Influences of upland timber harvest on aquatic invertebrate communities in seasonal ponds: efficacy of forested buffers

We assessed community responses of aquatic invertebrates in 16 small, seasonal ponds in a forested region of north central Minnesota, USA, to evaluate potential influences of timber harvest and efficacy of uncut forested buffers in adjacent uplands. Invertebrate data gathered before (2000) and during the first 4 years following clearcut timber harvest (2001–2004) indicated that tree removal was followed by shifts in aquatic invertebrate communities in adjacent seasonal ponds. Retention of forested buffers appeared to partially mitigate influences of tree removal, but benefits of buffers may be limited by wind throw or other factors. Additional research is needed to clarify relationships between ecological characteristics of seasonal ponds and upland silviculture activities, and to better document efficacy and longevity of forested buffers.     

A community-based framework for aquatic ecosystem models

Here, we communicate a point of departure in the development of aquatic ecosystem models, namely a new community-based framework, which supports an enhanced and transparent union between the collective expertise that exists in the communities of traditional ecologists and model developers. Through a literature survey, we document the growing importance of numerical aquatic ecosystem models while also noting the difficulties, up until now, of the aquatic scientific community to make significant advances in these models during the past two decades. Through a common forum for aquatic ecosystem modellers we aim to (i) advance collaboration within the aquatic ecosystem modelling community, (ii) enable increased use of models for research, policy and ecosystem-based management, (iii) facilitate a collective framework using common (standardised) code to ensure that model development is incremental, (iv) increase the transparency of model structure, assumptions and techniques, (v) achieve a greater understanding of aquatic ecosystem functioning, (vi) increase the reliability of predictions by aquatic ecosystem models, (vii) stimulate model inter-comparisons including differing model approaches, and (viii) avoid ??re-inventing the wheel??, thus accelerating improvements to aquatic ecosystem models. We intend to achieve this as a community that fosters interactions amongst ecologists and model developers. Further, we outline scientific topics recently articulated by the scientific community, which lend themselves well to being addressed by integrative modelling approaches and serve to motivate the progress and implementation of an open source model framework.     

Neuronal protein NP 185 is developmentally regulated,initially expressed during synaptogenesis,and localized in synaptic terminals

Evidence is presented here that demonstrates the presence of NP185 (AP₃) in neuronal cells, specifically within syn-aptic terminals of the central nervous system and in the peripheral nervous system, particularly in the neuro-muscular junction of adult chicken muscle. Biochemical results obtained in our laboratories indicate that NP185 is associated with brain synaptic vesicles, with clathrin-coated vesicles, and with the synaptosomal plasma membrane. Also, NP185 binds to tubulin and clathrin light chains and the binding is regulated by phosphorylation (Su et al., 1991). Based on these properties and the data reported here, we advance the postulate that NP185 fulfills multiple functions in synaptic terminals. One function is that of a plasma membrane docking or channel protein, another of a signaling molecule for brain vesicles to reach the synaptic terminal region, and a third is that of a recycling molecule by binding to protein components on the lipid bilayer of the synaptic plasma membrane during the process of endocytosis. In support of these premises, a thorough study of NP185 using the developing chick brain, adult mouse brain, and chicken straited muscle was begun by temporally and spatially mapping the expression and localization of NP185 in evolving and mature nerve endings. To achieve these objectives, monoclonal antibodies to NP185 were used for immunocytochemistry in tissue sections of chicken and mouse cerebella. The distribution of NP185 was compared with those of other cytoskeletal and cytoplasmic proteins of axons and synapses, namely synaptophysin, vimentin, neurofilament NF68, and the intermediate filaments of glial cells (GFAP). The data indicate that expression of NP185 temporally coincides with synaptogenesis, and that the distribution of this protein is specific for synaptic terminal buttons of the CNS and the PNS.     

The effect of arrestin conformation on the recruitment of c-Raf1, MEK1, and ERK1/2 activation

Arrestins are multifunctional signaling adaptors originally discovered as proteins that "arrest" G protein activation by G protein-coupled receptors (GPCRs). Recently GPCR complexes with arrestins have been proposed to activate G protein-independent signaling pathways. In particular, arrestin-dependent activation of extracellular signal-regulated kinase 1/2 (ERK1/2) has been demonstrated. Here we have performed in vitro binding assays with pure proteins to demonstrate for the first time that ERK2 directly binds free arrestin-2 and -3, as well as receptor-associated arrestins-1, -2, and -3. In addition, we showed that in COS-7 cells arrestin-2 and -3 association with β(2)-adrenergic receptor (β2AR) significantly enhanced ERK2 binding, but showed little effect on arrestin interactions with the upstream kinases c-Raf1 and MEK1. Arrestins exist in three conformational states: free, receptor-bound, and microtubule-associated. Using conformationally biased arrestin mutants we found that ERK2 preferentially binds two of these: the "constitutively inactive" arrestin-Δ7 mimicking microtubule-bound state and arrestin-3A, a mimic of the receptor-bound conformation. Both rescue arrestin-mediated ERK1/2/activation in arrestin-2/3 double knockout fibroblasts. We also found that arrestin-2-c-Raf1 interaction is enhanced by receptor binding, whereas arrestin-3-c-Raf1 interaction is not.     

Edited transcripts compete with unedited mRNAs for trans-acting editing factors in higher plant chloroplasts

The dog, as a model for cardiovascular function, has been widely used in the pharmacological analysis of PDE inhibitors, particularly those thought to target the heart. However biochemical analyses of dog heart PDE have been largely performed on mixed enzyme populations, sequence information is lacking and no PDE from dog heart has been cloned. We have characterized a completely purified PDE1 enzyme from dog heart using dye-affinity, Mono-Q and calmodulin-affinity chromatography. The enzyme was stimulated 3-4-fold by calmodulin ([S]=0.5 microM) and, in the absence of calmodulin, exhibited biphasic kinetics with a low K(m) of 1.2 microM and 0.53 microM for cAMP and cGMP, with respective V(max) values of 283 and 146 nmoles min(-1) mg(-1). Internal peptides from this enzyme were used to design degenerate PCR primers. Subsequent 3'-RACE, 5'-RACE and high fidelity PCR were then used to produce a full length gene identified as PDE1A1 by sequence identity to human and bovine sequences. Northern analysis using the dog heart cDNA as a probe suggested the presence of an additional form of PDE1, in heart only, separate from the PDE1A group which was present in both heart and skeletal muscle. Multiple forms of human PDE1A are known to exist and PDE1B is present in human heart muscle. The findings here extend the PDE1 data to the dog and contribute to our understanding of the molecular biology of PDE1A in this species.     

Hemagglutinin Receptor Specificity and Structural Analyses of Respiratory Droplet-Transmissible H5N1 Viruses

Two ferret-adapted H5N1 viruses capable of respiratory droplet transmission have been reported with mutations in the hemagglutinin receptor-binding site and stalk domains. Glycan microarray analysis reveals that both viruses exhibit a strong shift toward binding to “human-type” α2-6 sialosides but with notable differences in fine specificity. Crystal structure analysis further shows that the stalk mutation causes no obvious perturbation of the receptor-binding pocket, consistent with its impact on hemagglutinin stability without affecting receptor specificity.     

CD4-directed peptide vaccination augments an antitumor response, but efficacy is limited by the number of CD8+ T cell precursors

Peptide vaccination is an immunotherapeutic strategy being pursued as a method of enhancing Ag-specific antitumor responses. To date, most studies have focused on the use of MHC class I-restricted peptides, and have not shown a correlation between Ag-specific CD8(+) T cell expansion and the generation of protective immune responses. We investigated the effects of CD4-directed peptide vaccination on the ability of CD8(+) T cells to mount protective antitumor responses in the DUC18/CMS5 tumor model system. To accomplish this, we extended the amino acid sequence of the known MHC class I-restricted DUC18 rejection epitope from CMS5 to allow binding to MHC class II molecules. Immunization with this peptide (tumor-derived extracellular signal-regulated kinase-II (tERK-II)) induced Ag-specific CD4(+) T cell effector function, but did not directly prime CD8(+) T cells. Approximately 31% of BALB/c mice immunized with tERK-II were protected from subsequent tumor challenge in a CD40-dependent manner. Priming of endogenous CD8(+) T cells in immunized mice was detected only after CMS5 challenge. Heightened CD4(+) Th cell function in response to tERK II vaccination allowed a 12-fold reduction in the number of adoptively transferred CD8(+) DUC18 T cells needed to protect recipients against tumor challenge as compared with previous studies using unimmunized mice. Furthermore, tERK-II immunization led to a more rapid and transient expansion of transferred DUC18 T cells than was seen in unimmunized mice. These findings illustrate that CD4-directed peptide vaccination augments antitumor immunity, but that the number of tumor-specific precursor CD8(+) T cells will ultimately dictate the success of immunotherapy.     

Testing for parallel allochronic isolation in lake–stream stickleback

The evolution of reproductive isolation (RI) is a critical step shaping progress towards speciation. In the context of ecological speciation, a critical question is the extent to which specific reproductive barriers important to RI evolve rapidly and predictably in response to environmental differences. Only reproductive barriers with these properties (importance, rapidity, predictability) will drive the diversification of species that are cohesively structured by environment type. One candidate barrier that might exhibit such properties is allochrony, whereby populations breed at different times. We studied six independent lake–stream population pairs of threespine stickleback (Gasterosteus aculeatus Linnaeus, 1758) that are known from genetic studies to show RI. However, the specific reproductive barriers driving this RI have proven elusive, leading to a ‘conundrum of missing reproductive isolation’. We here show that breeding times differ among some of the populations, but not in a consistent manner between lakes and streams. Moreover, the timing differences between lake and stream populations within each pair could account for only a small proportion of total RI measured with neutral genetic markers. Allochrony cannot solve the conundrum of missing reproductive isolation in lake–stream stickleback.