophobic side chain in place of Asp, essentially obliterated all binding and catalysis.Key words: β-galactosidase, sodium, potassium, binding, aspartic acid.
Implementing artificial neural network models for real-time water colour forecasting in a water treatment plant
Authors: Zhang Qing J , Cudrak Audrey A , Shariff Riyaz , Stanley Stephen J
Published: April 2004

Artificial neural network (ANN) technology has evolved from the experimental stage into actual industrial applications. To achieve this significant transition, careful planning and adjustment are required. This paper illustrates such an example in the water treatment industry. The project objective is to upgrade the ANN models from a previous research project and install the system on-line in the Rossdale Water Treatment Plant in Edmonton, Alberta, Canada, to forecast raw water colour one day ahead. The article discusses the important issues and techniques to upgrade the neural network model to the actual application. Furthermore, sufficient communication is also required between the designers and the users to address the applicability and user friendly issues in model implementation. Failure in communication can render the whole process ineffective. Possible improvements are also recommended for the future on-line applications. Key words: artificial neural network, river raw water, forecasting, water treatment.
Application of back-propagation neural network modeling for free residual chlorine, total trihalomethanes and trihalomethanes speciation
Authors: Rodriguez Manuel J , Sérodes Jean-B.
Published: April 2004

The application of back-propagation neural networks (BPNNs) was assessed for modeling residual chlorine decay, total THM concentrations (TTHMs), and three individual THM species (CHCl3, CHBrCl2, and CHBr2Cl) in water that was chlorinated under laboratory-scale conditions. Data for modeling chlorine decay and TTHM were generated in chlorination experiments carried out with water collected in water utilities of the Quebec City region, whereas data for THM species were provided by the US Geological Survey for the Mississippi River and its tributaries. The BPNN models were compared with conventional models developed with exactly the same data. Results showed that the ability of BPNN to model residual chlorine decay is, in general terms, comparable to the ability of kinetic first- and second-order models. For TTHMs and THM speciation, however, the ability of BPNN was clearly higher in comparison with multivariate regression models, in particular when brominated disinfection by-products (DBPs) (CHBrCl2 and CHBr2Cl) were modeled. The successful application of BPNN presented in this study opens the door to other potential applications of BPNN for field-scale data concerning THMs as well as for other relevant disinfection by-products. Key words: back-propagation neural networks, drinking water, chlorination, residual chlorine decay, trihalomethanes, prediction.
Modeling of hourly NOx concentrations using artificial neural networks
Authors: Hasham Faizal A , Kindzierski Warren B , Stanley Stephen J
Published: April 2004

Modeling of ambient air quality is an important component of urban air quality management. Artificial neural network (ANN) modeling may offer advantages in understanding processes that follow nonlinear, complex relationships. Artificial neural network modeling is a black-box method where relationships describing complex situations are not necessarily known. The ANN models learn patterns based on historical data, and then conduct simulations based upon these patterns. The objective of this study was to evaluate the feasibility of using an ANN to predict ambient hourly concentrations of oxides of nitrogen (NOx) in an industrial corridor adjacent to Edmonton, Alberta. A standard 4-layer back-propagation network was used to predict ambient hourly NOx concentrations using industry stack emission rates, meteorological data, and traffic counts as input variables. The resulting model fit (R2 of 0.63) and precision of model prediction (root mean square error of 1.8 × 103 ppm as NO2) suggested that ANN modeling shows promise for predicting NOx behaviour; however, further work is necessary to improve its forecasting ability.Key words: urban air quality, airshed, modeling, artificial neural network.
A neural network approach to selecting indicators for a sustainable ecosystem
Authors: Rowland J , Andrews W S , Creber K A.M
Published: April 2004

The determination of indicators for monitoring the natural environment to ensure the ecological sustainability of areas under stress is problematic and prone to bias. Further, because of limitations in the resources available for data collection, such indicators should be selected carefully and their number confined to the minimum required to effectively monitor the system under study. Statistical techniques have traditionally been used to help select indicators, but often data lack the requirements of valid statistical analyses: minimal noise, variables linearly separable, variables able to be assigned numerical values, and high dimensionality. In this study, an alternate robust technique, artificial neural networks, is used to examine ecosystem data in multidimensional space and to select the minimum number of measured indicators that have the greatest weight in maintaining a sustainable ecosystem. The case study involves selecting indicators for ensuring the completion of ecologically sustainable army training conducted in a mixed grass prairie ecosystem. A neural network model was created that reduced the number of required measured indicators from 62 to 12, while minimizing researcher bias. Key words: ecological sustainability indicators, neural networks, military training.
Artificial neural network modelling of oil sands extraction processes
Authors: Zhang Qing J , Sawatzky Ronald P , Wallace E Dean , London Michael J , Stanley Stephen J
Published: April 2004

Although the artificial neural network (ANN) approach has been used in various disciplines of engineering since the 1980s, its use in the oil sands extraction industry is quite new and has great potential. This paper demonstrates two important ANN modelling techniques that will be very useful to the oil sand industry through a case study. First, the ANN pattern recognition approach is illustrated to categorize a large oil sand processing database consisting of experimental data from the last 20 years. Second, within each category, the authors demonstrate how to follow a general protocol to build ANN models that are capable of predicting the primary recovery and the primary froth quality for the oil sand treatment process with fairly good accuracy. To verify the reliability of the ANN models, besides the regular statistical and graphical analysis, the authors also conducted a sensitivity analysis to test the response logic, parameter interaction, and extrapolation capability of the ANN models. As shown in the paper, these tests are both satisfactory and interesting. With sufficient accuracy and robustness in performance, these ANN models can be used to evaluate both the qualitative and the quantitative response of the oil sand treatment process to the known key parameters, which, in turn, can then be used to optimize the oil sand treatment process. Furthermore, these ANN models can be linked with the geological survey results to estimate the production potential of an oil sand ore field and to manage the cost of stockpiling the chemicals for the treatment process.Key words: oil sand processing, artificial neural network, pattern recognition.
Genetic and phenotypic traits of streptomycetes used to characterize antibiotic activities of field-collected microbes
Authors: Davelos Anita L , Xiao Kun , Flor Jennifer M , Kinkel Linda L
Published: April 2004

Although antibiotic production may contribute significantly to microbial fitness, there is limited information on the ecology of antibiotic-producing microbial populations in soil. Indeed, quantitative information on the variation in frequency and intensity of specific antibiotic inhibitory and resistance abilities within soil microbial communities is lacking. Among the streptomycetes, antibiotic production is highly variable and resistance to antibiotics is highly specific to individual microbial strains. The objective of this work was to genetically and phenotypically characterize a reference collection of streptomycetes for use in distinguishing inhibition and resistance phenotypes of field-collected microbes. Specifically, we examined inhibition and resistance abilities of all isolates in all possible pairwise combinations, genetic relatedness using BOX-PCR and 16S rDNA sequence analyses, nutrient utilization profiles, and antibiotic induction among all possible three-way combinations of isolates. Each streptomycete isolate possessed a unique set of phenotypic and genetic characteristics. However, there was little correspondence between phenotypic and genetic traits. This collection of reference isolates provides the potential for distinguishing 1024 inhibition and resistance phenotypes in field-collected microbes. Relationships between the genetic and phenotypic characteristics examined may provide preliminary insight into the distinct strategies that microbes use in optimizing their fitness in natural environments.Key words: antibiotic inhibition, resistance, nutrient utilization, BOX-PCR, 16S rDNA.
Expression and purification of immunologically reactive DPPD, a recombinantMycobacterium tuberculosisskin test antigen, usingMycobacterium smegmatisandEscherichia colihost cells
Authors: Liu Chao , Flamoe Eric , Chen Hong-Jing , Carter Darrick , Reed Steven G , Campos-Neto Antonio
Published: April 2004

DPPD is a Mycobacterium tuberculosis recombinant antigen that elicits specific delayed type hypersensitivity reactions similar in size and morphological aspects to that elicited by purified protein derivative, in both guinea pigs and humans infected with M. tuberculosis. In addition, earlier clinical studies with DPPD suggested that this molecule could improve the specificity of the tuberculin skin test, which is used as an important aid for the diagnosis of tuberculosis. However, these studies could only be performed with DPPD engineered as a fusion molecule with another Mycobacterium spp. protein because no expression of DPPD could be achieved as a single molecule or as a conventional fusion protein in any commercial system. Although recombinant fusion proteins are in general suitable for several biological studies, they are by definition not ideal for studies involving highly purified and defined polypeptide sequences. Here, we report two alternative approaches for the expression of immunologically reactive recombinant genuine DPPD. The first approach used the rapidly growing, nonpathogenic Mycobacterium smegmatis as host cells transformed with the pSMT3 plasmid vector containing the full-length DPPD gene. The second approach used Escherichia coli transformed with the pET-17b plasmid vector containing the DPPD gene engineered in a three-copy fusion manner in tandem with itself. Though at low levels, expression and purification of immunologically reactive DPPD in M. smegmatis could be achieved. More abundant expression and purification of DPPD as a homo-trimer molecule was achieved in E. coli ([Formula: see text]2 mg/L of bacterial broth cultures). Interestingly, expression could only be achieved in host cells transformed with the DPPD gene containing its leader peptide. However, the expressed proteins lacked the leader sequence, which indicates that processing of the M. tuberculosis DPPD gene was accurately achieved and necessary in both M. smegmatis and E. coli. More importantly, the delayed type hypersensitivity reactions elicited by purified molecules in guinea pigs infected with M. tuberculosis were indistinguishable from that elicited by purified protein derivative. Because the DPPD gene is present only in the tuberculosis-complex organisms of the Mycobacterium genus, these highly purified molecules should be helpful in identifying individuals sensitized with tubercle bacilli.Key words: Mycobacterium tuberculosis, Mycobacterium smegmatis skin test, DTH, DPPD.