Alternate routes to the cell surface underpin insulin-regulated membrane trafficking of GLUT4

Insulin-stimulated delivery of glucose transporters (GLUT4) from specialized intracellular GLUT4 storage vesicles (GSVs) to the surface of fat and muscle cells is central to whole-body glucose. This translocation and subsequent internalization of GLUT4 back into intracellular stores transits numerous small membrane-bound compartments (internal GLUT4-containing vesicles; IGVs) including GSVs, but the function of these different compartments is not clear. Cellugyrin and sortilin define distinct populations of IGV; sortilin-positive IGVs represent GSVs, but the function of cellugyrin-containing IGVs is unknown. Here we demonstrate a role for cellugyrin in intracellular sequestration of GLUT4 in HeLa cells and have used a proximity ligation assay to follow changes in pairwise associations between cellugyrin, sortilin, GLUT4 and membrane trafficking machinery following insulin-stimulation of 3T3-L1 adipoctyes. Our data suggest that insulin stimulates traffic from cellugyrin- to sortilin- membranes, and that cellugyrin-IGVs provide an insulin-sensitive reservoir to replenish GSVs following insulin-stimulated exocytosis of GLUT4. Furthermore, our data support the existence of a pathway from cellugyrin-membranes to the surface of 3T3-L1 adipocytes that bypasses GSVs under basal conditions, and that insulin diverts traffic away from this into GSVs

DEKAS - An evolutionary case-based reasoning system to support protection scheme design

This paper describes a decision support system being developed in conjunction with two UK utility companies to aid the design of electrical power transmission protection systems. A brief overview of the application domain is provided, followed by a description of the work carried out to date concerning the development and deployment of the Design Engineering Knowledge Application System (DEKAS). The paper then discusses the provision of intelligent decision support to the design engineer through the application of case-based reasoning (CBR). The key benefits from this will be outlined in conjunction with a relevant case study

Targeting long non-coding RNAs (lncRNAs) with oligonucleotides in cancer therapy

This document is the Accepted Manuscript version of a published work that appeared in final form in Translational Cancer Research. To access the final edited and published work see http://dx.doi.org/10.21037/tcr.2016.10.63No abstrac

Regulation of apoptosis by long non-coding RNA GAS5 in breast cancer cells: implications for chemotherapy.

The final publication is available at Springer via http://dx.doi.org/10.1007/s10549-014-2974-yThe putative tumour suppressor and apoptosis-promoting gene, growth arrest-specific 5 (GAS5), encodes long ncRNA (lncRNA) and snoRNAs. Its expression is down-regulated in breast cancer, which adversely impacts patient prognosis. In this preclinical study, the consequences of decreased GAS5 expression for breast cancer cell survival following treatment with chemotherapeutic agents are addressed. In addition, functional responses of triple-negative breast cancer cells to GAS5 lncRNA are examined, and mTOR inhibition as a strategy to enhance cellular GAS5 levels is investigated. Breast cancer cell lines were transfected with either siRNA to GAS5 or with a plasmid encoding GAS5 lncRNA and the effects on breast cancer cell survival were determined. Cellular responses to mTOR inhibitors were evaluated by assaying culture growth and GAS5 transcript levels. GAS5 silencing attenuated cell responses to apoptotic stimuli, including classical chemotherapeutic agents; the extent of cell death was directly proportional to cellular GAS5 levels. Imatinib action in contrast, was independent of GAS5. GAS5 lncRNA promoted the apoptosis of triple-negative and oestrogen receptor-positive cells but only dual PI3K/mTOR inhibition was able to enhance GAS5 levels in all cell types. Reduced GAS5 expression attenuates apoptosis induction by classical chemotherapeutic agents in breast cancer cells, providing an explanation for the relationship between GAS5 expression and breast cancer patient prognosis. Clinically, this relationship may be circumvented by the use of GAS5-independent drugs such as imatinib, or by restoration of GAS5 expression. The latter may be achieved by the use of a dual PI3K/mTOR inhibitor, to improve apoptotic responses to conventional chemotherapies.Breast Cancer Campaign U

The hormone response element mimic sequence of GAS5 lncRNA is sufficient to induce apoptosis in breast cancer cells.

Growth arrest-specific 5 (GAS5) lncRNA promotes apoptosis, and its expression is down-regulated in breast cancer. GAS5 lncRNA is a decoy of glucocorticoid/related receptors; a stem-loop sequence constitutes the GAS5 hormone response element mimic (HREM), which is essential for the regulation of breast cancer cell apoptosis. This preclinical study aimed to determine if the GAS5 HREM sequence alone promotes the apoptosis of breast cancer cells. Nucleofection of hormone-sensitive and -insensitive breast cancer cell lines with a GAS5 HREM DNA oligonucleotide increased both basal and ultraviolet-C-induced apoptosis, and decreased culture viability and clonogenic growth, similar to GAS5 lncRNA. The HREM oligonucleotide demonstrated similar sequence specificity to the native HREM for its functional activity and had no effect on endogenous GAS5 lncRNA levels. Certain chemically modified HREM oligonucleotides, notably DNA and RNA phosphorothioates, retained pro-apoptotic. activity. Crucially the HREM oligonucleotide could overcome apoptosis resistance secondary to deficient endogenous GAS5 lncRNA levels. Thus, the GAS5 lncRNA HREM sequence alone is sufficient to induce apoptosis in breast cancer cells, including triple-negative breast cancer cells. These findings further suggest that emerging knowledge of structure/function relationships in the field of lncRNA biology can be exploited for the development of entirely novel, oligonucleotide mimic-based, cancer therapies.Breast Cancer No

GAS5 lncRNA Modulates the Action of mTOR Inhibitors in Prostate Cancer Cells

Background There is a need to develop new therapies for castrate-resistant prostate cancer (CRPC) and growth arrest-specific 5 (GAS5) long non-coding RNA (lncRNA), which riborepresses androgen receptor action, may offer novel opportunities in this regard. GAS5 lncRNA expression declines as prostate cancer cells acquire castrate-resistance, and decreased GAS5 expression attenuates the responses of prostate cancer cells to apoptotic stimuli. Enhancing GAS5 lncRNA expression may therefore offer a strategy to improve the effectiveness of chemotherapeutic agents. GAS5 is a member of the 5' terminal oligopyrimidine gene family, and we have therefore examined if mTOR inhibition can enhance cellular GAS5 levels in prostate cancer cells. In addition, we have determined if GAS5 lncRNA itself is required for mTOR inhibitor action in prostate cancer cells, as recently demonstrated in lymphoid cells. Method The effects of mTOR inhibitors on GAS5 lncRNA expression and cell proliferation were determined in a range of prostate cancer cell lines. Transfection of cells with GAS5 siRNA and plasmid constructs was performed to determine the involvement of GAS5 lncRNA in mTOR inhibitor action. Results Treatment with rapamycin and rapalogues increased cellular GAS5 levels and inhibited culture growth in both androgen-dependent (LNCaP) and androgen-sensitive (22Rv1) cell lines, but not in androgen-independent (PC-3 and DU145) cells. GAS5 silencing in both LNCaP and 22Rv1 cells decreased their sensitivity to growth inhibition by mTOR inhibitors. Moreover, transfection of GAS5 lncRNA sensitized PC-3 and DU145 cells to mTOR inhibitors, resulting in inhibition of culture growth. Conclusion mTOR inhibition enhances GAS5 transcript levels in some, but not all, prostate cancer cell lines. This may in part be related to endogenous levels of GAS5 expression, which tend to be lower in prostate cancer cells representative of advanced disease, particularly since current findings demonstrate a role for GAS5 lncRNA in mTOR inhibitor action in prostate cancer cells

Galaxy-Galaxy Lensing in the Hubble Deep Field: The Halo Tully-Fisher Relation at Intermediate Redshift

A tangential distortion of background source galaxies around foreground lens galaxies in the Hubble Deep Field is detected at the 99.3% confidence level. An important element of our analysis is the use of photometric redshifts to determine distances of lens and source galaxies and rest-frame B-band luminosities of the lens galaxies. The lens galaxy halos obey a Tully-Fisher relation between halo circular velocity and luminosity; the typical lens galaxy, at a redshift z = 0.6, has a circular velocity of 210 +/-40 km/s at M_B = -18.5, if q_0 = 0.5. Control tests, in which lens and source positions and source ellipticities are randomized, confirm the significance level of the detection quoted above. Furthermore, a marginal signal is also detected from an independent, fainter sample of source galaxies without photometric redshifts. Potential systematic effects, such as contamination by aligned satellite galaxies, the distortion of source shapes by the light of the foreground galaxies, PSF anisotropies, and contributions from mass distributed on the scale of galaxy groups are shown to be negligible. A comparison of our result with the local Tully-Fisher relation indicates that intermediate-redshift galaxies are fainter than local spirals by 1.0 +/- 0.6 B mag at a fixed circular velocity. This is consistent with some spectroscopic studies of the rotation curves of intermediate-redshift galaxies. This result suggests that the strong increase in the global luminosity density with redshift is dominated by evolution in the galaxy number density.Comment: Revised version with minor changes. 13 pages, 7 figures, LaTeX2e, uses emulateapj and multicol styles (included). Accepted by Ap

A definitive merger-AGN connection at z~0 with CFIS: mergers have an excess of AGN and AGN hosts are more frequently disturbed

The question of whether galaxy mergers are linked to the triggering of active galactic nuclei (AGN) continues to be a topic of considerable debate. The issue can be broken down into two distinct questions: 1) Can galaxy mergers trigger AGN? 2) Are galaxy mergers the dominant AGN triggering mechanism? A complete picture of the AGN-merger connection requires that both of these questions are addressed with the same dataset. In previous work, we have shown that galaxy mergers selected from the Sloan Digital Sky Survey (SDSS) show an excess of both optically-selected, and mid-IR colour-selected AGN, demonstrating that the answer to the first of the above questions is affirmative. Here, we use the same optical and mid-IR AGN selection to address the second question, by quantifying the frequency of morphological disturbances in low surface brightness r-band images from the Canada France Imaging Survey (CFIS). Only ~30 per cent of optical AGN host galaxies are morphologically disturbed, indicating that recent interactions are not the dominant trigger. However, almost 60 per cent of mid-IR AGN hosts show signs of visual disturbance, indicating that interactions play a more significant role in nuclear feeding. Both mid-IR and optically selected AGN have interacting fractions that are a factor of two greater than a mass and redshift matched non-AGN control sample, an excess that increases with both AGN luminosity and host galaxy stellar mass.Comment: Accepted for publication in MNRA

The Next Generation Virgo Cluster Survey. XII. Stellar Populations and Kinematics of Compact, Low-Mass Early-Type Galaxies from Gemini GMOS-IFU Spectroscopy

We present Gemini GMOS-IFU data of eight compact low-mass early-type galaxies (ETGs) in the Virgo cluster. We analyse their stellar kinematics, stellar population, and present two-dimensional maps of these properties covering the central 5"x 7" region. We find a large variety of kinematics: from non- to highly-rotating objects, often associated with underlying disky isophotes revealed by deep images from the Next Generation Virgo Cluster Survey. In half of our objects, we find a centrally-concentrated younger and more metal-rich stellar population. We analyze the specific stellar angular momentum through the lambdaR parameter and find six fast-rotators and two slow-rotators, one having a thin counter-rotating disk. We compare the local galaxy density and stellar populations of our objects with those of 39 more extended low-mass Virgo ETGs from the SMAKCED survey and 260 massive ($M>10^{10}$\Msun) ETGs from the A3D sample. The compact low-mass ETGs in our sample are located in high density regions, often close to a massive galaxy and have, on average, older and more metal-rich stellar populations than less compact low-mass galaxies. We find that the stellar population parameters follow lines of constant velocity dispersion in the mass-size plane, smoothly extending the comparable trends found for massive ETGs. Our study supports a scenario where low-mass compact ETGs have experienced long-lived interactions with their environment, including ram-pressure stripping and gravitational tidal forces, that may be responsible for their compact nature.Comment: Accepted in ApJ, 19 pages, 10 figure

Regulation of the cell cycle and cell death by protein phosphatase 4 in breast cancer cell lines

Background At the molecular level, cell death is often regulated by the level of phosphorylation of particular proteins, i.e. by the balance of between opposing kinase and phosphatase activities on those proteins. Protein phosphatase 4 (PP4) is a PP2A-related serine/threonine phosphatase. PP2A has already been implicated in the control of cell proliferation, cell cycle and tumorigenesis. Using a functional expression cloning strategy, we have previously identified the catalytic subunit of PP4 (PP4c) as an important gene influencing the regulation of both apoptosis and cell proliferation in human leukaemic cell lines and in normal lymphocytes. The aims of this study were to examine the effects of PP4c overexpression and silencing on the cell death and survival of breast cancer cell lines. Method MCF7 and MDA-MB-231 cells were transfected with pcDNA3.1 encoding PP4c (pcDNA3-PP4c) or siRNAs to different PP4c sequences. Cells transfected with scrambled siRNA or empty vector were considered as controls. Culture viability, apoptosis and cell cycle were assessed post transfection. Results In MCF7 and metastatic MDA-MB-231 cells, PP4c over-expression exerted an inhibitory effect on cell proliferation, enhanced spontaneous apoptosis and decreased their colony forming ability. Conversely, siRNA mediated silencing of PP4 enhanced the proliferation and survival of MCF7 and MDA-MB-231 cells, affected cell cycle kinetics by enhancing the proportion of cells in S and G2/M phases, increased the colony forming ability and stimulated the anchorage independent growth. Conclusion PP4c promotes cell death and inhibits proliferation in breast cells, suggestive of a role of PP4c as tumour suppressor gene. Down regulation of PP4c expression increases cell survival, proliferation and anchorage independent growth of breast cancer cells, indicating a potential link between the PP4c expression levels, tumorigenesis and metastasis