Unique among the intracellular lipid binding proteins, acyl CoA binding protein (ACBP) exclusively binds long chain fatty acyl CoAs (LCFA-CoAs). To test if ACBP is an essential protein in mammals, the ACBP gene was ablated by homologous recombination in mice. While ACBP heterozygotes appeared phenotypically normal, intercrossing of the heterozygotes did not result in any live homozygous deficient (null) ACBP^(-/-) pups. Heterozygous and wild type embryos were detected at all postimplantation stages, but no homozygous ACBP null embryos were obtained– suggesting that an embryonic lethality occurred at a preimplantation stage of development, or that embryos never formed. While ACBP null embryos were not detected at any blastocyst stage, ACBP null embryos were detected at the morula (8- cell), cleavage (2-cell), and zygote (1-cell) preimplantation stages. Two other LCFACoA binding proteins, sterol carrier protein-2 (SCP-2) and sterol carrier protein-x (SCPx) were significantly upregulated at these stages. These findings demonstrate for the first time that ACBP is an essential protein required for embryonic development and its loss of function may be initially compensated by concomitant upregulation of two other LCFA-CoA binding proteins only at the earliest preimplantation stages. The fact that ACBP is the first known intracellular lipid binding protein whose deletion results in embryonic lethality suggests its vital importance in mammals.

WNT ligands are secreted proteins that act as signals between cells. WNTs activate several interconnected signaling pathways that are required for embryonic development as well as tissue homeostasis in adults. The X-chromosomal Porcn gene encodes a membrane-bound O-acyl transferase that is required for the acylation of all 19 WNT ligands encoded in the mammalian genome. Non-acylated WNTs fail to be secreted from the producing cell and thus do not activate downstream signaling targets. In my thesis research, I have investigated the function of Porcn in mouse embryonic development. In vitro, I have shown that Porcn is required for canonical WNT signaling in ES cells and further, for their differentiation into endodermal and mesodermal derivatives. Taking advantage of a mouse line carrying a conditional (floxed) Porcn allele that I have generated, I have focused my studies on the early embryonic roles of Porcn using Cre recombinase-mediated and X chromosome inactivation-based ablation of Porcn function in vivo. I have found that the earliest requirement for Porcn in mouse development is the induction of gastrulation. In contrast to findings from in vitro studies, I have provided evidence that Porcn is not required for pre-implantation development in vivo. Dissecting embryonic and extra- embryonic roles of Porcn, I have been able to show that Porcn is required in the extra-embryonic chorion in order to mediate chorio-allantoic fusion, whereas ablation in the extra-embryonic visceral endoderm had no apparent effects. The extra-embryonic requirement for Porcn results in a parent-of-origin effect in Porcn heterozygous females due to X chromosome inactivation. In contrast to the placentation defect causing embryonic lethality of maternal allele mutants, deletion of the paternal allele caused variable fetal defects resulting in perinatal lethality with only rare survivors to adulthood. Both fetuses and adults represent a mouse model for Focal Dermal Hypoplasia (FDH), the syndrome caused by mutations in the human PORCN gene. My studies highlight the importance of PORCN-mediated WNT signaling for gastrulation, placentation, and fetal development, but suggest that endogenous WNT secretion does not play an essential role in either implantation or blastocyst lineage specification.

WNT ligands are secreted proteins that act as signals between cells. WNTs activate several interconnected signaling pathways that are required for embryonic development as well as tissue homeostasis in adults. The X-chromosomal Porcn gene encodes a membrane-bound O-acyl transferase that is required for the acylation of all 19 WNT ligands encoded in the mammalian genome. Non-acylated WNTs fail to be secreted from the producing cell and thus do not activate downstream signaling targets. In my thesis research, I have investigated the function of Porcn in mouse embryonic development. In vitro, I have shown that Porcn is required for canonical WNT signaling in ES cells and further, for their differentiation into endodermal and mesodermal derivatives. Taking advantage of a mouse line carrying a conditional (floxed) Porcn allele that I have generated, I have focused my studies on the early embryonic roles of Porcn using Cre recombinase-mediated and X chromosome inactivation-based ablation of Porcn function in vivo. I have found that the earliest requirement for Porcn in mouse development is the induction of gastrulation. In contrast to findings from in vitro studies, I have provided evidence that Porcn is not required for pre-implantation development in vivo. Dissecting embryonic and extra- embryonic roles of Porcn, I have been able to show that Porcn is required in the extra-embryonic chorion in order to mediate chorio-allantoic fusion, whereas ablation in the extra-embryonic visceral endoderm had no apparent effects. The extra-embryonic requirement for Porcn results in a parent-of-origin effect in Porcn heterozygous females due to X chromosome inactivation. In contrast to the placentation defect causing embryonic lethality of maternal allele mutants, deletion of the paternal allele caused variable fetal defects resulting in perinatal lethality with only rare survivors to adulthood. Both fetuses and adults represent a mouse model for Focal Dermal Hypoplasia (FDH), the syndrome caused by mutations in the human PORCN gene. My studies highlight the importance of PORCN-mediated WNT signaling for gastrulation, placentation, and fetal development, but suggest that endogenous WNT secretion does not play an essential role in either implantation or blastocyst lineage specification.

Preimplantation embryo development in the mouse is a time of rapid cellular morphological and molecular changes leading to embryo implantation for the generation of offspring. The Mager lab studies these events occuring between fertilization and implantation in order to better understand the initial events which set the stage for all future aspects of development. The result of this research impacts many scientific disciplines including in-vitro based means of embryo culture, establishment of epigenetic marks, differentiation and cellular reprogramming and can be used in translational research for the improvement of in-vitro culture techniques and develop novel therapies such as cell replacement in the case of macular degeneration (Bin, L., 2009). Through the use of in-vitro embryo culture, RNA interference (RNAi) approaches and daily observations, gene function required in preimplantation embryo development can be determined. In the initial published body of work evaluating gene knockdown using our RNAi approach (Maserati M 2011), WDR74 was characterized in preimplantation embryo development. We now understand that WDR74 is implicated in RNA production and/or stability as gene knockdown at the 1 cell stage significantly depletes mRNA within the embryo by the morula stage. Furthermore, double knockdown of Trp53 and Wdr74 results in a partial rescue of blastocyst formation suggesting p53 mediated apoptosis in the failure to make a blastocyst phenotype. The initial characterization of 4 RNA processing genes (SF3b14, SF3b1/SAP155, Rpl7l1 and Rrp7a) required for blastocyst formation was later evaluated. The results of this work has been submitted for publication and will be published soon in the journal Zygote. SF3b14 and SF3b1, identified as being part of the splicesome complex, disproportionally contributes to gene transcription of those genes containing more than 1 exon verifying a role in RNA splicing. Rpl7l1, identified by GO terms as a possible ribosomal gene, was found to be present in the cytoplasm and, surprisingly, in the nucleus. It is surmised this gene influences polymerase 2 activity as Rpl7l1 gene knockdown embryos demonstrate reduced active polymerase 2 activity at the morula stage. Rrp7a was identified as being critical in blastocyst formation and is present in the cytoplasm while excluded from the nucleus. Based on location and GO terms, this suggests a role in translation. Taken together, these 4 genes act in 3 different ways impacting RNA production, splicing or translation promoting blastocyst formation in the mouse. The final gene evaluated in this work was Bcl-6 corepressor (Bcor). As opposed to our previous work with RNA processing factors, this gene knockdown does not result in a failure to make a blastocyst. Bcor knockdown increases the rate of physiologically normal blastocysts in both murine and bovine models. Although further characterization must be done, temporary Bcor gene knockdown might be a useful improvement of in-vitro embryo culture systems including murine, bovine, equine and possibly even human. This manuscript is divided into 4 chapters, the first of which is a review of preimplantation embryo development. This covers selected and relevant events between fertilization and just before implantation of the embryo into the uterus. I mainly focus on events after fertilization and the necessary changes required for zygotic genome transcription and lineage specification. The second chapter characterizes WDR74, a gene we identified as critical in the formation of a blastocyst in a reverse genetic screen. As state before, we assess WDR74 function with the developing embryo and conclude the protein plays a role in RNA production and/or stability of RNA transcripts. We also test to rescue blastocyst formation in WDR74 knockdown embryos in an attempt to further evaluate WDR74 function. We continue the characterization of genes whose temporary reduction causes the failure of blastocyst formation in the third chapter. Here we report on four additional RNA processing genes in a body of work which has been published in the journal Zygote. Since these genes contained similar GO terms, we assumed they may all function in a similar way so they were assayed together as a group. As function of these genes were unknown, we determined protein localization within the cell, function in RNA splicing, alternative splicing and to determine if the failure to make a blastocyst is due to lineage specification. In the final chapter, BCOR gene expression is characterized in preimplantation embryo development as in the former 2 chapters. However, the result of this gene knockdown does not lead to the failure to make a blastocyst, rather this improves the number of blastocysts formed during the correct physiological time; the same time that blastocysts form invivo. Undoubtedly, this could lead to possible commercial applications which are reviewed along with the preliminary data we have been able to collect thus far. Specifically, the continuation of the BCOR gene knockdown research in preimplantation embryo development is pitched in the form of academic and international business collaboration with InvitroBrasil for the production of cloned bovine, equine and ICSI in equine.

A gestação inicial da égua é um período fascinante que abrange numerosas e intensas mudanças em seu desenvolvimento, muitas das quais são únicas para a espécie equina. Esse desenvolvimento depende da manutenção da função lútea, do estabelecimento de um ambiente uterino e de uma interação precisa e orquestrada entre o concepto e o ambiente uterino. O objetivo deste estudo foi verificar as alterações histológicas do endométrio e a produção histotrófica em éguas cíclicas e prenhes nos dias 7, 10 e 13 pós-ovulação. No primeiro ciclo, biópsias endometriais de 30 éguas foram coletadas no dia 7 (n = 10), 10 (n = 10) e 13 (n = 10) constituindo o grupo éguas cíclicas. No segundo ciclo, as mesmas éguas foram cobertas por um garanhão fértil, acompanhadas diariamente até detectar a ovulação, considerada o dia 0. Foram coletadas biópsias endometriais nos dias7 (n 10), 10 (n 10) e 13 (n 10). Imediatamente após a coleta, o útero foi lavado e as éguas em que foi obtido embrião, foram inseridas no grupo de éguas prenhes. Um maior calibre dos vasos sanguíneos foi observado em prenhez comparados às éguas cíclicas do dia 7 aos 13. No sétimo dia pós-ovulação, uma grande perda de células ciliadas foi evidente no grupo de éguas prenhes, comparadas ao grupo de éguas cíclicas, as células do epitélio endometrial estavam mais protusas e uma pequena quantidade de secreção histotrófica entre as dobras endometriais foi observada. No décimo dia de prenhez, secreção histotrófica glandular e do epitélio luminal estavam mais presentes comparadas às éguas do grupo cíclico. No dia 13 de prenhes, foi observado um grande conteúdo de histotrofo nas aberturas glandulares que estavam cercadas por células ciliares. Ocorreram alterações no ambiente uterino logo após a entrada do embrião no útero. No estroma e no lúmen, essas modificações parecem visar fornecer a nutrição necessária para o desenvolvimento inicial do embrião e estas mudanças nas estruturas celulares irão interagir na sinalização embrionária, futura fixação, implantação e placentação. / The early pregnancy of mare is a fascinating period that encompasses numerous and intense changes in its development, many of which are unique to the equine species. This development depends on the maintenance of the luteal function, the establishment of a favorable uterine environment and a precise and orchestrated interaction between the concept and the uterine environment. The aim of this study was to evaluate histological changes in the endometrium in days 7, 10 and 13 post-ovulation in pregnant and cyclic mares. In the first cycle, endometrial biopsies from 30 cyclic mares (Cyclic group) were collected on days 7, 10 and 13 post-ovulation. In the second cycle, the same mares were bred by a fertile stallion. At days 7, 10 and 13 post-ovulation intrauterine biopsies were collected. Immediately after sample collection, the mare‟s uteri were flushed, and those mares with embryo recovery were assigned to the Pregnant group. A larger blood vessel caliber was observed in pregnant mares than in cyclic from day 7 to 13. On the 7th day a large loss of ciliated cells was evident in the group of pregnant mares in comparison with the Cyclic group and the superficial cells of the endometrium were more protruded, and a small amount of histotrophic material between the folds was observed. On the 10th day of pregnancy, the glandular histotrophic secretion and the secretion of luminal epithelium became more intense than the secretion of cyclic mares. On the 13th day of pregnancy, a very large amount of histotroph was observed within large glandular openings surrounded by ciliated cells. Changes occurred in the uterine environment thereupon the entry of the embryo into the uterus. In the stroma and in the lumen, these modifications seem aim to provide the necessary nutrition for the initial development of the embryo and to promote changes at cellular structures that will interact in the embryonic signaling and future fixation, implantation and placentation.

A gestação inicial da égua é um período fascinante que abrange numerosas e intensas mudanças em seu desenvolvimento, muitas das quais são únicas para a espécie equina. Esse desenvolvimento depende da manutenção da função lútea, do estabelecimento de um ambiente uterino e de uma interação precisa e orquestrada entre o concepto e o ambiente uterino. O objetivo deste estudo foi verificar as alterações histológicas do endométrio e a produção histotrófica em éguas cíclicas e prenhes nos dias 7, 10 e 13 pós-ovulação. No primeiro ciclo, biópsias endometriais de 30 éguas foram coletadas no dia 7 (n = 10), 10 (n = 10) e 13 (n = 10) constituindo o grupo éguas cíclicas. No segundo ciclo, as mesmas éguas foram cobertas por um garanhão fértil, acompanhadas diariamente até detectar a ovulação, considerada o dia 0. Foram coletadas biópsias endometriais nos dias7 (n 10), 10 (n 10) e 13 (n 10). Imediatamente após a coleta, o útero foi lavado e as éguas em que foi obtido embrião, foram inseridas no grupo de éguas prenhes. Um maior calibre dos vasos sanguíneos foi observado em prenhez comparados às éguas cíclicas do dia 7 aos 13. No sétimo dia pós-ovulação, uma grande perda de células ciliadas foi evidente no grupo de éguas prenhes, comparadas ao grupo de éguas cíclicas, as células do epitélio endometrial estavam mais protusas e uma pequena quantidade de secreção histotrófica entre as dobras endometriais foi observada. No décimo dia de prenhez, secreção histotrófica glandular e do epitélio luminal estavam mais presentes comparadas às éguas do grupo cíclico. No dia 13 de prenhes, foi observado um grande conteúdo de histotrofo nas aberturas glandulares que estavam cercadas por células ciliares. Ocorreram alterações no ambiente uterino logo após a entrada do embrião no útero. No estroma e no lúmen, essas modificações parecem visar fornecer a nutrição necessária para o desenvolvimento inicial do embrião e estas mudanças nas estruturas celulares irão interagir na sinalização embrionária, futura fixação, implantação e placentação. / The early pregnancy of mare is a fascinating period that encompasses numerous and intense changes in its development, many of which are unique to the equine species. This development depends on the maintenance of the luteal function, the establishment of a favorable uterine environment and a precise and orchestrated interaction between the concept and the uterine environment. The aim of this study was to evaluate histological changes in the endometrium in days 7, 10 and 13 post-ovulation in pregnant and cyclic mares. In the first cycle, endometrial biopsies from 30 cyclic mares (Cyclic group) were collected on days 7, 10 and 13 post-ovulation. In the second cycle, the same mares were bred by a fertile stallion. At days 7, 10 and 13 post-ovulation intrauterine biopsies were collected. Immediately after sample collection, the mare‟s uteri were flushed, and those mares with embryo recovery were assigned to the Pregnant group. A larger blood vessel caliber was observed in pregnant mares than in cyclic from day 7 to 13. On the 7th day a large loss of ciliated cells was evident in the group of pregnant mares in comparison with the Cyclic group and the superficial cells of the endometrium were more protruded, and a small amount of histotrophic material between the folds was observed. On the 10th day of pregnancy, the glandular histotrophic secretion and the secretion of luminal epithelium became more intense than the secretion of cyclic mares. On the 13th day of pregnancy, a very large amount of histotroph was observed within large glandular openings surrounded by ciliated cells. Changes occurred in the uterine environment thereupon the entry of the embryo into the uterus. In the stroma and in the lumen, these modifications seem aim to provide the necessary nutrition for the initial development of the embryo and to promote changes at cellular structures that will interact in the embryonic signaling and future fixation, implantation and placentation.

Cochlear implantation is a proven and accepted option for young children with profound hearing loss. Cochlear implantation requires a professional team which should inform, guide, support and collaborate with parents constantly throughout the process. Existing programs for children with hearing loss and their families are generally designed on the basis of what experts believe they should contain, rather than on what parents actually wish to receive, which may often lead to a mismatch between the professionals view and the parents’ views of parental needs. In order to ensure that parental needs are appropriately addressed it is imperative that professionals investigate and understand the individual needs and desires of the parents with whom they collaborate. This can be managed by carefully tailoring information to their individual needs and presenting information in an accessible format at the time it is most appropriate and digestible. The aim of this research study was to develop a pre-implantation tool to rate the individual support and information needs of parents of young cochlear implant candidates. Within the context of applied research, a qualitative descriptive intervention research design was used in the study. Ten parents of children with cochlear implants participated in a semi-structured interview to investigate their need for information and support during the pre-implantation phase of cochlear implantation. Their responses were analysed and compared to relevant literature in order to develop the pre-implantation rating tool for parents of cochlear implant candidates. The rating tool consists of ten areas for information and support. These areas are as follows: general, technical, surgery, social support, financial, communication options, education, outcomes, rehabilitation and parental role. Parents are able to rate which areas of information and support is important to them and what they would like to discuss with the professional involved. Parents are also encouraged to identify any area of information and support that is not included in the rating tool that they would want information on from the cochlear implant team. This rating tool was evaluated by eight speech-language pathologist/audiologist working in six cochlear implant programmes in South Africa to determine the value of the rating tool. Positive responses were given about the adaptability of the tool to identify individual needs for support and information and the tool would be useful to guide speech-language pathologist/audiologists to identify needs of parents that should initially be addressed. Respondents agreed that the rating tool provides an opportunity to express parent’s individual needs for information and support; that the tool correlates with a family centered approach and would be useful to include in cochlear implant programs. The majority of participants felt the rating tool possibly will be effective in identifying information and support needs of parents before cochlear implantation and respondents would be willing to implement the rating tool in their cochlear implant programme. The positive response from professionals working in the field of cochlear implantation validates the effectiveness of the rating tool. AFRIKAANS : Kogleêre inplantering is ‘n beproefde en aanvaarde opsie vir jong kinders met ‘n uitermatige gehoorverlies. Kogleêre inplanting vereis dat ‘n professionele span ouers deur die proses inlig, lei en ondersteun. Huidige programme vir kinders met gehoorverlies en hulle gesinne, is oor die algemeen gebaseer op grond van inligting wat volgens kundiges belangrik is om in te sluit. Hierdie programme is nie noodwendig gebaseer op inligting ouers graag wil ontvang nie. Dit kan lei tot ‘n verskil tussen die perspektief van professionele persone teenoor die van die ouer oor ouer- behoeftes aan inligting. Om te verseker dat ouers se behoeftes effektief aangespreek word, is dit noodsaaklik om dit te ondersoek en die individuele behoeftes van ouers te verstaan. Dit kan gedoen word deur inligting aan te pas volgens die individuele behoeftes van ouers en die inligting te verskaf in ‘n toeganklike wyse op ‘n gepaste tyd wanneer dit geskik is en die ouer die inligting kan prosesseer. Die doel van hierdie navorsing studie was om ‘n pre-inplantering instrument te ontwikkel om die individuele behoeftes aan inligting en ondersteuning van ouers van jong kogleêre inplantings kandidate te bepaal. Binne die konteks van toegepaste navorsing is ‘n kwalitatiewe beskrywings intervensie navorsingsontwerp gebruik. Tien ouers van kinders met kogleêre inplantings het deelgeneem aan ‘n semi-gestruktureerde onderhoud. Die onderhoud het ouers se behoefte aan inligting en ondersteuning tydens die pre-inplanterings fase van kogleêre inplantasie ondersoek. Die resultate is geanaliseer en vergelyk met relevante literatuur om sodoende die pre-inplantering bepaling instrument vir ouers van kogleêre kandidate te ontwerp. Die instrument bestaan uit tien areas van inligting en ondersteuning. Hierdie areas is as volg: algemeen, tegnies, chirurgie, sosiale ondersteuning, finansieel, kommunikasie opsies, onderrig, rehabilitasie en ouer rol. Ouers kan bepaal watter areas van inligting en ondersteuning vir hulle belangrik is en wat hulle graag wil bespreek met die professionele persone betrokke by die kogleêre inplanting proses. Ouers word ook aangemoedig om enige area van inligting en ondersteuning te identifiseer wat moontlik nie ingesluit is in die instrument nie, maar wat hulle graag met die kogleêre span wil bespreek. Agt spraak-taal patoloë/oudioloë van ses kogleêre inplantings programme in Suid-Afrika het die instrument geëvalueer om die waarde daarvan te bepaal. Positiewe insette is gegee oor die aanpasbaarheid van die instrument om die individuele behoeftes vir inligting en ondersteuning te bepaal; dat die instrument betekenisvol is om die spraak-taal patoloog/oudioloog te lei om die behoeftes van ouers te identifiseer en aan te spreek; dat die instrument ooreenstem met ‘n familie- gesentreede benadering en dat die instrument effektief ingesluit kan word in kogleêre inplantings programme. Die meeste deelnemers het aangedui dat die instrument effektief sal wees in die identifisering van inligting en ondersteunings behoeftes van ouers voor ‘n kogleêre inplanting. Deelnemers het aangedui dat hulle bereid sal wees om die instrument te implementeer in hulle kogleêre inplantingsprogram. Die positiewe respons van spraak-taal patoloë en oudioloë dui op die geldigheid en effektiwiteit van die instrument. / Dissertation (MCommunication Pathology)--University of Pretoria, 2010. / Speech-Language Pathology and Audiology / unrestricted

With pre-implantation genetic diagnosis (PGD), genetic testing and selective transfer of embryos is possible. In the future, germ-line gene therapy (GLGT) applied to embryos before implantation, in order to introduce missing genes or replace mutant ones, may be possible. The objective of this dissertation is to analyse moral aspects of these technologies, as described by eighteen British, Italian and Swedish gynaecologists and geneticists. The objective is systematised into three parts: research interviews and qualitative analysis, philosophical analysis, and elaboration of a framework that supports the combination of analytic methods. PGD was described as positive since it enabled some couples at risk for a genetic disease to have a child without the disease. PGD was described as in different senses ‘better’ than methods for prenatal diagnosis and selective termination of pregnancy. It was also described as positive since it provided couples at risk with one more option, even if it did not result in the birth of a healthy child. However, interviewees were concerned about the difficulty of defining and evaluating genetic disease. They were also concerned about patients’ choices, and about exaggerated use or misuse. Whereas PGD gave rise to ambivalence in terms of how to understand, describe and evaluate it, GLGT was often described as unrealistic or undesirable. The results of the qualitative analysis are used in a philosophical analysis of the concepts of choice, autonomous choice, ambivalence, trust and ambivalence in trust relations. A set of distinct characteristics of each concept are elaborated. The results of the philosophical analysis are used in the discussion of the results of the qualitative analysis. The study shows that the technologies imply both ‘new’ ways to perform ‘old’ medical practices and ‘new’ practices. Old moral questions are reformulated. New moral questions are added. Against the background of this, the concept of genetic identity is discussed. Key words: empirical ethics, pre-implantation genetic diagnosis, germ-line gene therapy, qualitative research, philosophical analysis, medical progress, genetic disease, choice, autonomous choice, ambivalence, trust, genetic identity.

Präzise regulierte Genexpression, ist der Schlüssel zu erfolgreicher Embryonal-entwicklung. Die Expression von Zelltyp-spezifischen Transkriptionsfaktoren kann durch räumliche Interaktionen von Promotoren und Enhancern im Nukleus kontrolliert werden, aber auch durch 3D Faltung der DNA in größere organisatorische Einheiten wie “Topologically Associating Domains” (TADs) oder “A/B compartments”. Um die 3D Faltung in den Zelltypen des prä-implantations Embryos zu untersuchen, nutze ich ES und XEN Zellen, die stark dem Epiblast und dem primitiven Endoderm in der inneren Zellmasse des E4.5 Embryos ähneln. Um den Zusammenhang zwischen 3D DNA Faltung und zellulärer Identität zu erforschen, habe ich GAM, ATAC-seq und RNA-seq Daten von ES und XEN Zellen produziert. Um die Genom-Architektur im Embryo zu untersuchen, habe ich außerdem die GAM Methode an den Mausembryo angepasst und kann dadurch erstmals genomweit DNA-Faltung in den spezifischen Zelltypen der inneren Zellmasse des prä-implantations Embryos zeigen. ES und XEN Zellen zeigen viele differentiell exprimierte Gene, sowie starke Veränderungen in der Chromatin-Organisation, beispielweise in der Bildung von reprimierten Chromatinnetzwerken in ESCs, die wichtige XEN Gene wie Gata6 und Lama1 enthalten, während diese nicht aktiv sind. XEN-spezifische Genexpression ist oft mit der Präsenz von XEN-spezifischen “TAD boundaries” gekoppelt. Der Sox2 Locus zeigt eine ESC-spezifische Organisation mit aktiven Genen, und Regionen die von den Transkriptionsfaktoren SOX2, NANOG und OCT4 gebunden sind. Die starke Reorganisation der Genom-Architektur in wichtigen Loci wie Gata6 und Sox2 konnte ich mit in vivo GAM Daten bestätigen und finde ähnliche Unterschiede zwischen den beiden Zelltypen der inneren Zellmasse wie im in vitro Model. Diese Ergebnisse zeigen, wie wichtig es ist, Zelltypen getrennt zu untersuchen und, dass eine Verbindung zwischen zellulärer Identität und der Faltung des Genoms in der Embryonalentwicklung besteht. / Tightly controlled gene regulation is key to functional metazoan embryonic development. The expression of cell-fate determining transcription factors orchestrates the establishment of the various lineages of the embryo. Gene expression is often regulated via specific chromatin organisation. To investigate cell type-specific differences in chromatin folding in early embryonic development, I used in vitro models of the two distinct cell populations in the blastocyst ICM. In mouse ES and XEN cells, I mapped 3D genome conformation using Genome Architecture Mapping (GAM), chromatin accessibility using ATAC-seq, and gene expression using total RNA-seq. To enable the mapping of 3D genome folding directly in the blastocyst ICM, I adapted GAM for cell type-specific selection of nuclei, by integrating immunofluorescence detection of markers, and generated the first genome-wide chromatin contact maps that distinguish ICM cell types. I report that the ES and XEN cell lineages undergo abundant large scale rearrangements of genome architecture and exhibit high numbers of differentially expressed genes. For example, extra-embryonic endoderm genes, such as Lama1 and Gata6, form silent hubs in ESCs, potentially connecting maintenance of pluripotency to 3D structure of the genome. Further, I show that the expression of XEN cell-specific genes relates to the formation of XEN cell-specific TAD boundaries. Chromatin contacts at the Sox2 locus exhibit an ESC-specific organisation around binding of pluripotency transcription factors OCT4, NANOG and SOX2, into hubs of high gene activity. The observations detected in in vitro models, were investigated in smaller GAM datasets produced using the in vivo counterparts in the ICM. Overall, in vivo data confirmed the high degree of chromatin rearrangement among the two cell types, specifically in loci of lineage driving genes. The findings from in vivo data further underscore the connection of genome topology and cellular identity.