Identification and characterization of a novel gene associated with hereditary retinal disorder(s)

Bakgrunn: Retinitis pigmentosa (RP) er kjent som den vanligst forekommende arvelige retinale dystrofi med en prevalens på nesten 1:4000 på verdensbasis. RP skyldes i utgangspunktet tap av stav fotoreseptorer, ledsaget av redusert nattsyn og innsnevring av synsfeltet. Senere oppstår tap av tapp fotoreseptorer som fører til tap av sentralsyn. I en familie der to søsken ble diagnostisert med ikke-syndromisk RP, fant vi en homozygot mutasjon i genet all-trans retinoic acid induced differentiation factor (ATRAID) som førte til en tidlig terminering av proteinet. Mål: ATRAID er et lite karakterisert protein. Målet var å karakterisere ATRAID mer detaljert og å finne støtte for dets rolle i utviklingen av RP. Måloppgave 1: Funksjonell karakterisering av ATRAID. Måloppgave 2: Undersøk subcellulær lokalisering av ATRAID. Måloppgave 3: Finn den mulige kausale mutasjonen som forårsaker ikke-syndromisk RP i en bestemt familie. Videre å undersøk fenotypen til mus som ikke utrykker Atraid. Metoder: Oppgave 1: In silico analyse, Cellekultur, Konstruksjon av retrovirale vektorer, Transduksjon, Transient transfeksjon, Subcellulær fraksjonering, Inhibitorbehandling, Antigen «retrieval», Immunfluorescensanalyse, Immunoblotanalyse. Oppgave 2: Cellekultur, Konstruksjon av retrovirale vektorer, Transduksjon, Antigen «retrieval», Immunfluorescensanalyse. Oppgave 3: Klinisk undersøkelse, Eksomsekvensering Cellekultur, Immunoblotanalyse, Dyremodell, Histopatologisk farging, Immunfluorescensanalyse, Transmisjonselektronmikroskopi. Resultater: Artikkel 1: Hovedisoformen av ATRAID er en N-glykosylert form av Isoform C (Iso C). Isoform A ser ut til å bli raskt nedbrutt av proteasomer etter syntesen. I celler transfektert med et konstrukt som er predikert å uttrykke Isoform B, var det isteden Iso C som ble syntetisert. Iso C viser en punktformet fordeling i cytoplasmaet med høy konsentrasjon i det juxtanukleære området. Høy konsentrasjon av ATRAID ble sett i Golgi-området. I tillegg fant vi at lokaliseringen av proteinet overlappet med både RAB11, en markør for resirkulerende endosomer, og de lysosymale markørproteinene LAMP1 og LAMP2. Artikkel 2: Lokalisering av ATRAID Iso C er ikke begrenset til lysosomene. Den følger tett lokaliseringen av transferrin og transferrinreseptorene som er lokalisert til clathrin-belagte vesikler. Dessuten finnes det sammen eller nær RAB5, EEA1, RAB7 og RAB9 som er markører for henholdsvis tidlige og sene endosomer. Artikkel 3: To søsken viste typiske trekk ved ikke-syndromisk retinitis pigmentosa, med tap av perifert synsfelt, reduserte fullfelts elektroretinogram amplituder, og tap av RPE og fotoreseptorer visualisert ved OCT. Eksomsekvensering avdekket en variant, NM_001170795.3(ATRAID), c.120dupG, p.Ser41Glufs*13, som fører til prematur terminering av ATRAID protein syntese. I en Atraid-/- musemodell ble tegn på RPE og fotoreseptor degenerasjon sett hos to av fem mus. Konklusjoner: I dette prosjektet fant vi at hoved isoformen av ATRAID (Iso C) var til stede ikke bare i lysosomer, men også i endosomer, noe som tyder på at den kan spille en rolle i det endosomale-lysosomale systemet eller i andre cellulære trafikkveier. Hos våre RP pasienter resulterte den observerte ATRAID-varianten i tap av proteinet. Varianter i ATRAID har ikke vært assosiert med RP tidligere. Nedbrytning av fotoreseptorer ble sett hos to av fem Atraid-/- mus. Ytterligere studier er nødvendig for å bekrefte om musene er en passende modell for RP. Vår hypotese er at tap av ATRAID kan være assosiert med utvikling av RP, blant annet gjennom dysregulering av de endosomale, lysosomale, eller andre cellulære trafikkveier, enten i retinalt pigmentepitel eller fotoreseptorer.Background: Retinitis pigmentosa, with a prevalence of almost 1:4000 worldwide, is known as the most common inherited retinal dystrophy. It is initially characterized by loss of rod photoreceptors accompanied by reduced night vision and constriction of the visual field. Later, loss of cone photoreceptors occurs which leads to loss of central vision. In a family where two siblings were diagnosed with non-syndromic RP, we found a homozygous mutation in all‐trans retinoic acid‐induced differentiation factor (ATRAID) gene leading to a premature termination of the protein. Objectives: ATRAID is a poorly characterized protein. The aim was to characterize ATRAID in greater detail and to provide support for its role in the development of RP. Aims paper 1: Functional characterization of ATRAID. Aims paper 2: Investigate subcellular localization of ATRAID. Aims paper 3: Find the causative mutation responsible for non-syndromic RP in a particular family and examine the phenotype of an Atraid-/- mouse line. Methods: Paper 1: In silico analysis, Cell culture, Construction of retroviral vectors, Transduction, Transient transfection, Subcellular fractionation, Inhibitor treatment, Antigen retrieval, Immunofluorescence analysis, Immunoblot analysis. Paper 2: Cell culture, Construction of retroviral vectors, Transduction, Antigen retrieval, Immunofluorescence analysis. Paper 3: Clinical examination, Exome sequencing, Cell culture, Immunoblot analysis, Animal model, Histopathology staining, Immunofluorescence analysis, Transmission electron microscopy. Results: Paper 1: The main isoform of ATRAID is Isoform C (Iso C) which is highly N-glycosylated. Isoform A appears to be rapidly degraded by proteasomes after its production. In cells transfected with a construct predicted to express Isoform B, the protein detected was indistinguishable from Iso C. Iso C shows a punctuate distribution in the cytoplasm with high concentration in the juxtanuclear area. High concentration of ATRAID was seen in the Golgi area and its overlap with RAB11, marker of recycling endosomes and LAMP1 and LAMP2, lysosomal markers were found. Paper 2: ATRAID Iso C is not restricted to the lysosomes. It follows closely the localization of transferrin and the transferrin receptors that are localized to clathrin-coated vesicles. Moreover, it is found together or near RAB5, EEA1, RAB7 and RAB9 that are markers of early and late endosomes, respectively. Paper 3: The two siblings showed typical features of non-syndromic retinitis pigmentosa with loss of peripheral visual field, reduced full-field electroretinogram amplitudes, and loss of RPE and photoreceptors on OCT. Exome sequencing revealed a variant, NM_001170795.3(ATRAID), c.120dupG, p.Ser41Glufs*13, that leads to loss of ATRAID expression. In Atraid-/- mouse models, signs of RPE and photoreceptor degeneration were seen in 2/5 mice. Conclusions: In this project, we found that the main isoform of ATRAID (Iso C) was present not only in lysosomes but also in endosomes showing that it may play a role in the endosomal–lysosomal pathway or other cellular trafficking pathways. In our non-syndromic retinitis pigmentosa patients, the observed ATRAID variant resulted in a loss of ATRAID. Variants in ATRAID have not been associated with RP previously. Degradation of photoreceptors was seen in two out of five Atraid-/- mice. Further studies are needed to confirm if the mice are a suitable model for RP. We hypothesize that loss of ATRAID could be associated with the development of RP, among others through dysregulation of the endosomal lysosomal or other cellular trafficking pathways either in the retinal pigment epithelium or the photoreceptors.Doktorgradsavhandlin

Diving into the Depths of Belladonna: Uncovering the Secrets of the Deadly Nightshade

Tropane alkaloids, such as hyoscyamine and scopolamine, are vital medicinal compounds found in various therapeutic formulations. These alkaloids are synthesized within the roots of specific genera of the Solanaceae family through a complex, partially understood multistep pathway. In efforts to deepen our comprehension of tropane alkaloid biosynthesis, a novel de novo transcriptome assembly was crafted for Deadly Nightshade (Atropa belladonna). A crucial intermediate in the biosynthesis of hyoscyamine and scopolamine is littorine, formed by the condensation of tropine and phenyllactic acid. Phenyllactic acid originates from phenylalanine through transamination to phenylpyruvate. Through transcriptome analysis, a distinct aromatic amino acid aminotransferase (ArAT), designated Ab-ArAT4, was identified. This enzyme coexists with known tropane alkaloid biosynthesis genes in the roots of A. belladonna. Silencing Ab-ArAT4 resulted in diminished synthesis of hyoscyamine and scopolamine due to reduced phenyllactic acid levels. In vitro studies revealed that recombinant Ab-ArAT4 primarily catalyzes the initial step of phenyllactic acid synthesis: the transamination of phenylalanine to phenylpyruvate

Control of modular multilevel converters in high voltage direct current power systems

This thesis focuses on a comprehensive analysis of Modular Multilevel Converters (MMC) in High Voltage Direct Current (HVDC) applications from the viewpoint of presenting new mathematical dynamic models and designing novel control strategies. In the first step, two new mathematical dynamic models using differential flatness theory (DFT) and circulating currents components are introduced. Moreover, detailed step-by-step analysis-based relationships are achieved for accurate control of MMCs in both inverter and rectifier operating modes. After presenting these new mathematical equations-based descriptions of MMCs, suitable control techniques are designed in the next step. Because of the nonlinearity features of MMCs, two nonlinear control strategies based on direct Lyapunov method (DLM) and passivity theory-based controller combined with sliding mode surface are designed by the use of circulating currents componentsbased dynamic model to provide a stable operation of MMCs in HVDC applications under various operating conditions. The negative effects of the input disturbance, model errors and system uncertainties are suppressed by defining a Lyapunov control function to reach the integralproportional terms of the flat output errors that should be finally added to the initial inputs. Simulation results in MATLAB/SIMULINK environment verify the positive effects of the proposed dynamic models and control strategies in all operating conditions of the MMCs in inverter mode, rectifier mode and HVDC applications.Esta tese visa proceder a uma análise abrangente de conversores multinível modulares (MMC) para transmissão a alta tensão em corrente contínua (HVDC), almejando apresentar novos modelos matemáticos em sistemas dinâmicos e projetar novas estratégias de controlo. Na primeira etapa são introduzidos dois novos modelos matemáticos dinâmicos que usam differential flatness theory e as componentes de correntes circulantes. Ainda, é estabelecida uma modelação matemática para o controlo preciso dos MMCs, operando em modo inversor ou modo retificador. Depois de apresentar as novas equações matemáticas, as técnicas de controlo mais adequadas são delineadas. Devido às características não lineares dos MMCs, são projetadas duas estratégias de controlo não-lineares baseadas no método direto de Lyapunov e no controlo do tipo passivity theory-based combinado com controlo por modo de deslizamento através do uso de modelos dinâmicos baseados em correntes circulantes para fornecer uma operação estável aos MMCs em aplicações de HVDC sob várias condições de operação. Os efeitos negativos das perturbações de entrada, erros de modelação e incertezas do sistema são suprimidos através da definição da função de controlo de Lyapunov para alcançar os termos de integraçãoproporcionalidade dos erros de saída para que possam finalmente ser adicionados às entradas iniciais. Os resultados da simulação computacional realizados em ambiente MATLAB/SIMULINK verificam os efeitos positivos dos modelos dinâmicos propostos e das novas estratégias de controlo em todas as condições de operação dos MMCs no modo inversor, retificador e em aplicações HVDC

Enhancing material selection and inflow efficiency for sustainable equipment manufacturing : a case study

The purpose of this thesis is to investigate strategies for optimizing material inflows and increasing resource efficiency in equipment manufacturing. The focus of this study is on improving sustainability in supply chain practices, aligning operations with European Sustainability Reporting Standards (ESRS) regulations, specifically the ESRS E5. This study utilizes a case study at The Company to identify key methodologies for tracking material flows, improving data accuracy, and implementing circular economy principles. It will also assess how these strategies can help reduce waste, energy consumption, and emissions while remaining compliant with ESRS requirements. The research will also examine how better resource management can boost supply chain resilience, visibility, and overall sustainability. Semi-structured interviews were used to identify the main factors. The Delphi method was used to determine the main factors using expert opinions, followed by material flow analysis (MFA). In this study, factors Enhancing material selection and inflow efficiency for Sustainable Equipment manufacturing factors include assessment and monitoring, manufacturability, economics, accessibility, and the environment. Also, the MFA method, which is enhanced by CNN and RNN models (with 98% and 96% accuracy, respectively), has been addressed by improving material flow tracking and identifying inefficiencies such as excess inventory due to forecast errors

Initializing cloud-based music tutorial based on AI

Based on cloud, with the help of artificial intelligence and machine learning algorithms, PiaTut is developing as an advanced tutorial to turn piano tutorial into education for all. Google-based, PiaTut engages users with instant feedback, melodies produced within the manager, and personalized lessons splitting users by their difficulties upon the enrolment. Unlike most other tutoring systems, PiaTut not only learns from the user but also provides multiple machine learning models including the LSTM neural networks specifically trained on a variety of piano compositions. This guarantees both effective and entertaining learning processes that allow for the effective development of knowledge and skills on a safe, international level, without geographical limitations. As a platform based on the cloud service, PiaTut shows that machine learning can provide high-quality learning experience to learners in piano education without barriers of time, space, and cost at an unprecedented level. The next versions will encompass offering more instruments, improving genre-based models, and improving the ML algorithms for better user experience. This work further emphasizes how the use of AI and cloud computing has revolutionized the use of educational technology by bridging the gap between the conventional effectiveness of teaching and utilizing the facilities of the modern technology. PiaTut will include more musical instruments, genre-specific models, and advanced ML algorithms in the future to further enhance its role as an innovative tool for global music education

Diving into the Depths of Belladonna: Uncovering the Secrets of the Deadly Nightshade

Tropane alkaloids, such as hyoscyamine and scopolamine, are vital medicinal compounds found in various therapeutic formulations. These alkaloids are synthesized within the roots of specific genera of the Solanaceae family through a complex, partially understood multistep pathway. In efforts to deepen our comprehension of tropane alkaloid biosynthesis, a novel de novo transcriptome assembly was crafted for Deadly Nightshade (Atropa belladonna). A crucial intermediate in the biosynthesis of hyoscyamine and scopolamine is littorine, formed by the condensation of tropine and phenyllactic acid. Phenyllactic acid originates from phenylalanine through transamination to phenylpyruvate. Through transcriptome analysis, a distinct aromatic amino acid aminotransferase (ArAT), designated Ab-ArAT4, was identified. This enzyme coexists with known tropane alkaloid biosynthesis genes in the roots of A. belladonna. Silencing Ab-ArAT4 resulted in diminished synthesis of hyoscyamine and scopolamine due to reduced phenyllactic acid levels. In vitro studies revealed that recombinant Ab-ArAT4 primarily catalyzes the initial step of phenyllactic acid synthesis: the transamination of phenylalanine to phenylpyruvate

Exploring the History, Uses, and Dangers of Belladonna: Unveiling the Mysteries of the Deadly Nightshade

Belladonna (Atropa belladonna) is a plant with a rich history, renowned for its beauty and lethality. This review synthesizes existing literature to provide an overview of belladonna, covering its historical significance, medicinal uses, toxicity, regulatory status, and cultural references. Belladonna's alkaloids, such as atropine and scopolamine, have medicinal value but pose significant risks due to their high toxicity. Recent research explores their pharmacotherapeutic potential, yet controversies persist regarding their safety and regulation. Safety precautions are paramount due to the plant's extreme toxicity. This review underscores the need for informed decision-making and further research to navigate the complexities of belladonna's use in contemporary healthcare

O papel dos novos processos de registro na emissão de documentos de propriedade

In different societies, the advancement and growth of countries in all economic, industrial, cultural, etc. areas has been directly related to the extent of development in the field of modern communication technologies, and therefore countries are on the path to the development of these modern sciences, and new scientific technologies all aspects of life of the individual and social, has set a record of popular influence.One of the most sophisticated systems in the world is the registration system of each country, which is based on legal requirements and legal obligations, and plays an essential role in legal health and judicial stability. What is of great importance in the organization of registration of documents and real estate, which itself is a subdivision of the judiciary, is the registration of the rights of individuals and their property, and the registration organization from the traditional system to the new system of cadastre with interest the acquisition of advanced equipment and is undergoing tremendous transformation in this organization and operation of the cadastre area, issuance of property documents for the cadastre of land and property, the issuance and development of electronic signatures, the launch of an electronic inquiry system Register and E-commerce Real Estate Transactions, Setting Up and Enforcement of Electronic Registration System ( Immediate ) Effect New registration registrations.En diferentes sociedades, el avance y crecimiento de países en todas las áreas económicas, industriales, culturales, etc., se ha relacionado directamente con el grado de desarrollo en el campo de las tecnologías modernas de comunicación, y por lo tanto, los países están en el camino hacia el desarrollo de estos las ciencias modernas y las nuevas tecnologías científicas, todos los aspectos de la vida de los individuos y las redes sociales, han establecido un récord de influencia popular.Uno de los sistemas más sofisticados del mundo es el sistema de registro de cada país, que se basa en requisitos legales y obligaciones legales, y desempeña un papel esencial en la salud legal y la estabilidad judicial. Lo que es de gran importancia en la organización del registro de documentos y bienes inmuebles, que a su vez es una subdivisión de la judicatura, es el registro de los derechos de las personas y sus bienes, y la organización de registro del sistema tradicional al nuevo sistema de Catastro con interés en la adquisición de equipos avanzados y está experimentando una tremenda transformación en esta organización y operación del área de catastro, emisión de documentos de propiedad para el catastro de tierras y propiedades, la emisión y desarrollo de firmas electrónicas, el lanzamiento de un sistema de consulta electrónica Registro y comercio electrónico Transacciones inmobiliarias, establecimiento y aplicación del sistema electrónico de registro Efecto (inmediato) ¬ Nuevos registros de registro.Em diferentes sociedades, o avanço e crescimento dos países em todas as áreas econômicas, industriais, culturais, etc. tem estado diretamente relacionado com a extensão do desenvolvimento no campo das modernas tecnologias de comunicação e, portanto, os países estão no caminho para o desenvolvimento dessas tecnologias. ciências modernas e novas tecnologias científicas todos os aspectos da vida do indivíduo e social, estabeleceu um recorde de influência popular.Um dos sistemas mais sofisticados do mundo é o sistema de registro de cada país, baseado em requisitos legais e obrigações legais, e desempenha um papel essencial na saúde jurídica e na estabilidade judicial. O que é de grande importância na organização do registro de documentos e imóveis, que em si é uma subdivisão do Judiciário, é o registro dos direitos dos indivíduos e de suas propriedades, e a organização do registro do sistema tradicional ao novo sistema de registro. cadastrar com interesse a aquisição de equipamentos avançados e está passando por uma tremenda transformação nesta organização e operação da área de cadastro, emissão de documentos de propriedade para o cadastro de terras e propriedades, emissão e desenvolvimento de assinaturas eletrônicas, lançamento de sistema eletrônico de averiguação Cadastro e Transações Imobiliárias de E-commerce, Implantação e Fiscalização Imediata do Sistema de Registro Eletrônico ¬ Novos registros cadastrais

Collaborative systems and business models to support the forest products industry

La concurrence internationale, notamment dans l'industrie des produits forestiers, est en constante augmentation et elle oblige les entreprises à produire des produits novateurs, à satisfaire la demande de plus en plus personnalisée des clients et à chercher l'accès à de nouveaux marchés. Néanmoins, les entreprises n'ont pas nécessairement la capacité d'atteindre ces réalisations individuellement. Par conséquent, établir une collaboration avec d'autres organisations pourrait les aider à résoudre des problèmes, qui peuvent ne pas être résolus individuellement. Dans cette recherche, la pertinence de la collaboration dans l'industrie des produits forestiers et des modèles d'affaires possibles pour l'établissement d'une cour de triage forestier sont étudiés, dans le but d'aider les investisseurs potentiels, les analystes commerciaux et les gestionnaires à avoir une analyse approfondie et rapide des aspects commerciaux clés de ces activités. La première partie de cette recherche examine les principaux moteurs, avantages, facilitateurs et défis des collaborations dans l'industrie des produits forestiers, afin d'aider les entreprises à identifier les mécanismes de collaboration les mieux adaptés à leur initiative de collaboration particulière. Un cadre conceptuel est élaboré pour identifier les diverses composantes affectant les mécanismes de collaboration pour créer une collaboration réussie dans l'industrie des produits forestiers. La deuxième partie de cette recherche examine les aspects commerciaux critiques de l'établissement d'une cour de triage forestier pour faciliter l'accès à la ressource tout en supportant une collaboration entre les parties prenantes. Pour atteindre cet objectif, le modèle d'affaires proposé par Osterwalder et Pigneur (2010) est appliqué pour développer des modèles spécifiques au cas d'une cour de triage. Deux canevas de modèles d'affaires différents sont donc proposés et peuvent être applicables aux entreprises pour analyser l'aspect commercial de la mise en œuvre d'une cour de triage forestier. Une analyse de planification stratégique (FFOM) est développée pour répertorier les forces, les faiblesses, les opportunités et les menaces associées à ces modèles.International competition, particularly in the forest products industry, is constantly increasing and it forces companies to produce innovative products, satisfy more customized demand, and develop new markets. Nevertheless, firms do not necessarily have the ability to reach these achievements individually. Therefore, establishing a collaboration with other organizations could help them in solving problems, which may not be solved individually. In this research, the relevance of collaboration in the forest products industry as well as possible business models to establish a log sort yard are investigated with the aim of giving potential investors, business analysts, and managers a detailed overview of the key business aspects of these business units. The first part of this research investigates the main drivers, benefits, facilitators, and challenges of collaborations in the forest products industry, in order to help firms identify the best-fit collaborative mechanisms for their particular collaborative initiative. A conceptual framework is elaborated to identify the various components affecting the collaboration mechanisms needed to create successful collaboration in the forest products industry. The second part of this research investigates the critical business aspects of establishing a log sort yard related to its specific goal and support collaboration. To reach this goal, the Business Model Canvas proposed by Osterwalder and Pigneur (2010) is applied to develop possible business models for log sort yards. Two different business models are proposed which may be useful to firms to analyze the business aspect of implementing a log sort yard. A strategic planning analysis (SWOT analysis) is developed to list the strengths, weaknesses, opportunities and threats associated with these models