A critical analysis of local and global cultural factors in graphic wayfinding design: a case study of Beijing

The main intentions of this thesis are to analyse and explain changes in the function and graphic components of Beijing s wayfinding systems and to explain how the systems construct multiple cultural and political identities at different historical periods and in changing local/global contexts. In the thesis, the oversimplified one-way theory of the global-local dichotomy, in which the global power of the West is overwhelming and constantly dominant, and the local system of non-Western countries is passive and fragile, is challenged. Instead, this thesis seeks to examine the interactivity and correlation of the local and the global from two perspectives: mobility and reversibility. Looking at mobility is to consider the local and global and their nexus as different interconnections and networks that are constantly and unevenly changing. Reversibility, with which this thesis is most concerned, deals primarily with the reversible relationship of the local and global, namely, that either the local or the global can be dominant. This point is well illustrated by the evolution of Beijing s graphic wayfinding systems function and appearance. Beijing, as the capital of China, has undergone a radical transformation from the fall of the last Empire Qing (1912) to the establishment of the People s Republic of China (1949). The meaning of Beijing varies in accordance with the changes in its political and social structures. There have been five phases in Beijing s development: a well-planned imperial city; a capital city with a republican spirit; a totally industrialised but relatively isolated capital of a socialist country; an open and modernised Chinese-style socialist city; and a cosmopolitan city. In the course of this metamorphosis, what took place was a series of collisions, exchanges, fusions, and re-collisions between local power and global power. Along with the immense changes in Beijing, the role and appearance of the graphic wayfinding systems have also changed, especially those of road signs and doorplates, whose roles have been transformed from that of initial household register to orientation reference, to effective propaganda tool, and then on to the regeneration of a city. Finally, Beijing s graphic wayfinding design within its urban development has been reconfirmed as a useful instrument to support the new forms of visual narratives and consolidate the city brand of Beijing in the 21st century. This study probes into the political and cultural significances behind the changes of the graphic wayfinding systems of Beijing, as well as the interaction between the local and the global as reflected in the formation of these findings. The mutable and reversible relationship between the local and the global is illustrated and clarified through analysis and comparison of various functions and visual elements between Beijing s present graphic wayfinding systems and its early wayfinding signs, as well as decoding the different mainstream political or cultural ideologies that have deeply affected the function and design of Beijing graphic wayfinding systems at different periods

Allele-specific siRNA Therapy for Keratitis-Ichthyosis-Deafness Syndrome

Dominant mutations in the gene GJB2 cause keratitis-ichthyosis-deafness (KID) syndrome, a severe condition affecting the skin, cornea and inner ear. GJB2 encodes the protein connexin-26 (Cx26) which forms hemichannels or gap junction channels allowing the passage of signalling molecules. Approximately 80% of KID syndrome patients carry a c.148G>A (p.D50N) mutation in GJB2, which results in aberrant channel function. We hypothesised that silencing of the mutant allele in patient keratinocytes using allele-specific siRNA could correct the channel function. First, to confirm whether patient keratinocytes with only one wildtype GJB2 allele formed functional channels following allele-specific siRNA treatment, GJB2+/- keratinocytes were generated using CRISPR/Cas9. The scrape-loading dye transfer (SLDT) assay showed no distinguishable difference in gap junction intercellular communication (GJIC) between GJB2+/- and GJB2+/+ cells, suggesting normal GJIC in GJB2+/- keratinocytes. Nineteen siRNAs were designed and tested in HeLa cells expressing wildtype or mutant GJB2-GFP transgene. A lead siRNA, was discovered, which potently inhibited the mutant mRNA and protein without affecting wildtype GJB2 expression. The efficacy of the lead siRNA was assessed using keratinocytes derived from a KID syndrome patient (KID-KC) harbouring heterozygous c.148G>A mutation. These cells displayed pathological features of KID syndrome, with reduced gap junction plaque formation, impaired GJIC and hyperactive hemichannels confirmed by immunostaining, SLDT, patch clamp and neurobiotin uptake assays. Following treatment with the siRNA, selective silencing of mutant GJB2 allele in KID-KCs was confirmed at mRNA and protein levels. Significant improvement of GJIC and reversal of hemichannel activity were detected, with the latter corrected to a level comparable to that recorded in normal keratinocytes. Furthermore, RNA-Seq analysis showed that only six genes in the KID-KC transcriptome were significantly altered by the siRNA treatment, suggesting low-level off-target effects. In conclusion, allele-specific siRNA silencing of pathogenic dominant GJB2 mutation could be a potential therapeutic intervention for KID syndrome

State of the art of audio- and video based solutions for AAL

Working Group 3. Audio- and Video-based AAL ApplicationsIt is a matter of fact that Europe is facing more and more crucial challenges regarding health and social care due to the demographic change and the current economic context. The recent COVID-19 pandemic has stressed this situation even further, thus highlighting the need for taking action. Active and Assisted Living (AAL) technologies come as a viable approach to help facing these challenges, thanks to the high potential they have in enabling remote care and support. Broadly speaking, AAL can be referred to as the use of innovative and advanced Information and Communication Technologies to create supportive, inclusive and empowering applications and environments that enable older, impaired or frail people to live independently and stay active longer in society. AAL capitalizes on the growing pervasiveness and effectiveness of sensing and computing facilities to supply the persons in need with smart assistance, by responding to their necessities of autonomy, independence, comfort, security and safety. The application scenarios addressed by AAL are complex, due to the inherent heterogeneity of the end-user population, their living arrangements, and their physical conditions or impairment. Despite aiming at diverse goals, AAL systems should share some common characteristics. They are designed to provide support in daily life in an invisible, unobtrusive and user-friendly manner. Moreover, they are conceived to be intelligent, to be able to learn and adapt to the requirements and requests of the assisted people, and to synchronise with their specific needs. Nevertheless, to ensure the uptake of AAL in society, potential users must be willing to use AAL applications and to integrate them in their daily environments and lives. In this respect, video- and audio-based AAL applications have several advantages, in terms of unobtrusiveness and information richness. Indeed, cameras and microphones are far less obtrusive with respect to the hindrance other wearable sensors may cause to one’s activities. In addition, a single camera placed in a room can record most of the activities performed in the room, thus replacing many other non-visual sensors. Currently, video-based applications are effective in recognising and monitoring the activities, the movements, and the overall conditions of the assisted individuals as well as to assess their vital parameters (e.g., heart rate, respiratory rate). Similarly, audio sensors have the potential to become one of the most important modalities for interaction with AAL systems, as they can have a large range of sensing, do not require physical presence at a particular location and are physically intangible. Moreover, relevant information about individuals’ activities and health status can derive from processing audio signals (e.g., speech recordings). Nevertheless, as the other side of the coin, cameras and microphones are often perceived as the most intrusive technologies from the viewpoint of the privacy of the monitored individuals. This is due to the richness of the information these technologies convey and the intimate setting where they may be deployed. Solutions able to ensure privacy preservation by context and by design, as well as to ensure high legal and ethical standards are in high demand. After the review of the current state of play and the discussion in GoodBrother, we may claim that the first solutions in this direction are starting to appear in the literature. A multidisciplinary 4 debate among experts and stakeholders is paving the way towards AAL ensuring ergonomics, usability, acceptance and privacy preservation. The DIANA, PAAL, and VisuAAL projects are examples of this fresh approach. This report provides the reader with a review of the most recent advances in audio- and video-based monitoring technologies for AAL. It has been drafted as a collective effort of WG3 to supply an introduction to AAL, its evolution over time and its main functional and technological underpinnings. In this respect, the report contributes to the field with the outline of a new generation of ethical-aware AAL technologies and a proposal for a novel comprehensive taxonomy of AAL systems and applications. Moreover, the report allows non-technical readers to gather an overview of the main components of an AAL system and how these function and interact with the end-users. The report illustrates the state of the art of the most successful AAL applications and functions based on audio and video data, namely (i) lifelogging and self-monitoring, (ii) remote monitoring of vital signs, (iii) emotional state recognition, (iv) food intake monitoring, activity and behaviour recognition, (v) activity and personal assistance, (vi) gesture recognition, (vii) fall detection and prevention, (viii) mobility assessment and frailty recognition, and (ix) cognitive and motor rehabilitation. For these application scenarios, the report illustrates the state of play in terms of scientific advances, available products and research project. The open challenges are also highlighted. The report ends with an overview of the challenges, the hindrances and the opportunities posed by the uptake in real world settings of AAL technologies. In this respect, the report illustrates the current procedural and technological approaches to cope with acceptability, usability and trust in the AAL technology, by surveying strategies and approaches to co-design, to privacy preservation in video and audio data, to transparency and explainability in data processing, and to data transmission and communication. User acceptance and ethical considerations are also debated. Finally, the potentials coming from the silver economy are overviewed.publishedVersio

Consonant colour and vocalism in the history of Irish

Wydział AnglistykiNiniejsza praca poświęcona jest związanym ze sobą kwestiom barwienia spółgłosek i wokalizmu w historii języka Irlandzkiego, ze szczególną uwagą na okres staro-irlandzki. Głównym twierdzeniem pracy jest to, że język staro-irlandzki miał trzy odrębne serie spółgłosek, polegających na barwieniu, oraz minimalny (wertykalny) system samogłosek składający się z dwóch tylko komponentów. Obrona głównego twierdzenia ma podwójny charakter: empiryczny i typologiczny. Część typologiczna bazuje na kompleksowym przeglądzie minimalnych systemów samogłosek w językach świata. Część empiryczna oparta jest na opisie morfologii czasownika staro-irlandzkiego, w którym występują trzy barwy spółgłoskowe, ale tylko dwie samogłoski.This dissertation deals with the related questions of consonant colour and vocalism in the history of Irish, focusing particularly on the Old Irish period. It argues that Old Irish had three distinct series of consonant colour, and a vertical vowel system of only two members. This position is defended typologically, by means of a comprehensive survey of minimal and vertical vowel systems in the cross-linguistic literature, and also empirically, through a detailed description of Old Irish verbal morphology in terms of a phonological system with three consonant colours and only two vowels

Molecular Characterisation of Antimicrobial Arsenal of Dictyostelium discoideum

In Entamoeba histolytica, Amoebapores and lysozymes are known to confer antimicrobial functions. The structure and functions of Amoebapores and lysozymes are evolutionally conserved, as evidenced by their analogous in many other organisms. Amoebapores and its structural counterparts have five α-helices that are connected by disulfide bonds, involving cysteine residues. Any protein with this typical structure is categorised under the SAPLIP family (Saposin-like protein) and the domain is called SAPLIP domain. SAPLIPs are known for their diverse cellular functions. The Amoebapores (M Leippe 1997) from E. histolytica and their structural and functional analogues, NK-lysin (porcine) and granulysin (human) are known for their antimicrobial and cytolytic functions. By contrast, the name giving Saposins (human) are necessary for lipid metabolism. Dictyostelium discoideum, has a multi-protein complex in which one component, called countin has a SAPLIP domain. Countin is required for aggregate size determination during development. Earlier work in our laboratory has found that the D. discoideum genome harbours 17 genes, which encode for 33 putative Amoebapore-like peptides (Apls). In addition, the D. discoideum genome comprises 15 genes coding for four different classes of lysozymes. These classes are c-type lysozymes (LyC), phage-type lysozymes (LyT4), Entamoeba-type lysozymes (LyEh), and the amoeba lysozymes (Alys), which is a unique class of lysozymes found in D. discoideum. As D. discoideum amoebae feed on bacteria, they are at a risk to encounter pathogenic bacteria. Therefore, it has been postulated that the Apls and lysozymes may play a synergistic role to kill and degrade the bacteria. My study has focused on three apl genes (aplD, aplJ, and aplP) and two lysozyme genes (lyC2 and lyC3), based on collaborator reports that these genes are implicated in bacterial destruction. The phenotypic characterisations were carried out with the single gene KO mutants for the corresponding genes. All the gene disruptions were performed in wild type Ax2 background. Among all five KO mutants tested for growth on bacteria, the aplD¯ showed growth defects on several strains of K. pneumoniae. The aplP¯ and lyC2¯ were defective for growth only on the capsule defective mutant of Kp52145 (K¯). As the aplD¯ was defective for growth on different K. pneumoniae strains, further phenotypic characterisations were carried out for the aplD¯. In accordance with growth defects on K. pneumoniae, the aplD¯ was also defective for intracellular killing of K. pneumoniae (KpLM21). Moreover, the wild type Ax2 cells confronted with KpLM21 showed at least three fold up-regulation of the aplD transcript (qRT-PCR). Similar qRT-PCR experiments with axenic Ax2 revealed that the transcription of aplD is moderately up-regulated during axenic growth, whereas sturdily up-regulated during development and peaked at the slug stage. This spontaneous up-regulation of aplD transcript during Ax2 development, which was performed totally in the absence of bacteria, prompted me to do the slug infection experiments with the aplD¯. As the D. discoideum cells stop feeding at the onset of development they may need an armament to protect their developing structures from foreign intruders. In fact, the migrating slugs are highly prone to infection when compared with the other stages of development. Following this, the slug infection experiments were performed with a GFP expressing strain of K. pneumoniae (KpGFP) and an E. coli strain that expressed DsRed (E. coli DsRed). The Ax2 slugs were used as controls. Unlike Ax2 slugs, the aplD¯ slugs infected with KpGFP showed massive KpGFP accumulation which corroborates with growth and killing defects of aplD¯ with K. pneumoniae. In contrast, the aplD¯ slugs infected with E. coli DsRed were able to slough off E. coli DsRed in their slime trail. However, I could show that the aplD¯ slugs were unaffected in their ability to form S cells and to sequester toxin (EtBr). Apart from their reduced ability to defend against K. pneumoniae, the aplD¯ also showed several morphogenetic defects during development. This includes late stream breaks (12 h), delayed formation of smaller slugs (18 h) and fruiting bodies (~ 26 h), and a relatively reduced spore formation efficiency (sfe). Nevertheless, the spore morphology and spore viability were unaffected in aplD¯. These observations may point to an additional role for aplD during D. discoideum development. On the other hand, it is also possible that the aplD¯ is immunocompromised and therefore it is showing several developmental defects. In summary, this study shows that the aplD is crucial for the growth of D. discoideum on K. pneumoniae and also for the intracellular killing of K. pneumoniae. Additionally, aplD is required to prevent K. pneumoniae accumulation in the D. discoideum slugs.In Entamoeba histolytica sind die Amoebapores und die Lysozyme dafür bekannt, antimikrobielle Funktionen zu übernehmen. Die Struktur und die Funktion sowohl der Amoebapores als auch der Lysozyme sind evolutionär konserviert, was durch das Vorhandensein analoger Proteine in einer Vielzahl anderer Organismen bewiesen ist. Die Amoebapores und deren strukturelle Verwandte sind durch fünf α-Helices charakterisiert, die über Disulfidbrücken zwischen Cysteinresten miteinander verbunden sind. Jedes Protein, das diese typische Struktur zeigt, wird der SAPLIP-Familie (Saposin-like protein), den Saposin-ähnlichen Proteinen, zugeordnet. Die entsprechende Domäne wird demnach SAPLIP-Domäne genannt. Die SAPLIPs sind für ihre vielfältigen Funktionen bekannt. Die Amoebapores aus E. histolytica und deren strukturell und funktionell analoge Proteine, das NK-lysin aus dem Schwein und das Granulysin aus dem Menschen, sind für ihre antimikrobiellen und cytolytischen Funktionen bekannt. Die namengebenden Saposine des Menschen sind dagegen für den Lipidstoffwechsel wichtig. In Diyctyostelium discoideum gibt es eine Komponente eines Multi-Protein-Komplexes, das countin, das ebenfalls eine SAPLIP-Domäne besitzt. Countin ist notwendig für die Bestimmung der Aggregatgröße während der Entwicklungsphase von D. discoideum. Als Ergebnis einer früheren Arbeit in diesem Labor konnte gezeigt werden, dass das Genom von D. discoideum 17 Gene aufweist, die für 33 putative Amoebapore-ähnliche Peptide (Apls) codieren. Zudem finden sich im Genom von D. discoideum 15 Gene, die für Lysozyme aus vier verschiedenen Klassen codieren. Dabei handelt es sich um die Klassen der c-Typ Lysozyme (LyC), T4 Phagen-Lysozyme (LyT4), Lysozyme des Entamoeba-Typs (LyEh) und die Amoeba-Lysozyme (Alys), eine ausschließlich in D. discoideum vorkommende Lysozym-Klasse. Da das Amöbenstadium von D. discoideum Bakterien als Nahrungsquelle nutzt, besteht für diese auch ein hohes Risiko, auf pathogene Bakterien zu treffen. Daher wurde angenommen, dass die Apls und die Lysozyme eine synergistische Rolle beim Töten und Degradieren von Bakterien spielen könnten. In meiner Arbeit habe ich mich auf die Untersuchung von drei Apl-Genen (aplD, aplJ und aplP) sowie zwei Lysozym-Genen (lyC2 und lyC3) konzentriert, für die bereits von einer kooperierenden Arbeitsgruppe berichtet wurde, dass sie möglicherweise eine Rolle beim Abbau von Bakterien spielen. Die phänotypischen Charakterisierungen wurden mithilfe von Deletionsmutanten für die einzelnen Gene durchgeführt. Als Ausgangsstamm für die Herstellung dieser Mutanten wurde jeweils der Wildtyp-Stamm Ax2 verwendet. Alle fünf Deletionsmutanten wurden bezüglich ihrer Fähigkeit, auf Bakterien zu wachsen, untersucht. Dabei zeigte die aplD--Mutante Wachstumsdefekte auf den meisten der getesteten Stämme von Klebsiella pneumoniae, während die Mutanten aplP- und lyC2- nur einen Wachstumsdefekt auf dem Klebsiella-Stamm Kp52145 (K-) zeigten, der sich wiederum dadurch auszeichnet, dass er eine veränderte Kapsel aufweist. Da das Wachstum der aplD--Mutante auf verschiedenen Stämmen von K. pneumoniae gestört war, wurde diese Mutante im Folgenden genauer phänotypisch charakterisiert. In Übereinstimmung mit den Wachstumsdefekten auf K. pneumoniae zeigte sich, dass die aplD--Mutante zudem in ihrer Fähigkeit beeinträchtigt war, intrazelluläre K. pneumoniae (KpLM21) zu töten. Des Weiteren zeigten Zellen des Wildtyps Ax2, die mit KpLM21 konfrontiert wurden, eine mindestens dreifache Hochregulierung das aplD-Transkripts (qRT-PCR). Ähnliche qRT-PCR-Experimente mit axenischen Ax2-Zellen ergaben, dass die Transkription von aplD während des axenischen Wachstums nur leicht hochreguliert wird, wohingegen dies während der Entwicklungsphase sehr stark der Fall ist, mit einem Maximum im sogenannten Slug-Stadium. Die spontane Hochregulierung der aplD-Transkription während der Entwicklung von Ax2-Zellen passierte in der kompletten Abwesenheit von Bakterien. Diese Beobachtung veranlasste mich dazu, das Infektionsexperiment mit dem Slug-Stadium von aplD--Mutanten durchzuführen. Da D. discoideum Zellen mit dem Beginn der Entwicklungsphase aufhören zu fressen, ist es anzunehmen, dass sie Abwehrmoleküle benötigen, um ihre sich entwickelnden Strukturen vor fremden Eindringlingen zu schützen. Tatsächlich sind wandernde Slugs im Vergleich zu den anderen Stadien der Entwicklungsphase hoch anfällig für Infektionen. Daraufhin wurden Infektionsexperimente mit Slugs mit einem GFP exprimierenden Stamm von K. pneumoniae (KpGFP) und einem DsRed exprimierenden Stamm von E. coli (E. coli DsRed) durchgeführt. Slugs des Wildtyps Ax2 wurden dabei als Kontrolle verwendet. Anders als der Wildtyp wiesen die mit KpGFP infizierten aplD--Slugs eine massive Akkumulation der Bakterien auf, was mit den entsprechenden Wachstums- und Tötungsdefekten dieser Mutante übereinstimmt. Im Gegensatz dazu waren die mit E. coli DsRed infizierten aplD--Slugs in der Lage, diese Bakterien in ihre Schleimspur mit abzugeben. Zudem konnte ich zeigen, dass die aplD--Slugs nicht in ihrer Fähigkeit beeinträchtigt waren, S-Zellen zu bilden und Giftstoffe (EtBr) abzusondern. Neben der verminderten Fähigkeit, sich gegen K. pneumoniae zu verteidigen, zeigte die aplD--Mutante außerdem einige morphogenetischen Defekte während der Entwicklungsphase. Hierzu gehörten Unterbrechungen in den Hyphen des späten Streaming-Stadiums (12 h), die verzögerte Bildung von kleineren Slugs (18 h) und Fruchtkörpern (~26 h) sowie eine etwas reduzierte Sporenbildungseffizienz. Gleichwohl waren die Morphologie und die Viabilität der Sporen in der aplD--Mutante nicht beeinträchtig. Diese Beobachtungen könnten auf eine zusätzliche Rolle für AplD während der Entwicklungsphase hindeuten. Andererseits wäre es auch denkbar, dass die aplD--Mutante generell immunsupprimiert ist und somit mehrere Entwicklungsdefekte aufweist. Zusammenfassend konnte ich in dieser Arbeit zeigen, dass aplD von entscheidender Bedeutung für das Wachstum von D. discoideum auf K. pneumoniae und auch für das intrazelluläre Töten dieser Bakterien ist. Außerdem ist aplD notwendig, um die Akkumulation von K. pneumoniae in den Slugs von D. discoideum zu verhindern

Change blindness: eradication of gestalt strategies

Arrays of eight, texture-defined rectangles were used as stimuli in a one-shot change blindness (CB) task where there was a 50% chance that one rectangle would change orientation between two successive presentations separated by an interval. CB was eliminated by cueing the target rectangle in the first stimulus, reduced by cueing in the interval and unaffected by cueing in the second presentation. This supports the idea that a representation was formed that persisted through the interval before being 'overwritten' by the second presentation (Landman et al, 2003 Vision Research 43149–164]. Another possibility is that participants used some kind of grouping or Gestalt strategy. To test this we changed the spatial position of the rectangles in the second presentation by shifting them along imaginary spokes (by ±1 degree) emanating from the central fixation point. There was no significant difference seen in performance between this and the standard task [F(1,4)=2.565, p=0.185]. This may suggest two things: (i) Gestalt grouping is not used as a strategy in these tasks, and (ii) it gives further weight to the argument that objects may be stored and retrieved from a pre-attentional store during this task

Connectionist and Process Modelling of Long-Term Sequence: The Integration of Relative Judgements, Representation and Learning

A large amount of psychological research is devoted to the representation of sequences. It is a fundamental upon which most of the processes of cognition are based. Despite the amount of research into sequencing, there has been relatively little investigation of the types of representations generated in response to sequential information. These representations must allow operations to be performed on individual elements, as well as operations between and among elements. This thesis begins by describing the effects found when subjects are asked to make relative order judgements using sequences which are in long-term storage (e.g. the alphabet or number series). These effects are then used to examine some of the theories and models which have been developed, with a view toward generating a general purpose mechanism with the ability to model all of the different effects found with different types of stimuli. In the course of developing the new model, the neuropsychological findings in the area are examined in Chapter Two. Deficit studies and neuropsychological investigations are able to isolate which aspects of a task appear to be processed in different structures. If a patient loses the ability to perform one aspect of a task but not another, trying to model both of these aspects in one network may be counter-productive. The construction of a new model is begun in Chapter Three. This model is developed in the PDP environment as it offers the ability to change (learn) as a result of experience, and demands a more thorough definition of the mechanisms operating within the network. Chapter Four details a formal definition of the Serial Order Network (SON) model outlined in Chapter Three, including a section devoted to relative order judgements, called the Response Generation Network (RGN) and undertakes a comparison between the SON/RGN and Poltrock's (1990) random walk model described in Chapter One. A review of some of the sequence learning networks developed is undertaken in Chapter Five. This review is used to choose sequence learning networks, which may be used to learn the type of representation needed. These sequence learning networks are investigated in Chapter Six for their ability to learn the sequence incrementally. It is determined that not one of these networks is appropriate. Thus, in Chapter Seven a recitation mechanism is added directly to the representation in the SON. The resulting system is investigated, and it is determined that the system's success in recitation is not dependent on an idiosyncratic setting of the parameters in the network. The definition of the SON and complementary recitation network is not sufficient. The resulting mechanisms should also be compatible with the developmental literature for both children learning sequences for the first time, and adults learning novel sequences. A review of this literature is conducted in Chapter Eight. It is also necessary to explain how this SON representation can be developed, and how the model can be used to explain the seeming hierarchic nature of some sequences. In Chapter Nine a mechanism designed to mimic a hierarchical structure for the representation of a sequence is developed. A learning mechanism is defined for the resulting system. This system is then investigated for its ability to recite both hierarchic and non-hierarchic sequences, and to generate the relative order and developmental effects referred to in Chapters One and Nine. The model developed in this thesis is the only model existent which is able to explain sequence learning, representation and relative order effects, and represents an advance in the approach to modelling sequence information

Network based analysis to identify master regulators in prostate carcinogenesis

Prostate cancer (PCa) is the second most common tumor diagnosed in man, for which robust prognostic markers and novel targets for therapy are lacking. Major challenges in PCa therapeutical management arise from the marked intra and inter-tumors heterogeneity, hampering the discernment of molecular subtypes that can be used to guide treatment decisions. For this reason, virtually all patients undergoing standard of care androgen deprivation therapy for locally advanced or metastatic cancer, will eventually progress into the more aggressive and currently incurable form of PCa, referred to as castration resistant prostate cancer (CRPC). By exploiting the richness of information stored in gene-gene interactions, I tested the hypothesis that a gene regulatory network derived from transcriptomic profiles of PCa orthografts can reveal transcriptional regulators to be subsequently adopted as robust biomarkers or as target for novel therapies. Among the 1308 regulons identified from the preclinical models, Cox regression analysis coherently associated JMJD6 regulon activity with disease-free survival in three clinical cohorts, outperforming three published prognostic gene signatures (TMCC11, BROMO-10 and HYPOXIA-28). Given its potential role in a number of cancers, in-depth investigations of JMJD6 mediated function in PCa is warranted to test if it has a driver role in tumor progression. Encouraged by the predictive abilities of the gene regulatory network inferred from transcriptomics data, I explored the possibility of integrating the regulons structure with data from the proteomes of the same preclinical orthografts studied by RNA sequencing. This approach leverages the complementarity between gene and protein expression, to increase the robustness of the statistical analysis. Similar to gene-gene co-expression profiles, protein-protein co-expression data can provide a distinct representation of the molecular alterations underlying a biological phenotype. By implementing a pipeline to integrate modules derived from transcriptomic based regulons and proteinprotein interactions respectively from matched RNA-seq and quantitative proteomic data, I obtained 516 joint modules entailing a median of four protein complexes (range 1-41) per individual transcription factor regulon, providing new insight into its regulatory mechanisms. In the final step of the analysis, a permutation-based enrichment of the genes/proteins integrative modules implicated MID1 (an E3 ubiquitin ligase belonging to the family of tripartite motif containing protein) to be a driver transcriptional regulator in CRPC. In fact, MID1 module was the only candidate for which gene-gene and proteinprotein interactions were supported (p-value < 0.05) by both differentially expressed genes and proteins obtained from the CRPC vs PC contrast. Finally, I wished to test the usefulness of a network based investigation as a tool to identify predictors of treatment response. To this end, I obtained transcriptomics data from an in vivo subcutaneous xenograft treatment experiment (namely mychophenolic acid or abiraterone/ARN-509 as stand alone treatment or in combination) and determined which regulons were inferred to be active in the tumours following treatment. The androgen receptor positive human LNCaP C4-2b prostate cancer cells were injected into mice. The effects of treatment were assessed by collecting serial tumor sizes and by performing RNAseq at the designed endpoint of the study. Noteworthy, the gene graph enrichment analysis provided novel hypothesisbehind the anti- proliferative effect of mychophenolic acid (MPA), suggesting the SET proto-oncogene to be a target for MPA mediated suppression of proliferation. Of note, standard gene-set enrichment analysis, without input on specific gene-gene interactions, was not effective in prioritising the SET protooncogene, demonstrating the usefulness of the network based investigation. Collectively, data presented in this thesis provides an alternative perspective for the analysis of multi-omics profiles from PCa and highlights the importance of gene-gene and protein protein interactions in prostate cancer growth and progression