Thrombin induces Egr-1 expression in fibroblasts involving elevation of the intracellular Ca2+ concentration, phosphorylation of ERK and activation of ternary complex factor

Background

The Arthropods are a diverse group of organisms including Chelicerata (ticks, mites, spiders), Crustacea (crabs, shrimps), and Insecta (flies, mosquitoes, beetles, silkworm). The cattle tick, Rhipicephalus (Boophilus) microplus, is an economically significant ectoparasite of cattle affecting cattle industries world wide. With the availability of sequence reads from the first Chelicerate genome project (the Ixodes scapularis tick) and extensive R. microplus ESTs, we investigated evidence for putative RNAi proteins and studied RNA interference in tick cell cultures and adult female ticks targeting Drosophila homologues with known cell viability phenotype.

Results

We screened 13,643 R. microplus ESTs and I. scapularis genome reads to identify RNAi related proteins in ticks. Our analysis identified 31 RNAi proteins including a putative tick Dicer, RISC associated (Ago-2 and FMRp), RNA dependent RNA polymerase (EGO-1) and 23 homologues implicated in dsRNA uptake and processing. We selected 10 R. microplus ESTs with >80% similarity to D. melanogaster proteins associated with cell viability for RNAi functional screens in both BME26 R. microplus embryonic cells and female ticks in vivo. Only genes associated with proteasomes had an effect on cell viability in vitro. In vivo RNAi showed that 9 genes had significant effects either causing lethality or impairing egg laying.

Conclusion

We have identified key RNAi-related proteins in ticks and along with our loss-of-function studies support a functional RNAi pathway in R. microplus. Our preliminary studies indicate that tick RNAi pathways may differ from that of other Arthropods such as insects.     

Conserved tertiary base pairing ensures proper RNA folding and efficient assembly of the signal recognition particle Alu domain

Proper folding of the RNA is an essential step in the assembly of functional ribonucleoprotein complexes. We examined the role of conserved base pairs formed between two distant loops in the Alu portion of the mammalian signal recognition particle RNA (SRP RNA) in SRP assembly and functions. Mutations disrupting base pairing interfere with folding of the Alu portion of the SRP RNA as monitored by probing the RNA structure and the binding of the protein SRP9/14. Complementary mutations rescue the defect establishing a role of the tertiary loop–loop interaction in RNA folding. The same mutations in the Alu domain have no major effect on binding of proteins to the S domain suggesting that the S domain can fold independently. Once assembled into a complete SRP, even particles that contain mutant RNA are active in arresting nascent chain elongation and translocation into microsomes, and, therefore, tertiary base pairing does not appear to be essential for these activities. Our results suggest a model in which the loop–loop interaction and binding of the protein SRP9/14 play an important role in the early steps of SRP RNA folding and assembly.     

1H, 15N and 13C chemical shift assignments for the winged helix domains of two archeal MCM C-termini

High-fidelity replication guarantees the stable inheritance of genetic information stored in the DNA of living organisms. The minichromosome maintenance (MCM) complex functions as replicative DNA-unwinding helicase and has been identified as one key player in the replication process of archea and eukarya. Despite the availability of considerable structural information on archeal MCMs, such information was missing for their C-terminal domain. In order to obtain more detailed structural information, we assigned the NMR chemical shifts for backbone and side chain nuclei for the MCM C-terminal winged helix domains of the archeal species Methanothermobacter thermautrophicus and Sulfolobus solfataricus.     

Chemically moderated gamete preferences predict offspring fitness in a broadcast spawning invertebrate

Sperm chemoattraction, where sperm locate unfertilized eggs by following a concentration gradient of egg-derived chemoattractants, has been widely documented across numerous taxa. While marine invertebrates are favoured models for understanding the underlying mechanisms of sperm chemoattraction, the evolutionary forces underpinning the process remain enigmatic. Here, we show that in mussels (Mytilus galloprovincialis), chemically moderated gamete preferences promote assortative fertilizations between genetically compatible gametes. When offered the choice of egg clutches from two females, sperm exhibited consistent but differential ‘preferences’ for chemical cues secreted from conspecific eggs. Critically, our data reveal that the preferences shown by sperm during the egg-choice trials are highly predictive of early embryonic viability when eggs and sperm from the same individuals are mixed during standard (no-choice) fertilization assays. Moreover, we demonstrate that by experimentally separating chemoattractants from eggs, sperm swimming behaviour is differentially regulated by egg-derived chemoattractants, and that these changes in sperm behaviour are highly consistent with observed patterns of gamete preferences, fertilization and larval survival. Together, this integrated series of experiments reveals that the behaviour of sperm is fine-tuned to respond differentially to the chemical signals emitted from different conspecific eggs, and that these choices have measurable fitness benefits.     

Intensity-based protein identification by machine learning from a library of tandem mass spectra

Tandem mass spectrometry (MS/MS) has emerged as a cornerstone of proteomics owing in part to robust spectral interpretation algorithms. Widely used algorithms do not fully exploit the intensity patterns present in mass spectra. Here, we demonstrate that intensity pattern modeling improves peptide and protein identification from MS/MS spectra. We modeled fragment ion intensities using a machine-learning approach that estimates the likelihood of observed intensities given peptide and fragment attributes. From 1,000,000 spectra, we chose 27,000 with high-quality, nonredundant matches as training data. Using the same 27,000 spectra, intensity was similarly modeled with mismatched peptides. We used these two probabilistic models to compute the relative likelihood of an observed spectrum given that a candidate peptide is matched or mismatched. We used a 'decoy' proteome approach to estimate incorrect match frequency, and demonstrated that an intensity-based method reduces peptide identification error by 50-96% without any loss in sensitivity.     

Multipotent capacity of immortalized human bronchial epithelial cells

While the adult murine lung utilizes multiple compartmentally restricted progenitor cells during homeostasis and repair, much less is known about the progenitor cells from the human lung. Translating the murine stem cell model to humans is hindered by anatomical differences between species. Here we show that human bronchial epithelial cells (HBECs) display characteristics of multipotent stem cells of the lung. These HBECs express markers indicative of several epithelial types of the adult lung when experimentally tested in cell culture. When cultured in three different three-dimensional (3D) systems, subtle changes in the microenvironment result in unique responses including the ability of HBECs to differentiate into multiple central and peripheral lung cell types. These new findings indicate that the adult human lung contains a multipotent progenitor cell whose differentiation potential is primarily dictated by the microenvironment. The HBEC system is not only important in understanding mechanisms for specific cell lineage differentiation, but also for examining changes that correlate with human lung diseases including lung cancer.     

Integrating field surveys and remote sensing data to study distribution, habitat use and conservation status of the herpetofauna of the Comoro Islands

We studied the non-marine reptile and amphibian species of the volcanic Comoro archipelago in the Western Indian Ocean, a poorly known island herpetofauna comprising numerous microendemic species of potentially high extinction risk and widespread, non-endemic and often invasive taxa. According to our data, the Comoro islands are inhabited by two amphibian species and at least 28 species of reptiles although ongoing genetic studies and unconfirmed historical records suggest an even higher species diversity. 14 of the 28 currently recognized species of terrestrial reptiles (50%) and the two amphibians are endemic to a single island or to the Comoro archipelago. The majority of species are most abundant at low elevation. However, a few endemic species, like the gekkonid lizards Paroedura sanctijohannis and Phelsuma nigristriata, are more common in or even confined to higher altitudes. We created habitat maps from remotely sensed data in combination with detailed species distribution maps produced using comprehensive data from field surveys between 2000 and 2010, literature, and historical locality records based on specimens in zoological collections. Using these data, we assessed the conservation status of the endemic terrestrial reptiles and amphibians according to the IUCN Red List criteria. Our results show that although little area of natural forest remains on the Comoros, many species are abundant in degraded forest or plantations. Competition and predation by invasive species appears to be the most important threat factor for the endemic herpetofauna, together with habitat degradation and destruction, which further favours invasive species. We propose the status Endangered for three species, Vulnerable for one species, Near Threatened for six species, Least Concern for four and Data Deficient for two species. The endemic subspecies Oplurus cuvieri comorensis is proposed for the status Critically Endangered. Based on the results of this study, seven areas of importance for reptile and amphibian conservation on the Comoros are identified. This study shows how remote sensing data can contribute to increasing accuracy and objectiveness of conservation assessments.     

Lack of relation between venous plasma total catecholamine concentrations and ventricular arrhythmias after acute myocardial infarction.

Electrocardiographic tracings were recorded continuously to monitor ventricular tachycardia and R-on-T and R-on-apex-T ventricular premature beats, and repeated estimations of venous plasma total catecholamine concentrations were carried out in 26 patients admitted to a coronary care unit with acute myocardial infarction. No relation existed between the increased catecholamine concentrations found in these patients and the incidence of ventricular arrhythmias occurring six to 48 hours after the onset of symptoms.     

Sites of synthesis of plasma proteins in the foetal rat

1. The foetal rat of 16 or more days incorporates ¹⁴C-labelled amino acids into all the demonstrable plasma protein fractions in vivo. 2. Slices of foetal rat liver incubated in vitro incorporate ¹⁴C-labelled amino acids into the main plasma protein fractions, including the foetal-specific `post-albumin'. 3. Slices of placenta are unable to incorporate ¹⁴C-labelled amino acids into plasma proteins in vitro. 4. Liver slices from maternal rats incubated in vitro incorporate ¹⁴C-labelled amino acids into plasma proteins. The presence of post-albumin cannot be demonstrated after incubation. 5. Liver slices from foetal rats, but not from adult rats, contain demonstrable amounts of haemoglobin into which ¹⁴C-labelled amino acids are incorporated.     

Fine scale movements and activity areas of white sharks (Carcharodon carcharias) in Mossel Bay, South Africa

Previous work on white sharks indicate the species show seasonally limited movement patters, at certain aggregation sites small areas may play vital roles in the life history of a large amount of the population. Acoustic telemetry was used to estimate habitat use of white sharks, Carcharodon carcharias, while aggregating at Mossel Bay, South Africa. Total range of all shark tracks combined accumulated 782 h and covered an area of 93.5 km² however, within this range, sharks were found to highly utilise a core habitat (50 % Kernel, K50) of just 1.05 km² over a reef system adjacent to a river mouth. Individual tracks revealed additional core habitats, some of which were previously undocumented and one adjacent to a commercial harbor. Much was found to be dependent on the size of the shark, with larger sharks (>400 cm) occupying smaller activity areas than subadult (300–399 cm) and juvenile (<300 cm) conspecifics, while Index of Reuse (IOR) and Index of Shared Space (IOSS) were both found to increase with shark size. Such results provide evidence that larger white sharks are more selective in habitat use, which indicates they have greater experience within aggregation sites. Furthermore, the focused nature of foraging means spatially restricted management strategies would offer a powerful tool to aid enforcement of current protective legislation for the white shark in similar environments of limited resources and capacity.