Cloning and expression of a human kinesin heavy chain gene: interaction of the COOH-terminal domain with cytoplasmic microtubules in transfected CV-1 cells

To understand the interactions between the microtubule-based motor protein kinesin and intracellular components, we have expressed the kinesin heavy chain and its different domains in CV-1 monkey kidney epithelial cells and examined their distributions by immunofluorescence microscopy. For this study, we cloned and sequenced cDNAs encoding a kinesin heavy chain from a human placental library. The human kinesin heavy chain exhibits a high level of sequence identity to the previously cloned invertebrate kinesin heavy chains; homologies between the COOH-terminal domain of human and invertebrate kinesins and the nonmotor domain of the Aspergillus kinesin-like protein bimC were also found. The gene encoding the human kinesin heavy chain also contains a small upstream open reading frame in a G-C rich 5' untranslated region, features that are associated with translational regulation in certain mRNAs. After transient expression in CV-1 cells, the kinesin heavy chain showed both a diffuse distribution and a filamentous staining pattern that coaligned with microtubules but not vimentin intermediate filaments. Altering the number and distribution of microtubules with taxol or nocodazole produced corresponding changes in the localization of the expressed kinesin heavy chain. The expressed NH2-terminal motor and the COOH-terminal tail domains, but not the alpha-helical coiled coil rod domain, also colocalized with microtubules. The finding that both the kinesin motor and tail domains can interact with cytoplasmic microtubules raises the possibility that kinesin could crossbridge and induce sliding between microtubules under certain circumstances.     

Sorption of fluorescent polystyrene microplastic particles to edible seaweed <Emphasis Type="Italic">Fucus vesiculosus</Emphasis>

Increased global demands for food have raised interest for seaweed as a healthy and sustainable food source. At the same time, the large amounts of microplastic in the oceans have raised concern in relation to pollution of seafood including sea vegetables. The aim of this study was to examine sorption of fluorescent polystyrene (PS) microplastic particles to edible macroalga (seaweed) Fucus vesiculosus, and to investigate to what extent adsorbed PS particles could be washed off, using an industrial relevant method. PS microplastic particles (diameter of 20 μm) were used in a concentration of 2.65 mg L^?1 (corresponding to 597 particles per mL) in filtrated seawater (50 mL) to treat F. vesiculosus distal tips in blue cap flasks (100 mL) placed in a rotary box for 2 h. Results showed sorption of PS microplastic particles to F. vesiculosus analysed by microscopy and a significant reduction of 94.5% by washing. These results were based on high microplastic concentrations, not comparable to natural conditions/concentrations. Nonetheless, this study provides methodological and mechanistic insights into procedures for investigating the sorption of microplastics to seaweed, for which there is currently no established standardised method.     

Evolutionary relationship between human major histocompatibility complexHLA-DR haplotypes

 HLA-DR haplotypes of the human major histocompatibility complex are organized in five different groups. They can be identified based on the serological specificity expressed by the polymorphic DRB1 locus and by the presence of a characteristic set of DRB genes. The nucleotide sequences of introns 4 and 5 of the two DRB genes (DRB1 ^* 01 and DRB6 ^* 01) from a DR1 haplotype and the three DRB genes (DRB1 ^* 15, DRB6 ^* 15, and DRB5 ^* 15), from a DR51 haplotype were determined. This study identified endogenous retroviral long terminal repeat elements (ERV9 LTR) located at identical positions in intron 5 of the DRB1 genes in both the DR1 and DR51 haplotypes. Phylogenetic analyses revealed a close evolutionary relationship between these two haplotypes. The DRB5 gene, unique for the DR51 haplotype, may have been lost by a recent gene deletion event creating the DR1 haplotype. A model for the evolution of the human DR haplotypes involving separate duplication and contraction events is presented. Received: 10 October 1995 / Revised: 22 November 1995     

Spatial and temporal variability of saphrophytic bacteria in the surface waters of Paranaguá and Antonina Bays,Paraná, Brazil

Spatial and temporal variability in the concentration of saprophytic bacteria was investigated in surface waters of Paranaguá and Antonina bays.Seven points along a profile from the entrance of Paranaguá Bay to the innermost part of Antonina Bay, were sampled monthly from November 1985 to November 1986. Temperature, dissolved oxygen, pH and precipitation data were also measured.ZoBell 2216E culture medium was utilized with freshwater and 32% salinity water.Principal component analysis of biotic and abiotic data showed a gradient from the innermost bay to the baymouth bar with an increase in bacteria concentration at Antonina Bay. Significant variation was not observed among sampling stations at Paranaguá Bay, except for February 1986 when there was a homogeneous increase of bacteria throughout both bays in association with an increase in temperature and precipitation. There was also no strong variation of saprophyte numbers at sampling locations near Paranaguá city, despite domestic and harbour waste input.We conclude that the outer region of the estuary is highly influenced by the adjacent ocean and that the inner part has typical estuarine characteristics. The region may be considered little polluted.     

Primate DRB genes from the DR3 and DR8 haplotypes contain ERV9 LTR elements at identical positions

The HLA-DRB genes of the human major histocompatibility complex constitute a multigene family with a varying number of DRB genes in different haplotypes. To gain further knowledge concerning the evolutionary relationship, the complete nucleotide sequence was determined for a region spanning introns 4 and 5 of the three DRB genes (DRB1*0301, DRB2 and DRB3*0101) from a DR52 haplotype and the single DRB gene (DRB1*08021) in the DR8 haplotype. These analyses identified an endogenous retroviral long terminal repeat element (ERV9 LTR3), inserted at identical positions in intron 5 of the functional DRB genes in these two haplotypes. Comparison of the nucleotide sequence from introns 4 and 5 including the ERV9 LTR elements revealed a strong similarity between the three expressed DRB genes. The DRB3*0101 and DRB1*08021 genes were most similar in this comparison. These findings provide further evidence for a separate duplication in a primordial DR52 haplotype followed by a gene contraction event in the DR8 haplotype. A homologous element was found in a chimpanzee DRB gene from a DR52 haplotype. This represents the first characterized ERV9 LTR element in a nonhuman species. The corresponding introns of the DRB genes in the DR4 haplotype contain no ERV9 LTRs. In contrast, these genes have insertions of distinct Alu repeats, implying distinct evolutionary histories of DR52 and DR53 haplotypes, respectively. Phylogenetic analyses of DRB introns from DR52, DR53, and DR8 haplotypes showed a close relationship between the DRB2 and DRB4 genes. Thus, the ancestral DR haplotype that evolved to generate the DR52 and DR53 haplotypes most likely shared a primordial common DRB gene.The nucleotide sequence data reported in this paper have been submitted to the EMBL nucleotide sequence database and have been assigned the accession numbers X82660–X82663     

Prediction of ovarian response to exogenous gonadotropin stimulation: utilization for collection of primate oocytes for fertilization in vitro

The predictive value of 2 tests for ovarian response to controlled ovarian hyperstimulation in the cynomolgus monkey model was evaluated. The tests utilized were: 1) the cycle Day 3 (Day 1 = onset of menses) FSH value and 2) the acute estradiol (E(2)) response to a GnRH agonists (GnRHa) administered on Day 3. Both tests were performed during the cycle preceding control ovarian hyperstimulation. Subsequently, monkeys (n = 26) were stimulated with Metrodin(T) (Days 2-6, 25IU/d) and Pergonal(T) (Day 7 to hCG administration, 25IU/d). Laparoscopic oocyte retrieval was performed 32 to 34 after hCG administration. Analysis of the data revealed that Day 3 FSH values could not predict whether an animal would respond well to control ovarian hyperstimulation in a subsequent cycle (P = 0.77). However, the E(2) change 24 h post-GnRHa administration was significantly greater for animals responding well to control ovarian hyperstimulation compared with the animals deleted after 6 d of stimulation (P = 0.042). The mean change in E(2) levels in animals taken to aspiration was 97.8 pg/ml compared with only 21.6 pg/ml for the deleted animals. This differential response of E(2) production after GnRHa treatment was used to correctly identify (by discriminant analysis) 78% of the animals subsequently deleted for poor response. Thus, the increase in serum E(2) level after GnRHa, but not the basal FSH level, was found to be predictive of ovarian response to stimulation in the cynomolgus monkey.     

Comparison of analytical methods for profiling <Emphasis Type="Italic">N-</Emphasis> and <Emphasis Type="Italic">O-</Emphasis>linked glycans from cultured cell lines

The Human Disease Glycomics/Proteome Initiative (HGPI) is an activity in the Human Proteome Organization (HUPO) supported by leading researchers from international institutes and aims at development of disease-related glycomics/glycoproteomics analysis techniques. Since 2004, the initiative has conducted three pilot studies. The first two were N- and O-glycan analyses of purified transferrin and immunoglobulin-G and assessed the most appropriate analytical approach employed at the time. This paper describes the third study, which was conducted to compare different approaches for quantitation of N- and O-linked glycans attached to proteins in crude biological samples. The preliminary analysis on cell pellets resulted in wildly varied glycan profiles, which was probably the consequence of variations in the pre-processing sample preparation methodologies. However, the reproducibility of the data was not improved dramatically in the subsequent analysis on cell lysate fractions prepared in a specified method by one lab. The study demonstrated the difficulty of carrying out a complete analysis of the glycome in crude samples by any single technology and the importance of rigorous optimization of the course of analysis from preprocessing to data interpretation. It suggests that another collaborative study employing the latest technologies in this rapidly evolving field will help to realize the requirements of carrying out the large-scale analysis of glycoproteins in complex cell samples.     

Evolution of brain–body allometry in Lake Tanganyika cichlids

Brain size is strongly associated with body size in all vertebrates. This relationship has been hypothesized to be an important constraint on adaptive brain size evolution. The essential assumption behind this idea is that static (i.e., within species) brain–body allometry has low ability to evolve. However, recent studies have reported mixed support for this view. Here, we examine brain–body static allometry in Lake Tanganyika cichlids using a phylogenetic comparative framework. We found considerable variation in the static allometric intercept, which explained the majority of variation in absolute and relative brain size. In contrast, the slope of the brain–body static allometry had relatively low variation, which explained less variation in absolute and relative brain size compared to the intercept and body size. Further examination of the tempo and mode of evolution of static allometric parameters confirmed these observations. Moreover, the estimated evolutionary parameters indicate that the limited observed variation in the static allometric slope could be a result of strong stabilizing selection. Overall, our findings suggest that the brain–body static allometric slope may represent an evolutionary constraint in Lake Tanganyika cichlids.     

A Concept of Bayesian Regulation in Fisheries Management

Stochastic variability of biological processes and uncertainty of stock properties compel fisheries managers to look for tools to improve control over the stock. Inspired by animals exploiting hidden prey, we have taken a biomimetic approach combining catch and effort in a concept of Bayesian regulation (BR). The BR provides a real-time Bayesian stock estimate, and can operate without separate stock assessment. We compared the performance of BR with catch-only regulation (CR), alternatively operating with N-target (the stock size giving maximum sustainable yield, MSY) and F-target (the fishing mortality giving MSY) on a stock model of Baltic Sea herring. N-targeted BR gave 3% higher yields than F-targeted BR and CR, and 7% higher yields than N-targeted CR. The BRs reduced coefficient of variance (CV) in fishing mortality compared to CR by 99.6% (from 25.2 to 0.1) when operated with F-target, and by about 80% (from 158.4 to 68.4/70.1 depending on how the prior is set) in stock size when operated with N-target. Even though F-targeted fishery reduced CV in pre-harvest stock size by 19–22%, it increased the dominant period length of population fluctuations from 20 to 60–80 years. In contrast, N-targeted BR made the periodic variation more similar to white noise. We discuss the conditions when BRs can be suitable tools to achieve sustainable yields while minimizing undesirable fluctuations in stock size or fishing effort.