Real-time imaging of the dynamics of secretory granules in growth cones

Secretory granules containing a hybrid protein consisting of the regulated secretory protein tissue plasminogen activator and an enhanced form of green fluorescent protein were tracked at high spatial resolution in growth cones of differentiated PC12 cells. Tracking shows that granules, unlike synaptic vesicles, generally are mobile in growth cones. Quantitative analysis of trajectories generated by granules revealed two dominant modes of motion: diffusive and directed. Diffusive motion was observed primarily in central and peripheral parts of growth cones, where most granules diffused two to four orders of magnitude more slowly than comparably sized spheres in dilute solution. Directed motion was observed primarily in proximal parts of growth cones, where a subset of granules underwent rapid, directed motion at average speeds comparable to those observed for granules in neurites. This high-resolution view of the dynamics of secretory granules in growth cones provides insight into granule organization and release at nerve terminals. In particular, the mobility of granules suggests that granules, unlike synaptic vesicles, are not tethered stably to cytoskeletal structures in nerve terminals. Moreover, the slow diffusive nature of this mobility suggests that secretory responses involving centrally distributed granules in growth cones will occur slowly, on a time scale of minutes or longer.     

Identifying the target cell in primary simian immunodeficiency virus (SIV) infection: highly activated memory CD4(+) T cells are rapidly eliminated in early SIV infection in vivo

It has recently been shown that rapid and profound CD4(+) T-cell depletion occurs almost exclusively within the intestinal tract of simian immunodeficiency virus (SIV)-infected macaques within days of infection. Here we demonstrate (by three- and four-color flow cytometry) that this depletion is specific to a definable subset of CD4(+) T cells, namely, those having both a highly and/or acutely activated (CD69(+) CD38(+) HLA-DR(+)) and memory (CD45RA(-) Leu8(-)) phenotype. Moreover, we demonstrate that this subset of helper T cells is found primarily within the intestinal lamina propria. Viral tropism for this particular cell type (which has been previously suggested by various studies in vitro) could explain why profound CD4(+) T-cell depletion occurs in the intestine and not in peripheral lymphoid tissues in early SIV infection. Furthermore, we demonstrate that an acute loss of this specific subset of activated memory CD4(+) T cells may also be detected in peripheral blood and lymph nodes in early SIV infection. However, since this particular cell type is present in such small numbers in circulation, its loss does not significantly affect total CD4(+) T cell counts. This finding suggests that SIV and, presumably, human immunodeficiency virus specifically infect, replicate in, and eliminate definable subsets of CD4(+) T cells in vivo.     

Activation of lymphocyte signaling by the R1 protein of rhesus monkey rhadinovirus

Rhesus monkey rhadinovirus (RRV) is a gamma-2 herpesvirus that exhibits a considerable degree of similarity to the human Kaposi's sarcoma-associated herpesvirus (KSHV). The R1 protein of RRV is distantly related to the K1 protein of KSHV, and R1, like K1, can contribute to cell growth transformation. In this study we analyzed the ability of the cytoplasmic tail of R1 to function as a signal transducer. The cytoplasmic domain of the R1 protein contains several tyrosine residues whose phosphorylation is induced in cells expressing Syk kinase. Expression of a CD8 chimera protein containing the extracellular and transmembrane domains of CD8 fused to the cytoplasmic domain of R1 mobilized intracellular calcium and induced cellular tyrosine phosphorylation in B cells upon stimulation with anti-CD8 antibody. None of the CD8-R1 cytoplasmic deletion mutants tested were able to mobilize intracellular calcium or to induce tyrosine phosphorylation to a significant extent upon addition of anti-CD8 antibody. Expression of wild-type R1 protein activated nuclear factor of activated T lymphocytes (NFAT) eightfold in B cells in the absence of antibody stimulation; expression of the CD8-R1C chimera strongly induced NFAT activity (60-fold) but only upon the addition of anti-CD8 antibody. We conclude that the cytoplasmic domain of R1 is capable of transducing signals that elicit B-lymphocyte activation events. The signal-inducing properties of R1 appear to be similar to those of K1 but differ in that the required sequences are distributed over a much longer stretch of the cytoplasmic domain (>150 amino acids). In addition, the induction of calcium mobilization was considerably longer in duration and stronger with R1 than with K1.     

A limited survey of aflatoxins and fumonisins in retail maizebased products in the UK using immunoassay detection

A total of 27 maize-based products destined for human consumption were collected from retail outlets within the city of Glasgow in the UK and were analysed for the presence of aflatoxins using immunoaftinity column chromatography with fluorescence detection and for fumonisins by competitive ELISA. Aflatoxins were detected at a trace level below 4 in eight (30%) of the 27 samples tested, no sample contained aflatoxins at a high level although one sample of sweetcorn did contain aflatoxins at a level of 5-10 Fumonisins were detected in eight (30%) of the samples at levels from 1 to 8mgkg^-1 and a further eight samples contained fumonisin at a level below 1 mgkg^-1 but above the detectable level. The highest concentration of fumonisins was found in a sample of fine corn meal at 8-12mgkg^-1.     

Partial protein domains: evolutionary insights and bioinformatics challenges

Protein domains are generally thought to correspond to units of evolution. New research raises questions about how such domains are defined with bioinformatics tools and sheds light on how evolution has enabled partial domains to be viable.With the rapid expansion in the number of determined protein sequences - over 92 million in UniProt in March 2015 - an ever-increasing number of biologists are using bioinformatics tools for annotation of these sequences. One widely used strategy is to identify occurrences of Pfam families within the sequence of interest [1]. A Pfam family is a multiple sequence alignment of the occurrences of a particular domain both in different species and in different regions of the same protein. The concept underpinning Pfam is that proteins typically comprise one or more domains (regions), each of which is an evolutionary unit that generally has a well-defined biological function. A significant sequence similarity between a query protein and a Pfam family provides the basis for annotations. Two recent articles [2,3] in Genome Biology evaluate the implications of having the query sequence only matching part of a Pfam family, which is an intriguing finding, given that a Pfam family is considered to be an evolutionary unit.     

The evolutionary emergence of stochastic phenotype switching in bacteria

Stochastic phenotype switching - or bet hedging - is a pervasive feature of living systems and common in bacteria that experience fluctuating (unpredictable) environmental conditions. Under such conditions, the capacity to generate variable offspring spreads the risk of being maladapted in the present environment, against offspring likely to have some chance of survival in the future. While a rich subject for theoretical studies, little is known about the selective causes responsible for the evolutionary emergence of stochastic phenotype switching. Here we review recent work - both theoretical and experimental - that sheds light on ecological factors that favour switching types over non-switching types. Of particular relevance is an experiment that provided evidence for an adaptive origin of stochastic phenotype switching by subjecting bacterial populations to a selective regime that mimicked essential features of the host immune response. Central to the emergence of switching types was frequent imposition of 'exclusion rules' and 'population bottlenecks' - two complementary faces of frequency dependent selection. While features of the immune response, exclusion rules and bottlenecks are likely to operate in many natural environments. Together these factors define a set of selective conditions relevant to the evolution of stochastic switching, including antigenic variation and bacterial persistence.     

Scats and den contents as indicators of the diet of stoats (Mustela erminea) in the Tasman Valley,South Canterbury,New Zealand

Stoats are significant predators of native fauna in New Zealand. They occur in many habitat types and consume a wide range of prey. The diet of stoats in the Tasman River, South Canterbury, was studied by analysis of scats and den contents. Analysis of 206 scats showed that stoats ate mainly lagomorphs, birds and invertebrates. Minor components included mice, lizards, fish and hedgehogs. Stoats ate more birds in spring than in autumn, and female stoats ate more invertebrates than did males. The contents of 219 dens collected in the same area at the same time provided further information. Birds and lagomorphs occurred at high frequency in dens, and other components were minor. Remains in dens were larger than in scats and allowed identification of many more prey items to species level. Den contents revealed a potentially substantial impact of stoats on threatened shorebirds locally; this impact was not detected by analysis of scats.     

The first 10 years of the dialysis-transplantation program at The Hospital for Sick Children,Toronto. 1: Predialysis and dialysis.

Renal dialysis and transplantation have been used for many years for adults with kidney failure but only recently for children. In May 1967 a renal-dialysis-transplantation program was established at The Hospital for Sick Children, Toronto for patients aged 6 to 18 years living within 240 km of Toronto. In 1973, children aged 1 to 5 years began to be accepted into the program, and by August 1977, 90 children (mean age 11 years) from all parts of Canada had been admitted to the program. The creation of vascular access in very small patients is difficult; the most successful types of access have been central shunts (established above the knee or the elbow) and bovine grafts. Specially made dialysis equipment is necessary for young patients. Young children should only be accepted in a dialysis-transplantation program that has a medical staff expert in meeting the specific needs of such children.