The influence of moss on grazers in high‐altitude streams: food,refuge or both?

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Effectiveness of specific RNA-mediated interference through ingested double-stranded RNA in Caenorhabditis elegans

Background  

In Caenorhabditis elegans, injection of double-stranded RNA (dsRNA) results in the specific inactivation of genes containing homologous sequences, a technique termed RNA-mediated interference (RNAi). It has previously been shown that RNAi can also be achieved by feeding worms Escherichia coli expressing dsRNA corresponding to a specific gene; this mode of dsRNA introduction is conventionally considered to be less efficient than direct injection, however, and has therefore seen limited use, even though it is considerably less labor-intensive.     

The Drosophila pericentrin-like protein is essential for cilia/flagella function, but appears to be dispensable for mitosis

Centrosomes consist of a pair of centrioles surrounded by an amorphous pericentriolar material (PCM). Proteins that contain a Pericentrin/AKAP450 centrosomal targeting (PACT) domain have been implicated in recruiting several proteins to the PCM. We show that the only PACT domain protein in Drosophila (the Drosophila pericentrin-like protein [D-PLP]) is associated with both the centrioles and the PCM, and is essential for the efficient centrosomal recruitment of all six PCM components that we tested. Surprisingly, however, all six PCM components are eventually recruited to centrosomes during mitosis in d-plp mutant cells, and mitosis is not dramatically perturbed. Although viable, d-plp mutant flies are severely uncoordinated, a phenotype usually associated with defects in mechanosensory neuron function. We show that the sensory cilia of these neurons are malformed and the neurons are nonfunctional in d-plp mutants. Moreover, the flagella in mutant sperm are nonmotile. Thus, D-PLP is essential for the formation of functional cilia and flagella in flies.     

Roles for 147 embryonic lethal genes on C.elegans chromosome I identified by RNA interference and video microscopy

Early embryonic development involves complex events such as the regulation of cell division and the establishment of embryonic polarity. To identify genes involved in these events, we collected four-dimensional time-lapse video recordings of the first three cell divisions and analysed terminal phenotypes after RNA interference of 147 embryonic lethal genes previously identified in a systematic screen of Caenorhabditis elegans chromosome I. Over half gave defects in early processes such as meiosis, the assembly or position of the first mitotic spindle, cytokinesis, and proper nuclear positioning. For some phenotypic classes, the majority of genes are involved in a shared biochemical process. In addition, we identified loss-of-function phenotypes for genes of unknown function, but for which homologues exist in other organisms, shedding light on the function of these uncharacterized genes. When applied to the whole genome, this approach should identify the vast majority of genes required for early cell processes, paving the way for a greatly improved understanding of these processes and their regulation at the molecular level.     

Stocking efficiency and the effects of diet preconditioning on the post-release adaptation of hatchery-reared juveniles of Atlantic salmon (Salmo salar L.) in an Atlantic temperate stream

This study explores the stocking efficiency of Atlantic salmon juveniles in a 4th order stream located at the southern limit of its Eastern Atlantic distribution. Moreover, with the idea of implementing new protocols and methods to improve ??traditional?? hatchery practices and rear a more ??wild?? like fish, the study examines the response of this species to natural diet preconditioning prior to release. The field study indicated that most juveniles disappeared from the stocked reaches within two months of their release, resulting in a recapture rate after a year of 0.0?C1.1?%. There were no differences in stocked juvenile??s density along the stream during the first months following stocking. The preconditioning experiments showed that juveniles of Atlantic salmon suffered a significant decrease in their condition factor when diet was changed from artificial pellets to live preys, either if the change occurred before (i.e., in a manipulative experiment) or after their release in the stream. Although weight, length, condition factor and specific growth rates reached by juveniles at the end of the rearing period were higher in individuals fed on pellets than in those fed on macroinvertebrates, differences disappeared only a week after their release in the stream. These results may have implications regarding the stocking success when releasing hatchery-reared Atlantic salmons in suboptimal environments. In particular, the study indicates that stocked salmon adaptation in the selected stream may not be improved by short conditioning periods to natural diet prior to their release, while we discuss potential explanations for the observed results.     

E-Cadherin-Dependent Stimulation of Traction Force at Focal Adhesions via the Src and PI3K Signaling Pathways

The interplay between cadherin- and integrin-dependent signals controls cell behavior, but the precise mechanisms that regulate the strength of adhesion to the extracellular matrix remains poorly understood. We deposited cells expressing a defined repertoire of cadherins and integrins on fibronectin (FN)-coated polyacrylamide gels (FN-PAG) and on FN-coated pillars used as a micro-force sensor array (μFSA), and analyzed the functional relationship between these adhesion receptors to determine how it regulates cell traction force. We found that cadherin-mediated adhesion stimulated cell spreading on FN-PAG, and this was modulated by the substrate stiffness. We compared S180 cells with cells stably expressing different cadherins on μFSA and found that traction forces were stronger in cells expressing cadherins than in parental cells. E-cadherin-mediated contact and mechanical coupling between cells are required for this increase in cell-FN traction force, which was not observed in isolated cells, and required Src and PI3K activities. Traction forces were stronger in cells expressing type I cadherins than in cells expressing type II cadherins, which correlates with our previous observation of a higher intercellular adhesion strength developed by type I compared with type II cadherins. Our results reveal one of the mechanisms whereby molecular cross talk between cadherins and integrins upregulates traction forces at cell-FN adhesion sites, and thus provide additional insight into the molecular control of cell behavior.     

Effectiveness of specific RNA-mediated interference through ingested double-stranded RNA in Caenorhabditis elegans

Background

In Caenorhabditis elegans, injection of double-stranded RNA (dsRNA) results in the specific inactivation of genes containing homologous sequences, a technique termed RNA-mediated interference (RNAi). It has previously been shown that RNAi can also be achieved by feeding worms Escherichia coli expressing dsRNA corresponding to a specific gene; this mode of dsRNA introduction is conventionally considered to be less efficient than direct injection, however, and has therefore seen limited use, even though it is considerably less labor-intensive.

Results

Here we present an optimized feeding method that results in phenotypes at least as strong as those produced by direct injection of dsRNA for embryonic lethal genes, and stronger for genes with post-embryonic phenotypes. In addition, the interference effect generated by feeding can be titrated to uncover a series of hypomorphic phenotypes informative about the functions of a given gene. Using this method, we screened 86 random genes on consecutive cosmids and identified functions for 13 new genes. These included two genes producing an uncoordinated phenotype (a previously uncharacterized POU homeodomain gene, ceh-6, and a gene encoding a MADS-box protein) and one gene encoding a novel protein that results in a high-incidence-of-males phenotype.

Conclusions

RNAi by feeding can provide significant information about the functions of an individual gene beyond that provided by injection. Moreover, it can be used for special applications for which injection or the use of mutants is sometimes impracticable (for example, titration, biochemistry and large-scale screening). Thus, RNAi by feeding should make possible new experimental approaches for the use of genomic sequence information.     

Cascading effects of predatory fish on the composition of benthic algae in high‐altitude streams

Cascading effects of predators can affect ecosystem properties by changing plant biomass, distribution and assemblage composition. Using data from field surveys and whole‐stream experiments we tested the hypothesis that predatory trout change assemblage composition of benthic algae in high‐elevation streams mediated by grazer behavior. Field surveys revealed that the taxonomic composition of algal assemblages differed significantly between streams that contained trout and those that were fishless; but comparisons of palatable versus unpalatable algal taxa between fish and fishless streams were equivocal because of high natural variability. Therefore, we tested for a behavioral (non‐consumptive) trophic cascade experimentally by adding brook trout chemical cues to six naturally fishless streams for 25 days and compared responses of grazers and algae to six reference streams without fish cues added. Algal response variables included rates of change in the abundance of three physiognomic categories, from most palatable (attached erect and prostrate diatoms) to least palatable (non‐diatoms), as determined from food selectivity analyses of the most common grazers (mayflies and caddisflies). Fish cues did not affect the mean densities or changes in densities of total grazers or any individual grazer species. However, in streams where fish cues were added, rates of accrual of attached erect diatoms, which was the preferred algal type for the grazer most vulnerable to trout predation (Baetis), were higher and their densities increased significantly faster with increasing densities of this grazer species than in reference streams. Results of his experiment support the hypothesis that predator induced suppression of grazer foraging behavior, rather than cascading effects of top predators on grazer density, may contribute to variation in the composition of algal assemblages among streams by allowing proliferation of most palatable algal species.     

The MEDiterranean Prediction And Classification System (MEDPACS): an implementation of the RIVPACS/AUSRIVAS predictive approach for assessing Mediterranean aquatic macroinvertebrate communities

In Spain, a national project known as GUADALMED, focusing on Mediterranean streams, has been carried out from 1998 to 2005 to implement the European water framework directive (WFD) requirements. One of the main objectives of the second phase of the project (2002–2005) was to develop a predictive system for the Spanish Mediterranean aquatic macroinvertebrate communities. A combined-season (spring, summer, and autumn) predictive model was developed by using the latest improvements on the selection of best predictor variables. Overall model performance measures were used to select the best discriminant function (DF) models, and also to evaluate their biases and precision. The final predictive model was based on the best five DF models. Each one of these models involved eight environmental variables. Final observed (O), expected (E), and O/E values for the number of macroinvertebrate families (NFAM) and two biotic indices (IBMWP and IASPT) were calculated by averaging their values, previously weighted by the quality of each DF model. Regression analyses among the final O and E values for the calibration dataset showed a high proximity to the ideal theoretical model, where the final E values explained 73–84% of the variation present in the macroinvertebrate communities of the Spanish Mediterranean watercourses. The ANOVA performed among the reference (calibration and validation) and test datasets showed clear differences for the O/E values. Finally, the assessments carried out by the predictive model were sensitive to anthropogenic pressure present in the study area and allowed the definition of five ecological status classes according to the WFD requirements. Handling editor: Richard H. Norris