Association of p60c-src with endosomal membranes in mammalian fibroblasts.

We have examined the subcellular localization of p60c-src in mammalian fibroblasts. Analysis of indirect immunofluorescence by three-dimensional optical sectioning microscopy revealed a granular cytoplasmic staining that co-localized with the microtubule organizing center. Immunofluorescence experiments with antibodies against a number of membrane markers demonstrated a striking co-localization between p60c-src and the cation-dependent mannose-6-phosphate receptor (CI-MPR), a marker that identifies endosomes. Both p60c-src and the CI-MPR were found to cluster at the spindle poles throughout mitosis. In addition, treatment of interphase and mitotic cells with brefeldin A resulted in a clustering of p60c-src and CI-MPR at a peri-centriolar position. Biochemical fractionation of cellular membranes showed that a major proportion of p60c-src co-enriched with endocytic membranes. Treatment of membranes containing HRP to alter their apparent density also altered the density of p60c-src-containing membranes. Similar density shift experiments with total cellular membranes revealed that the majority of membrane-associated p60c-src in the cell is associated with endosomes, while very little is associated with plasma membranes. These results support a role for p60c-src in the regulation of endosomal membranes and protein trafficking

Progress toward Public Access to Science

PLoS Chairman of the Board Harold Varmus applauds the newly enacted NIH public access policy as a positive step toward ensuring greater access to and better use of the scientific literature

The level of utilization of innovative activities of transport businesses in the Slovak Republic

The purpose of this article is to identify the primary difficulties of implementing innovative activities in Slovak transport businesses and to propose some recommendations for minimizing the occurrence of these problems, based on the analysis of literary sources and carried out research. This article summarizes results of the authors, who conducted research on a sample of 53 respondents in order to show the current situation in the Slovak transport business that use innovative activities. This article also contains recommendations for successful implementation of innovative activities in business. These recommendations should be a valuable tool for business managers in their attempts to implement innovative activities

Wnt/Wingless signaling through β-catenin requires the function of both LRP/Arrow and frizzled classes of receptors

BACKGROUND: Wnt/Wingless (Wg) signals are transduced by seven-transmembrane Frizzleds (Fzs) and the single-transmembrane LDL-receptor-related proteins 5 or 6 (LRP5/6) or Arrow. The aminotermini of LRP and Fz were reported to associate only in the presence of Wnt, implying that Wnt ligands form a trimeric complex with two different receptors. However, it was recently reported that LRPs activate the Wnt/β-catenin pathway by binding to Axin in a Dishevelled – independent manner, while Fzs transduce Wnt signals through Dishevelled to stabilize β-catenin. Thus, it is possible that Wnt proteins form separate complexes with Fzs and LRPs, transducing Wnt signals separately, but converging downstream in the Wnt/β-catenin pathway. The question then arises whether both receptors are absolutely required to transduce Wnt signals. RESULTS: We have established a sensitive luciferase reporter assay in Drosophila S2 cells to determine the level of Wg – stimulated signaling. We demonstrate here that Wg can synergize with DFz2 and function cooperatively with LRP to activate the β-catenin/Armadillo signaling pathway. Double-strand RNA interference that disrupts the synthesis of either receptor type dramatically impairs Wg signaling activity. Importantly, the pronounced synergistic effect of adding Wg and DFz2 is dependent on Arrow and Dishevelled. The synergy requires the cysteine-rich extracellular domain of DFz2, but not its carboxyterminus. Finally, mammalian LRP6 and its activated forms, which lack most of the extracellular domain of the protein, can activate the Wg signaling pathway and cooperate with Wg and DFz2 in S2 cells. We also show that the aminoterminus of LRP/Arr is required for the synergy between Wg and DFz2. CONCLUSION: Our study indicates that Wg signal transduction in S2 cells depends on the function of both LRPs and DFz2, and the results are consistent with the proposal that Wnt/Wg signals through the aminoterminal domains of its dual receptors, activating target genes through Dishevelled

The Classic: Integration of Deoxyribonucleic Acid Specific for Rous Sarcoma Virus after Infection of Permissive and Nonpermissive Hosts: (RNA tumor viruses/reassociation kinetics/duck cells)

A relatively simple but stringent technique was developed to detect the integration of virus-specific DNA into the genomes of higher organisms. In both permissive (duck) and nonpermissive (mammalian) cells which normally contain no nucleotide sequences specific for Rous sarcoma virus, transformation by the virus results in the appearance of DNA specific for Rous sarcoma virus covalently integrated into strands of host-cell DNA containing reiterated sequences. Early after infection of mouse or duck cells by Rous sarcoma virus, unintegrated DNA specific for the virus can be demonstrated

The terminal redundancy of the retrovirus genome facilitates chain elongation by reverse transcriptase

Transcription of DNA from the RNA genome of retroviruses by reverse transcriptase involves an unusual translocation of the growing chain from the 5' end to the 3' end of the RNA template. In order to elucidate the mechanism by which this translocation occurs, we have used chain termination to analyze nascent viral DNA synthesized in vitro by avian sarcoma virus, and we have determined the nucleotide sequence of appropriate regions of viral DNA isolated from infected cells and cloned into prokaryotic vectors. Our results provide direct experimental evidence for a previously proposed model in which a short terminal redundancy in viral RNA, and a DNA copy of the redundant sequence, are used to allow the growing DNA chain to move from the 5' to the 3' end of the template. Transcription of avian sarcoma virus RNA with purified reverse transcriptase also generates an anomalous product, a hairpin DNA that arises when the initial DNA transcript folds back on itself to continue synthesis. The foldback is mediated by an inverted repeat of 5 nucleotides in the sequence of nascent DNA. Anomalous hairpin DNA is not produced by detergent-activated virions. Thus, constituents of the virons or the configuration of encapsidated viral RNA must faciliatate correct transcription

Nanotechnology and the Developing World

How nanotechnology can be harnessed to address some of the world's most critical development problem

Air conditioning in emergency medical services

Bakalářská práce se zabývá návrhem vzduchotechnického systému pro zdravotnickou záchrannou službu. V objektu se řeší především odvětrávání garáží pro stav provozní, havarijní a požární, větrání zázemí zaměstnanců a zasedací místnosti.Bachelor thesis deals with HVAC system design for emergency medical services. This project was designed for garage ventilation, emergency and fire emergency ventilation in garage, then ventilation background of staff and meeting room.