Variants of human immunodeficiency virus type 1 that efficiently use CCR5 lacking the tyrosine-sulfated amino terminus have adaptive mutations in gp120, including loss of a functional N-glycan

By selecting the R5 human immunodeficiency virus type 1 (HIV-1) strain JR-CSF for efficient use of a CCR5 coreceptor with a badly damaged amino terminus [i.e., CCR5(Y14N)], we previously isolated variants that weakly utilize CCR5(Delta18), a low-affinity mutant lacking the normal tyrosine sulfate-containing amino-terminal region of the coreceptor. These previously isolated HIV-1(JR-CSF) variants contained adaptive mutations situated exclusively in the V3 loop of their gp120 envelope glycoproteins. We now have weaned the virus from all dependency on the CCR5 amino terminus by performing additional selections with HeLa-CD4 cells that express only a low concentration of CCR5(Delta18). The adapted variants had additional mutations in their V3 loops, as well as one in the V2 stem (S193N) and four alternative mutations in the V4 loop that eliminated the same N-linked oligosaccharide from position N403. Assays using pseudotyped viruses suggested that these new gp120 mutations all made strong contributions to use of CCR5(Delta18) by accelerating a rate-limiting CCR5-dependent conformational change in gp41 rather than by increasing viral affinity for this damaged coreceptor. Consistent with this interpretation, loss of the V4 N-glycan at position N403 also enhanced HIV-1 use of a different low-affinity CCR5 coreceptor with a mutation in extracellular loop 2 (ECL2) [i.e., CCR5(G163R)], whereas the double mutant CCR5(Delta18,G163R) was inactive. We conclude that loss of the N-glycan at position N403 helps to convert the HIV-1 envelope into a hair-trigger form that no longer requires strong interactions with both the CCR5 amino terminus and ECL2 but efficiently uses either site alone. These results demonstrate a novel functional role for a gp120 N-linked oligosaccharide and a high degree of adaptability in coreceptor usage by HIV-1.     

Long-term effects of hydropower installations and associated river regulation on River Shannon eel populations: mitigation and management

The Shannon, Ireland’s largest river, is used for hydroelectricity generation since 1929. Subsequently, the Electricity Supply Board assumed responsibility for management of its eel stocks, due to the impact of the hydro-dam on recruitment to the commercial fishery. In order to negate a decline in juvenile recruitment resulting from the installation of hydroelectric facilities, management was focused on stocking lakes with elvers and fingerling eels. These were trapped at the hydropower facilities and in estuarine tributaries during their up-stream migrations. Due to the decline of natural recruitment in more recent times, attempts have also been made to develop an estuarine glass eel fishery. Stock levels are then monitored through annual surveys of the population trends of juvenile (glass eel, elver), growing phase (yellow eel) and downstream migrating pre-spawners (silver eels). Survey results and fishery management programmes are reviewed in this article. In addition to the long-term effects the hydroelectric facilities have had on the stock levels, there is also an annual effect on the migratory patterns of downstream migratory silver eels. In the lower reaches of the river system flow rates are regulated by the hydroelectric stations. We review previous work that had highlighted the importance of flow in determining the timing of the silver eels migrations, and assess the relationship between flow and migration in more detail through the use of hydroacoustic and telemetric studies. Current research on seaward migrating silver eel populations, suggests that spawner escapement rates can most effectively be increased by trapping migrating eels at fishing weirs located up-stream of the power station and transporting them towards the estuary. Guest editors: R. L. Welcomme & G. Marmulla Hydropower, Flood Control and Water Abstraction: Implications for Fish and Fisheries     

Testing the antagonistic effect of follistatin on BMP family members in ovine granulosa cells

Follistatin was first demonstrated as an activin-binding protein, neutralizing its actions. However, there is emerging evidence that follistatin inhibits the action of other members of the transforming growth factor beta(TGFbeta) / bone morphogenetic protein (BMP) superfamily. Recently, numerous BMP factors have been shown to play important roles in regulating folliculogenesis and ovulation rate in mammals, and such a potential antagonistic role of follistatin is of particular interest in the context of ovarian function. Using a biological test based on progesterone production by ovine primary granulosa cells in culture, we show that follistatin was a strong antagonist of activin A, but not BMP-2 or BMP-4 actions. In contrast, noggin, a known specific BMP antagonist, had no effect on activin A but strongly neutralized BMP-2 and BMP-4 actions. BMP-6 action was only slightly reduced by both follistatin and noggin. Our data led to the conclusion that follistatin would not represent a determinant physiological modulator of the biological effect of BMP factors on granulosa cells.     

Multidrug-resistant Salmonella enterica serovar paratyphi A harbors IncHI1 plasmids similar to those found in serovar typhi

Salmonella enterica serovars Typhi and Paratyphi A cause systemic infections in humans which are referred to as enteric fever. Multidrug-resistant (MDR) serovar Typhi isolates emerged in the 1980s, and in recent years MDR serovar Paratyphi A infections have become established as a significant problem across Asia. MDR in serovar Typhi is almost invariably associated with IncHI1 plasmids, but the genetic basis of MDR in serovar Paratyphi A has remained predominantly undefined. The DNA sequence of an IncHI1 plasmid, pAKU_1, encoding MDR in a serovar Paratyphi A strain has been determined. Significantly, this plasmid shares a common IncHI1-associated DNA backbone with the serovar Typhi plasmid pHCM1 and an S. enterica serovar Typhimurium plasmid pR27. Plasmids pAKU_1 and pHCM1 share 14 antibiotic resistance genes encoded within similar mobile elements, which appear to form a 24-kb composite transposon that has transferred as a single unit into different positions into their IncHI1 backbones. Thus, these plasmids have acquired similar antibiotic resistance genes independently via the horizontal transfer of mobile DNA elements. Furthermore, two IncHI1 plasmids from a Vietnamese isolate of serovar Typhi were found to contain features of the backbone sequence of pAKU_1 rather than pHCM1, with the composite transposon inserted in the same location as in the pAKU_1 sequence. Our data show that these serovar Typhi and Paratyphi A IncHI1 plasmids share highly conserved core DNA and have acquired similar mobile elements encoding antibiotic resistance genes in past decades.     

Genotypic and phenotypic diversity of rhizobia isolated from Lathyrus japonicus indigenous to Japan

Sixty-one rhizobial strains from Lathyrus japonicus nodules growing on the seashore in Japan were characterized and compared to two strains from Canada. The PCR-based method was used to identify test strains with novel taxonomic markers that were designed to discriminate between all known Lathyrus rhizobia. Three genomic groups (I, II, and III) were finally identified using RAPD, RFLP, and phylogenetic analyses. Strains in genomic group I (related to Rhizobium leguminosarum) were divided into two subgroups (Ia and Ib) and subgroup Ia was related to biovar viciae. Strains in subgroup Ib, which were all isolated from Japanese sea pea, belonged to a distinct group from other rhizobial groups in the recA phylogeny and PCR-based grouping, and were more tolerant to salt than the isolate from an inland legume. Test strains in genomic groups II and III belonged to a single clade with the reference strains of R. pisi, R. etli, and R. phaseoli in the 16S rRNA phylogeny. The PCR-based method and phylogenetic analysis of recA revealed that genomic group II was related to R. pisi. The analyses also showed that genomic group III harbored a mixed chromosomal sequence of different genomic groups, suggesting a recent horizontal gene transfer between diverse rhizobia. Although two Canadian strains belonged to subgroup Ia, molecular and physiological analyses showed the divergence between Canadian and Japanese strains. Phylogenetic analysis of nod genes divided the rhizobial strains into several groups that reflected the host range of rhizobia. Symbiosis between dispersing legumes and rhizobia at seashore is discussed.     

Plasma glycoprotein profiling for colorectal cancer biomarker identification by lectin glycoarray and lectin blot

Colorectal cancer (CRC) remains a major worldwide cause of cancer-related morbidity and mortality largely due to the insidious onset of the disease. The current clinical procedures utilized for disease diagnosis are invasive, unpleasant, and inconvenient; hence, the need for simple blood tests that could be used for the early detection of CRC. In this work, we have developed methods for glycoproteomics analysis to identify plasma markers with utility to assist in the detection of colorectal cancer (CRC). Following immunodepletion of the most abundant plasma proteins, the plasma N -linked glycoproteins were enriched using lectin affinity chromatography and subsequently further separated by nonporous silica reversed-phase (NPS-RP)-HPLC. Individual RP-HPLC fractions were printed on nitrocellulose coated slides which were then probed with lectins to determine glycan patterns in plasma samples from 9 normal, 5 adenoma, and 6 colorectal cancer patients. Statistical tools, including principal component analysis, hierarchical clustering, and Z-statistics analysis, were employed to identify distinctive glycosylation patterns. Patients diagnosed with colorectal cancer or adenomas were shown to have dramatically higher levels of sialylation and fucosylation as compared to normal controls. Plasma glycoproteins with aberrant glycosylation were identified by nano-LC-MS/MS, while a lectin blotting methodology was used to validate proteins with significantly altered glycosylation as a function of cancer progression. The potential markers identified in this study for diagnosis to distinguish colorectal cancer from adenoma and normal include elevated sialylation and fucosylation in complement C3, histidine-rich glycoprotein, and kininogen-1. These potential markers of colorectal cancer were subsequently validated by lectin blotting in an independent set of plasma samples obtained from 10 CRC patients, 10 patients with adenomas, and 10 normal subjects. These results demonstrate the utility of this strategy for the identification of N -linked glycan patterns as potential markers of CRC in human plasma, and may have the utility to distinguish different disease states.     

Nuclear membrane receptors for ET-1 in cardiovascular function

Plasma membrane endothelin type A (ET(A)) receptors are internalized and recycled to the plasma membrane, whereas endothelin type B (ET(B)) receptors undergo degradation and subsequent nuclear translocation. Recent studies show that G protein-coupled receptors (GPCRs) and ion transporters are also present and functional at the nuclear membranes of many cell types. Similarly to other GPCRs, ET(A) and ET(B) are present at both the plasma and nuclear membranes of several cardiovascular cell types, including human cardiac, vascular smooth muscle, endocardial endothelial, and vascular endothelial cells. The distribution and density of ET(A)Rs in the cytosol (including the cell membrane) and the nucleus (including the nuclear membranes) differ between these cell types. However, the localization and density of ET-1 and ET(B) receptors are similar in these cell types. The extracellular ET-1-induced increase in cytosolic ([Ca](c)) and nuclear ([Ca](n)) free Ca(2+) is associated with an increase of cytosolic and nuclear reactive oxygen species. The extracellular ET-1-induced increase of [Ca](c) and [Ca](n) as well as intracellular ET-1-induced increase of [Ca](n) are cell-type dependent. The type of ET-1 receptor mediating the extracellular ET-1-induced increase of [Ca](c) and [Ca](n) depends on the cell type. However, the cytosolic ET-1-induced increase of [Ca](n) does not depend on cell type. In conclusion, nuclear membranes' ET-1 receptors may play an important role in overall ET-1 action. These nuclear membrane ET-1 receptors could be targets for a new generation of antagonists.