Sacred and Mortuary Landscapes in Iron Age Cyprus: A GIS Analysis

During the Archaic period (750-480 BC) the island of Cyprus underwent a dramatic transformation as new city-kingdoms rose to dominate the political landscape of the island. This shift resulted in increased competition for resources, establishment of political boundaries, and emergence of a pronounced social hierarchy within the new polities. While many of the large settlements that became centers of power during this time have been thoroughly studied, the manifestation of the large scale changes of the Archaic in the periphery have not been as fully investigated. The rural site of Athienou-Malloura, surveyed and excavated by the Athienou Archaeological Project includes a Cypro-Archaic sanctuary and nearby tombs on the hill of Maghara-Tepesi, four of which have been excavated. The present study compares the site of Athienou-Malloura to other comparable sites from around the island, in order to ascertain the distribution and role of rural sanctuaries and cemeteries during this period of increasing social complexity and political competition. The sites are compared to locations of the city-kingdoms, as well as access to natural resources such as arable soil and copper ore. It has been proposed both on Cyprus and elsewhere that grave monuments and religious sites partly functioned to create and enforce claims during turbulent growth periods. Building from more abstract speculations on Archaic Cypriot political boundaries, this study attempts to map a more nuanced view of the interplay between topography and human use of the landscape during this time

An investigation of genomic instability and its impact on cancer development and heterogeneity

Genomic instability (GIN), a genomic state facilitating large scale chromosomal rearrangements, is a hallmark of cancer. GIN can contribute to oncogenesis by disrupting genes, and leading to copy number aberrations (CNAs), the gain or loss of genomic segments. In this thesis I describe two projects linked by the overarching theme of GIN, outlined below: Project 1: Copy-number aberrations (CNAs) contribute to clonal diversity within cancer, with clinical implications. Breast cancer is one such example, but the effect of CNAs on gene expression in intra-tumour subclonal populations has not been properly characterised. Due to sequencing technology limits and lack of computational methods, it is difficult to assess CNAs at a subclonal level. Here, I have benchmarked the ‘InferCNV’ computational method and used it to infer single cell CNA profiles from 14 primary breast cancer single cell RNA-sequencing (scRNA-seq) datasets. I reveal diverse intratumoural heterogeneity involving at least four subclonal populations per tumour. Finally, I identify subclones with expression/CNA profiles indicative of metastatic potential, involving differential regulation of metastasis associated genes such as MUCL1, BST2 and IGFBP5. Project 2: High-grade serous ovarian cancer (HGSOC) is characterised by widespread GIN. Drivers of GIN include deficient DNA repair and amplification of Cyclin E1, however no major cause is known for one third of tumours. Deregulation of repetitive elements may contribute to GIN in HGSOC. It is difficult to investigate repetitive elements from sequencing data as they map to multiple places within the genome. I have quantified repetitive RNA in 99 high-grade serous ovarian cancer (HGSOC) and matched control RNA-seq datasets to determine their potential contribution to GIN. I identified retrotransposons which are deregulated in HGSOC, which may have been active during cancer development. Some of these retrotransposons were enriched at structural variant breakpoints, indicating potential causality. Finally, I identified retrotransposon-associated structural variants in proximity to deregulated oncogenes implicated in homologous DNA repair, which may have modulated their expression and contributed to cancer development. In summary, I have explored both a cause (retrotransposons) and consequence (CNA-based heterogeneity) of GIN in cancer, and shown how GIN can contribute to the modulation of cancer-associated genes which influence cancer development and outcomes

The botanical biofiltration of VOCs with active airflow: is removal efficiency related to chemical properties?

© 2019 Elsevier Ltd Botanical biofiltration using active green walls is showing increasing promise as a viable method for the filtration of volatile organic compounds (VOCs) from ambient air; however there is a high level of heterogeneity reported amongst VOC removal efficiencies, and the reasons for these observations have yet to be explained. Comparisons of removal efficiencies amongst studies is also difficult due to the use of many different VOCs, and systems that have been tested under different conditions. The current work describes a procedure to determine whether some of these differences may be related to the chemical properties of the VOCs themselves. This work used an active green wall system to test the single pass removal efficiency (SPRE) of nine different VOCs (acetone, benzene, cyclohexane, ethanol, ethyl acetate, hexane, isopentane, isopropanol and toluene) and explored which chemical properties were meaningful predictor variables of their biofiltration efficiencies. Ethanol was removed most efficiently (average SPRE of 96.34% ± 1.61), while benzene was least efficiently removed (average SPRE of 19.76% ± 2.93). Multiple stepwise linear regression was used to determine that the dipole moment and molecular mass were significant predictors of VOC SPRE, in combination accounting for 54.6% of the variability in SPREs amongst VOCs. The octanol water partition coefficient, proton affinity, Henry's law constant and vapour pressure were not significant predictors of SPRE. The most influential predictor variable was the dipole moment, alone accounting for 49.8% of the SPRE variability. The model thus allows for an estimation of VOC removal efficiency based on a VOC's chemical properties, and supports the idea that system optimisation could be achieved through methods that promote both VOC partitioning into the biofilter's aqueous phase, and substrate development to enhance adsorption.

PAM variants in patients with thyrotrophinomas, cyclical Cushing’s disease and prolactinomas

IntroductionGermline loss-of-function variants in PAM, encoding peptidylglycine α-amidating monooxygenase (PAM), were recently discovered to be enriched in conditions of pathological pituitary hypersecretion, specifically: somatotrophinoma, corticotrophinoma, and prolactinoma. PAM is the sole enzyme responsible for C-terminal amidation of peptides, and plays a role in the biosynthesis and regulation of multiple hormones, including proopiomelanocortin (POMC).MethodsWe performed exome sequencing of germline and tumour DNA from 29 individuals with functioning pituitary adenomas (12 prolactinomas, 10 thyrotrophinomas, 7 cyclical Cushing’s disease). An unfiltered analysis was undertaken of all PAM variants with population prevalence <5%.ResultsWe identified five coding, non-synonymous PAM variants of interest amongst seven individuals (six germline, one somatic). The five variants comprised four missense variants and one truncating variant, all heterozygous. Each variant had some evidence of pathogenicity based on population prevalence, conservation scores, in silico predictions and/or prior functional studies. The yield of predicted deleterious PAM variants was thus 7/29 (24%). The variants predominated in individuals with thyrotrophinomas (4/10, 40%) and cyclical Cushing’s disease (2/7, 29%), compared to prolactinomas (1/12, 8%).ConclusionThis is the second study to demonstrate a high yield of suspected loss-of-function, predominantly germline, PAM variants in individuals with pathological pituitary hypersecretion. We have extended the association with corticotrophinoma to include the specific clinical entity of cyclical Cushing’s disease and demonstrated a novel association between PAM variants and thyrotrophinoma. PAM variants might act as risk alleles for pituitary adenoma formation, with a possible genotype-phenotype relationship between truncating variants and altered temporal secretion of cortisol

Antigen-driven EGR2 expression is required for exhausted CD8 + T cell stability and maintenance

Chronic stimulation of CD8 T cells triggers exhaustion, a distinct differentiation state with diminished effector function. Exhausted cells exist in multiple differentiation states, from stem-like progenitors that are the key mediators of the response to checkpoint blockade, through to terminally exhausted cells. Due to its clinical relevance, there is substantial interest in defining the pathways that control differentiation and maintenance of these subsets. Here, we show that chronic antigen induces the anergy-associated transcription factor EGR2 selectively within progenitor exhausted cells in both chronic LCMV and tumours. EGR2 enables terminal exhaustion and stabilizes the exhausted transcriptional state by both direct EGR2-dependent control of key exhaustion-associated genes, and indirect maintenance of the exhausted epigenetic state. We show that EGR2 is a regulator of exhaustion that epigenetically and transcriptionally maintains the differentiation competency of progenitor exhausted cells. +This work was funded by National Institutes of Health Grant U19-AI100627, the Swiss National Science Foundation and the Novartis Foundation for Medical-Biological Research (S.S.G.), the Australian Cancer Research Foundation (for the Peter Mac Flow Cytometry and Molecular Genomics facilities) and by the National Health and Medical Research Council (NHMRC) through Program Grants 1016953 & 1113904, Ideas Grant APP2001719, Australia Fellowship 585490 (C.C.G.), Senior Principal Research Fellowships (1081858, C.C.G., 1139607, A.K.), and CJ Martin Early Career Fellowship 585518 (I.A.P.)

Regulation of human CD4+ T cell differentiation

Naive CD4+ T cells differentiate into specific effector subsets—Th1, Th2, Th17, and T follicular helper (Tfh)—that provide immunity against pathogen infection. The signaling pathways involved in generating these effector cells are partially known. However, the effects of mutations underlying human primary immunodeficiencies on these processes, and how they compromise specific immune responses, remain unresolved. By studying individuals with mutations in key signaling pathways, we identified nonredundant pathways regulating human CD4+ T cell differentiation in vitro. IL12Rβ1/TYK2 and IFN-γR/STAT1 function in a feed-forward loop to induce Th1 cells, whereas IL-21/IL-21R/STAT3 signaling is required for Th17, Tfh, and IL-10–secreting cells. IL12Rβ1/TYK2 and NEMO are also required for Th17 induction. Strikingly, gain-of-function STAT1 mutations recapitulated the impact of dominant-negative STAT3 mutations on Tfh and Th17 cells, revealing a putative inhibitory effect of hypermorphic STAT1 over STAT3. These findings provide mechanistic insight into the requirements for human T cell effector function, and explain clinical manifestations of these immunodeficient conditions. Furthermore, they identify molecules that could be targeted to modulate CD4+ T cell effector function in the settings of infection, vaccination, or immune dysregulation

An investigation of the cellular mechanisms regulated by CXCR7 during hepatic and pulmonary fibrosis.

Fibrosis of the lungs and liver, characterised by excessive extra-cellular matrix (ECM) deposition and scarring following injury, has a significant negative impact on health worldwide. The incidence of these conditions is increasing, however their development and cause are poorly understood. Immune signaling molecules known as chemokines and their receptors have been implicated in fibrogenesis. Our laboratory previously found the chemokine receptor CXCR7 to be implicated in lung and liver fibrosis, as blocking CXCR7 with specific antibodies or knocking CXCR7 out genetically reduced total collagen deposition and expression of other markers of fibrosis significantly in 2 models of lung fibrosis and 1 model of liver fibrosis. In the lung CXCR7 was detected in bronchial epithelial cells.Aims: In this study we sought to validate CXCR7 involvement in a second model of hepatic fibrosis and to define the cell types expressing CXCR7 in normal and fibrotic livers. Furthermore, we investigated the mechanisms regulated by CXCR7 in vitro.Results: During liver fibrosis Cxcr7 mRNA was upregulated in all models in parallel to several common markers of fibrosis. CXCR7 mRNA was also found to be upregulated in human cirrhotic liver. In a CCl4-induced liver fibrosis model, Cxcr7 KO prevented upregulation of several of these fibrosis markers.In the liver Cxcr7 was found to be expressed in the membrane of hepatocytes with a strong signal on the bile canaliculi. Cxcr7 could not be detected on endothelial cells or hepatic stellate cells, the fibroblasts of the liver.Treatment of transformed human bronchial epithelial Beas-2B cells in vitro with profibrogenic factor TGF-β resulted in upregulation of the mRNA of Cxcr7 and markers of fibrosis as expected, however blocking Cxcr7 or stimulating Cxcr7 with ligand did not have any effect.From this it can be concluded that CXCR7 is expressed in cells of epithelial origin in the liver (hepatocytes) and is upregulated during liver fibrosis. It was shown to have profibrotic effect in an additional model (CCl4-induced) of liver fibrosis. CXCL7 levels paralleled fibrotic marker levels both in vivo and in vitro. This study shows CXCR7 is a critical component of profibrogenic signaling and future work should focus on these cell types that express it to explore mechanisms of fibrotic activity by CXCR7. This will lead to a better understanding of lung and liver fibrogenesis and perhaps a potential therapeutic to block the receptor in human patients