New Roles for the Integrated Stress Response in Cancer and Proteostasis

The integrated stress response (ISR) is a highly conserved pathway that senses diverse stresses and responds by limiting total protein synthesis and redirecting translation to stress response transcripts. The ISR has the potential to modify a broad range of processes in cancer, but the major cancer-relevant functions of the pathway have remained elusive due to the complex and redundant nature of the upstream kinases that sense stress and activate the pathway. To overcome this challenge, we genetically targeted the central, regulatory node of the pathway, eIF2α-serine 51, directly, in primary squamous cell carcinoma cells to generate ISR-null SCCs. We surprisingly found that the ISR acted as a tumor suppressor early in tumorigenesis, but also promoted proteostasis in response to the chemotherapy and proteasome inhibitor, bortezomib. We found this second finding to be mechanistically linked to a previously-undescribed role for the ISR in regulating the microtubule cytoskeleton to promote the removal of aggregated proteins. As a second goal of this project we sought to develop small molecule inhibitors of the ISR for anticancer therapy, focusing on the alternative translation initiation factor, EIF2A, which we previously found to be critical for oncogenesis. Utilizing dual luciferase reporters for alternative translation (Atf4-firefly) and housekeeping translation (HBB-renilla) we performed a high throughput screen and have identified promising hits with potential as novel EIF2A inhibitors

Oncolytic HSV-mediated Regulation of the Host Hypoxia Response

Glioblastoma (GBM) is the most common and deadly primary brain tumor, accounting for over 10,000 new cancer diagnoses in the United States each year. The poor prognosis for GBM patients necessitates novel biological treatments. One such approach is the use of oncolytic herpes simplex virus 1 (oHSV). Like many novel treatments oHSV therapy causes off target effects that are not yet well understood. Our lab has demonstrated that oHSV treatment increases the vascularity of brain tumors. The goal of this study is to determine the mechanism by which oHSV treatment increases the vascularity of brain tumors. Preliminary data suggested the hypoxia inducible factor-1 alpha (HIF1α) may be activated in cells infected with oHSV, even in normal oxygen conditions. HIF1α is a transcription factor, which activates a variety of genes in response to a lack of oxygen. We hypothesized that HIF1α activation may be responsible for the increased vascularity of oHSV treated brain tumors. A screen of targetscan.org for the predicted target genes of herpes simplex virus 1 (HSV-1) revealed multiple miRNAs predicted to target a protein called, factor inhibiting HIF1α (FIH). This protein functionally inhibits HIF1α activation by preventing the binding of HIF1α to DNA. We believed that FIH would be negatively regulated in GBM cells infected with oHSV, thus allowing HIF1α activation. In this study, we demonstrate that HSV-1 expresses two miRNA molecules, which target and down regulate FIH. Transfection of miRNA inhibitors (antagomirs) was able to successfully abrogate the virus' ability to down regulate FIH as demonstrated by quantitative PCR and western blot. Moreover, transfection of HSV-1 miRNA mimics in the absence of virus was able to down regulate FIH protein levels (western blot) and activate the expression of a variety of HIF1α driven genes, including VEGF and IL-8 (quantitative PCR). However, after analyzing HIF1α promoter activity in a variety of cell lines, we determined that HIF1α was not activated during oHSV infection. The expression of VEGF and IL-8 was likely due to the activation of another transcription factor.Pelotonia Undergraduate Student FellowshipA five-year embargo was granted for this item.Academic Major: Biomedical Scienc

Oncolytic HSV-mediated Regulation of the Host Hypoxia Response

Biological Sciences: 2nd Place (The Ohio State University Denman Undergraduate Research Forum)Glioblastoma (GBM) is the most common and deadly primary brain tumor, accounting for over 10,000 new cancer diagnoses in the United States each year. The poor prognosis for GBM patients necessitates novel biological treatments. One such approach is the use of oncolytic herpes simplex virus 1 (oHSV). Like many novel treatments, oHSV therapy causes side effects that are not yet well understood. Our lab has demonstrated that oHSV treatment increases the vascularity of brain tumors. The goal of this study is to determine the mechanism by which oHSV treatment increases the vascularity of brain tumors. We have determined that the hypoxia inducible factor-1 alpha (HIF1α) is activated in cells infected with oHSV, even in normal oxygen conditions. HIF1α is a transcription factor which activates a variety of genes in response to a lack of oxygen. We believe that HIF1α activation may be responsible for the increased vascularity of oHSV treated brain tumors. A screen of targetscan.org for herpes simplex virus 1 (HSV-1) miRNAs and their predicted target genes revealed multiple miRNAs predicted to target a protein called, factor inhibiting HIF1α (FIH). This protein functionally inhibits HIF1α activation by preventing the binding of HIF1α to DNA. We hypothesized that FIH would be negatively regulated in GBM cells infected with oHSV, thus allowing HIF1α activation. In this study, we demonstrate that HSV-1 expresses two miRNA molecules, which target and down regulate FIH. Transfection of miRNA inhibitors (antagomirs) was able to successfully abrogate the virus' ability to downregulate FIH as demonstrated by quantitative PCR and western blot. Moreover, transfection of HSV-1 miRNA mimics in the absence of virus was able to downregulate FIH protein levels (western blot) and activate the expression of a variety of HIF1α driven genes, including VEGF and CCN1 (quantitative PCR). Our future aim is to determine if HSV-1 encodes for miRNA capable of binding to the 3' untranslated region (3' UTR) of FIH. For this study we will employ an FIH-3' UTR luciferase reporter vector. This experiment will demonstrate if the miRNA expressed by oHSV directly binds to the 3' UTR of FIH, thus inhibiting FIH gene expression, and activating HIF1α.Pelotonia Undergraduate FellowshipAcademic Major: Biomedical Scienc

Ethnic variations in incidence of asthma episodes in England & Wales:national study of 502,482 patients in primary care

BACKGROUND: Recent studies have demonstrated marked international variations in the prevalence of asthma, but less is known about ethnic variations in asthma epidemiology within individual countries and in particular the impact of migration on risk of developing asthma. Recent within country comparisons have however revealed that despite originating from areas of the world with a low risk for developing asthma, South Asian and Afro-Caribbean people in the UK are significantly (3× and 2× respectively) more likely to be admitted to hospital for asthma related problems than Whites. METHODS: Using data from the Fourth National Study of Morbidity Statistics in General Practice, a one-percent broadly representative prospective cohort study of consultations in general practice, we investigated ethnic variations in incident asthma consultations (defined as new or first consultations), and compared consultation rates between those born inside and outside the UK (migrant status). Logistic regression models were used to examine the combined effects of ethnicity and migration on asthma incident consultations. RESULTS: Results showed significantly lower new/first asthma consultation rates for Whites than for each of the ethnic minority groups studied (mean age-adjusted consultation rates per 1000 patient-years: Whites 26.4 (95%CI 26.4, 26.4); South Asians 30.4 (95%CI 30.3, 30.5); Afro-Caribbeans 35.1 (95%CI 34.9, 35.3); and Others 27.8 (27.7, 28.0). Within each of these ethnic groups, those born outside of the UK showed consistently lower rates of incident asthma consultations. Modelling the combined effects of ethnic and migrant status revealed that UK-born South Asians and Afro-Caribbeans experienced comparable risks for incident GP consultations for asthma to UK-born Whites. Non-UK born Whites however experienced reduced risks (adjusted OR 0.82, 95%CI 0.69, 0.97) whilst non-UK born South Asians experienced increased risks (adjusted OR 1.33, 95%CI 1.04, 1.70) compared to UK-born Whites. CONCLUSION: These findings strongly suggest that ethnicity and migration have significant and independent effects on asthma incidence. The known poorer asthma outcomes in UK South Asians and Afro-Caribbeans may in part be explained by the offspring of migrants experiencing an increased risk of developing asthma when compared to UK-born Whites. This is the first study to find heterogeneity for incident asthma consultations in Whites by migrant status

Book Reviews

Book Review 1Book Title: Early Life Histories of Fishes: New Developmental, Ecological and Evolutionary PerspectivesBook Author: Edited by E.K. BalonDr W. Junk, Dordrecht. 280 pp.Book Review 2Book Title: Comparative Aspects of Extracellular Acid-base BalanceBook Author: J.P. TruchotSpringer, 1987. 248 pp. 51 figures.Book Review 3Book Title: Insect Flight: Dispersal and MigrationBook Author: Edited by W. DanthanarayanaSpringer-Verlag, Berlin, 1986. 289 pp.Book Review 4Book Title:  The Mammalian Herbivore Stomach. Comparative Anatomy, Function and EvolutionBook Author: Peter LangerGustav Fischer, Stuttgart, 1988. 557 pages, 246 figures and 72 tablesBook Review 5Book Title:  Biology of the Integument. Vol. 2: VertebratesBook Authors: Edited by J. Bereiter-Hahn, A.G. Matoltsy & K.S. RichardsSpringer, Berlin, 1986. 855 pp.Book Review 6Book Title:  Advances in the Biology of Turbellarians and related PlatyhelminthesBook Author: Edited by Seth TylerDr. W. Junk Publishers, 1986. 357 pages; 253 figuresBook Review 7Book Title: Evolutionary Physiological EcologyBook Author: Edited by P. CalowCambridge University Press, Cambridge. 239 pp.Book Review 8Book Title: DragonfliesBook Author: Peter L. MillerCambridge University Press, Cambridge, New York and Melbourne, 1987. 84 pp

Geologic Map of the Ganiki Planitia Quadrangle (V–14), Venus

Our current research focuses on addressing four specific questions. Has the dominant style of volcanic expression within the quadrangle varied in a systematic fashion over time? Does the tectonic deformation within the quadrangle record significant regional patterns that vary spatially or temporally, and if so what are the scales, orientations and sources of the stress fields driving this deformation? If mantle upwelling and downwelling have played a significant role in the formation of Atla Regio and Atalanta Planitia as has been proposed, does the geology of Ganiki Planitia record evidence of northwest-directed lateral mantle flow connecting the two sites? Finally, can integration of the tectonic and volcanic histories preserved within the quadrangle help constrain competing resurfacing models for Venus

The integrated stress response remodels the microtubule-organizing center to clear unfolded proteins following proteotoxic stress

Cells encountering stressful situations activate the integrated stress response (ISR) pathway to limit protein synthesis and redirect translation to better cope. The ISR has also been implicated in cancers, but redundancies in the stress-sensing kinases that trigger the ISR have posed hurdles to dissecting physiological relevance. To overcome this challenge, we targeted the regulatory node of these kinases, namely, the S51 phosphorylation site of eukaryotic translation initiation factor eIF2α and genetically replaced eIF2α with eIF2α-S51A in mouse squamous cell carcinoma (SCC) stem cells of skin. While inconsequential under normal growth conditions, the vulnerability of this ISR-null state was unveiled when SCC stem cells experienced proteotoxic stress. Seeking mechanistic insights into the protective roles of the ISR, we combined ribosome profiling and functional approaches to identify and probe the functional importance of translational differences between ISR-competent and ISR-null SCC stem cells when exposed to proteotoxic stress. In doing so, we learned that the ISR redirects translation to centrosomal proteins that orchestrate the microtubule dynamics needed to efficiently concentrate unfolded proteins at the microtubule-organizing center so that they can be cleared by the perinuclear degradation machinery. Thus, rather than merely maintaining survival during proteotoxic stress, the ISR also functions in promoting cellular recovery once the stress has subsided. Remarkably, this molecular program is unique to transformed skin stem cells, hence exposing a vulnerability in cancer that could be exploited therapeutically

Elemental Abundances and Ionization States within the Local Interstellar Cloud Derived from HST and FUSE Observations of the Capella Line of Sight

We use ultraviolet spectra of Capella from the Hubble Space Telescope (HST) and Far Ultraviolet Spectroscopic Explorer (FUSE) satellites to study interstellar absorption lines from the Local Interstellar Cloud (LIC). Measurements of these lines are used to empirically determine the ionization states of carbon, nitrogen, and silicon in the LIC, for comparison with the predictions of theoretical photoionization models. We find that the observed ionization states are consistent with previously published photoionization predictions. Total abundances are determined for the elements mentioned above, and others, for comparison with solar abundances. Magnesium, aluminum, silicon, and iron are all depleted by at least a factor of 10 toward Capella. The abundances of carbon, nitrogen, and oxygen are essentially solar, although the error bars are large enough to also allow depletions of about a factor of 2 for these elements.Comment: 31 pages, 10 figures; AASTEX v5.0 plus EPSF extensions in mkfig.sty; accepted by Ap