Podcasts about PAX6

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.07.12.548679v1?rss=1 Authors: Duot, M., Viel, R., Viet, J., Le Goff-Gaillard, C., Paillard, L., Lachke, S., Gautier-Courteille, C., Reboutier, D. Abstract: The ocular lens, along with the cornea, focuses light on the retina to generate sharp images. Opacification of the lens, or cataract, is the leading cause of blindness worldwide. Presently, the best approach for cataract treatment is to surgically remove the diseased lens and replace it with an artificial implant. Although effective, this is costly and can have post-surgical complications. Toward identifying alternate treatments, it is imperative to develop organoid models relevant for lens studies and anti-cataract drug screening. Here, we demonstrate that by culturing mouse lens epithelial cells under defined 3-dimensional (3D) culture conditions, it is possible to generate organoids that display optical properties and recapitulate many aspects of lens organization at the tissue, cellular and transcriptomic levels. These 3D cultured lens organoids can be rapidly produced in large amounts. High-throughput RNA-sequencing (RNA-seq) on specific organoid regions isolated by laser capture microdissection (LCM) and immunofluorescence assays demonstrate that these lens organoids display spatiotemporal expression of key lens genes, e.g., Jag1, Pax6, Prox1, Hsf4 and Cryab. Further, these lens organoids are amenable to induction of opacities. Finally, knockdown of a cataract-linked RNA-binding protein encoding gene, Celf1, induces opacities in these organoids, indicating their use in rapidly screening for genes functionally relevant to lens biology and cataract. In sum, this lens organoid model represents a compelling new tool to advance the understanding of lens biology and pathology, and can find future use in the rapid screening of compounds aimed at preventing and/or treating cataract. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.09.18.508425v1?rss=1 Authors: Vogel, J. W., Alexander-Bloch, A., Wagstyl, K., Bertolero, M., Markello, R., Pines, A., Sydnor, V. J., Diaz-Papkovich, A., Hansen, J., Evans, A. C., Bernhardt, B., Misic, B., Satterthwaite, T., Seidlitz, J. Abstract: Cortical arealization arises during neurodevelopment from the confluence of molecular gradients representing patterned expression of morphogens and transcription factors. However, how these gradients relate to adult brain function, and whether they are maintained in the adult brain, remains unknown. Here we uncover three axes of topographic variation in gene expression in the adult human brain that specifically capture previously identified rostral-caudal, dorsal-ventral and medial-lateral axes of early developmental patterning. The interaction of these spatiomolecular gradients i) accurately predicts the location of unseen brain tissue samples, ii) delineates known functional territories, and iii) explains the topographical variation of diverse cortical features. The spatiomolecular gradients are distinct from canonical cortical functional hierarchies differentiating primary sensory cortex from association cortex, but radiate in parallel with the axes traversed by local field potentials along the cortex. We replicate all three molecular gradients in three independent human datasets as well as two non-human primate datasets, and find that each gradient shows a distinct developmental trajectory across the lifespan. The gradients are composed of several well known morphogens (e.g., PAX6 and SIX3), and a small set of genes shared across gradients are strongly enriched for multiple diseases. Together, these results provide insight into the developmental sculpting of functionally distinct brain regions, governed by three robust transcriptomic axes embedded within brain parenchyma. Copy rights belong to original authors. Visit the link for more info Podcast created by PaperPlayer

Dr Kat Arney takes a deep dive into our genetic make-up and tells the story of four pieces of human DNA: the fat gene, the Huntington gene, the CCR5 gene associated with HIV resistance, and PAX6, the eyeball gene.

REFERENCED STUDIES FOR ANDROGENETIC ALOPECIA Silva et al (starts at 1:10). Randomized clinical trial of low-dose oral minoxidil for the treatment of female pattern hair Loss: 0.25 mg versus 1 mg. Journal of the American Academy of Dermatology;  Online Jan 2022. Therianou et al (starts at 3;18). How Safe Is Prescribing Oral Minoxidil in Patients Allergic to Topical Minoxidil? Journal of the American Academy of Dermatology. J Am Acad Dermatol Feb 2022. James JF et al. (starts at 4:51) Efficacy and safety profile of oral spironolactone use for androgenic alopecia: A systematic review. J Am Acad Dermatol. Feb 2022 Plante et al (starts at 8:05). The Need for Potassium Monitoring in Women on Spironolactone for Dermatologic Conditions. J Am Acad Dermatol. 2022 Jan 21 Online;S0190-9622(22)00081-0. Özcan D (starts at 10:59). Pediatric androgenetic alopecia: a retrospective review of clinical  characteristics, hormonal assays and   metabolic syndrome risk factors in 23 patients An Bras Dermatol. 2022 Jan 12; online Zong et al (starts at 14:40). Prevalence of ocular anomalies is increased in women with polycystic ovary syndrome-exploration of association with PAX6 genotype. Ophthalmic Genetics. 2022 Jan 11;1-4. Online Deng et al (starts at 15:51). Androgen receptor mediated paracrine signaling induces regression of blood vessels in the dermal papilla in androgenetic alopecia. J Invest Dermatol. Jan 2022 online Li K et al (starts at 18:48). Association of fibrosis in the bulge portion with hair follicle miniaturization in androgenetic alopecia. J Am Acad Dermatol 2022; Jan;86(1):213-215.   REFERENCED STUDIES FOR ALOPECIA AREATA Di Fillipo et al (starts at 22:14). Efficacy of 308-nm excimer therapy in alopecia areata: a retrospective study with long-term follow-up Photodermatol Photoimmunol Photomed 2022 Jan 21.  online. Ting H-C et al (starts at 25:30). Association between alopecia areata and retinal diseases: A nationwide population-based cohort study. J Am Acad Dermatol  2021 Nov 1; online Abdelkader HA et al (starts at 26:24). Histone deacetylase 1 in patients with alopecia areata and acne vulgaris: An epigenetic alteration. Australas J Dermatol. 2022 Jan 25. Online.

Welcome to a new episode of EyePod Bayer! In this special episode, you will hear more about congenital aniridia, a rare but devastating pan-ocular disease resulting from mutations in the PAX6 gene. To bring together the various existing efforts to understand, manage, and treat aniridia, ANIRIDIA-NET was established. Stay tuned to hear more about it! Appearing in this podcast: Prof. Neil Lagali Prof. Dominique Bremond-Gignac Juliana Martinez Ivana Kildsgaard Questions? eyepod@bayer.com MA-M_AFL-DK-0071-2    

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.03.366914v1?rss=1 Authors: Ypsilanti, A. R., Pattabiraman, K., Catta-Preta, R., Golonzhka, O., Lindtner, S., Tang, K., Jones, I., Abnousi, A., Juric, I., Hu, M., Shen, Y., Dickel, D. E., Visel, A., Pennachio, L. A., Hawrylycz, M., Thompson, C., Zeng, H., Barozzi, I., Nord, A. S., Rubenstein, J. L. R. Abstract: We uncovered a transcription factor (TF) network that regulates cortical regional patterning. Screening the expression of hundreds of TFs in the developing mouse cortex identified 38 TFs that are expressed in gradients in the ventricular zone (VZ). We tested whether their cortical expression was altered in mutant mice with known patterning defects (Emx2, Nr2f1 and Pax6), which enabled us to define a cortical regionalization TF network (CRTFN). To identify genomic programming underlying this network, we performed TF ChIP-seq and chromatin-looping conformation to identify enhancer-gene interactions. To map enhancers involved in regional patterning of cortical progenitors, we performed assays for epigenomic marks and DNA accessibility in VZ cells purified from wild-type and patterning mutant mice. This integrated approach has identified a CRTFN and VZ enhancers involved in cortical regional patterning. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.05.18.102855v1?rss=1 Authors: Wu, J. P. H., Yeung, J., Wu, S.-R., Zoghbi, H. Y., Goldowitz, D. Abstract: Pou3f1 is a transcription factor involved in early neural differentiation. Cap Analysis Gene Expression (5'-CAGE) analysis reveals that Pou3f1 transcript is highly enriched in the developing cerebellum. Between embryonic (E) days E10.5 and E12.5, Pou3f1 expression is present prominently along the subpial stream (SS), suggesting that Pou3f1+ cells are glutamatergic cerebellar nuclear (CN) neurons. This finding was confirmed by immunofluorescent (IF) co-labeling of Pou3f1 and Atoh1, the master regulator of cells from the rhombic lip (RL) that are destined for neurons of the glutamatergic lineage, as well as in Atoh1-null tissues, in which Pou3f1 expression is absent. Interestingly, the expression of Pax6, another key molecule for CN neuron survival, does not co-localize with that of Pou3f1. In the Pax6-null Small Eye (Sey) mutant, which is characterized by a loss of many glutamatergic CN neurons, Pou3f1+ CN neurons are still present. Furthermore, Pou3f1-labeled cells do not co-express Tbr1, a well-established marker of glutamatergic CN neurons. These results highlight that Pou3f1+ cells are a distinct and previously unrecognized subtype of glutamatergic CN neurons that do not have the ''canonical'' sequence of Atoh1[->]Pax6[->]Tbr1 expressions. Instead, they express Atoh1, Pou3f1, and other markers of CN neurons, Brn2 and Irx3. These findings illustrate that glutamatergic CN neurons that arise from the RL are composed of molecularly heterogeneous subpopulations that are determined by at least two distinct transcriptional programs. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.04.24.059428v1?rss=1 Authors: Core, N., Erni, A., Mellon, P. L., Hoffmann, H. M., Beclin, C., Cremer, H. Abstract: Several subtypes of interneurons destined for the olfactory bulb are continuously generated after birth by neural stem cells located in the ventricular-subventricular zones of the lateral ventricles. Future neuronal identity depends on the positioning of pre-determined neural stem cells along the ventricle walls, which, in turn, depends on delimited expression domains of transcription factors and their cross regulatory interactions. However, mechanisms underlying positional identity of neural stem cells are still poorly understood. Here we show that the transcription factor Vax1 controls the production of two specific neuronal sub-types. First, it is directly necessary to generate Calbindin expressing interneurons from ventro-lateral progenitors. Second, it represses the generation of dopaminergic neurons by dorso-lateral progenitors through inhibiting Pax6 expression in the dorso-lateral wall. We provide evidence that this repression occurs via activation of microRNA miR-7, targeting Pax6 mRNA. Copy rights belong to original authors. Visit the link for more info

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Generation of the brain depends on proper regulation of progenitor proliferation and differentiation during development. Many such factors known to affect proliferation and differentiation are transcription factors. In particular, the transcription factor Pax6 has received much attention because of its potency to control various aspects of brain development. During development of the telencephalon Pax6 regulates patterning, cell proliferation and neurogenesis, but how Pax6 mediates and coordinates these diverse functions at the molecular level is not well understood. It has previously been demonstrated that the homeodomain of Pax6 plays a role in establishing the pallial-subpallial boundary. However it is not involved in other processes during telencephalic development as shown by the analysis of Pax64NEU mice, which are characterized by a point-mutation in the DNA-binding helix of the homeodomain. In order to gain more insights into the molecular network underlying the mild homeodomain function in the developing telencephalon, transcriptome analysis with Pax64NEU mice was performed. Almost no transcriptional changes were detected, suggesting that transcriptional regulation by the homeodomain of Pax6 has no major impact on forebrain development. Additionally, these results implied that the majority of effects exerted through Pax6 during telencephalic development are mediated by the bipartite paired-domain (PD). Therefore the main focus of this thesis was to examine the specific roles of the Pax6 paired-domain and its individual DNA-binding subdomains (PAI and RED) during forebrain development. The role of these DNA-binding domains was examined using mice with point-mutations in the PAI (Pax6Leca4, N50K) and RED (Pax6Leca2 R128C) subdomains and showed that the mutations in these subdomains exert opposing roles regulating proliferation in the developing cortex. While the mutated PAI domain resulted in reduced proliferation of both apical and basal progenitors, the mutated RED domain provoked increased proliferation. However, the PAI domain largely mediates the neurogenic function of Pax6. Additionally, genome-wide transcriptome analysis was able to unravel the key signatures mediated by the distinct domains. In summary, Pax6 exerts its key roles during forebrain development by use of distinct subdomains to regulate proliferation and differentiation. Thus Pax6 is able to coordinate and fine tune patterning, neurogenesis and proliferation in a simultaneous manner in different radial glial subpopulations. The transcriptional regulation through Pax6 may not only be restricted to protein coding genes, but may also include control of microRNA (miRNA) expression. Such small RNA molecules have recently been implicated in proliferation and differentiation during development, however expression and the role of single microRNAs is still poorly understood. Towards this end, miRNA expression profiling was performed using an embryonic stem cell differentiation system at different stages of neuronal differentiation in order to identify new miRNAs involved in radial glia specification and differentiation. This analysis revealed a number of microRNAs induced during differentiation from neural progenitors to neurons. Most strikingly only four miRNA candidates were found with exclusively high expression in progenitor cells. These data suggest that also Pax6 may play a role in transcriptional regulation beyond mRNAs.

Pax6 encodes a specific DNA-binding transcription factor that regulates the development of multiple organs, including the eye, brain and pancreas. Previous studies have shown that Pax6 regulates the entire process of ocular lens development. In the developing forebrain, Pax6 is expressed in ventricular zone precursor cells and in specific populations of neurons; absence of Pax6 results in disrupted cell proliferation and cell fate specification in telencephalon. In the pancreas, Pax6 is essential for the differentiation of α-, β- and δ-islet cells. To elucidate molecular roles of Pax6, chromatin immunoprecipitation experiments combined with high-density oligonucleotide array hybridizations (ChIP-chip) were performed using three distinct sources of chromatin (lens, forebrain and β-cells). ChIP-chip studies, performed as biological triplicates, identified a total of 5,260 promoters occupied by Pax6. 1,001 (133) of these promoter regions were shared between at least two (three) distinct chromatin sources, respectively. In lens chromatin, 2,335 promoters were bound by Pax6. RNA expression profiling from Pax6⁺/⁻ lenses combined with in vivo Pax6-binding data yielded 76 putative Pax6-direct targets, including the Gaa, Isl1, Kif1b, Mtmr2, Pcsk1n, and Snca genes. RNA and ChIP data were validated for all these genes. In lens cells, reporter assays established Kib1b and Snca as Pax6 activated and repressed genes, respectively. In situ hybridization revealed reduced expression of these genes in E14 cerebral cortex. Moreover, we examined differentially expressed transcripts between E9.5 wild type and Pax6⁻/⁻ lens placodes that suggested Efnb2, Fat4, Has2, Nav1, and Trpm3 as novel Pax6-direct targets. Collectively, the present studies, through the identification of Pax6-direct target genes, provide novel insights into the molecular mechanisms of Pax6 gene control during mouse embryonic development. In addition, the present data demonstrate that Pax6 interacts preferentially with promoter regions in a tissue-specific fashion. Nevertheless, nearly 20% of the regions identified are accessible to Pax6 in multiple tissues.

Insights into the developmental processes during which the brain forms from the neuroepithelium may provide a deeper understanding how the brain works. The Rho family of small GTPases is known for its many cell biological functions such as regulation of the cytoskeleton, gene expression, cell migration, adhesion, cell polarity and the cell cycle. All of these functions are of importance during the formation of the cerebral neocortex, which consists of the generation of its different cell types, their migration to their destination and their maturation to a functional network. These roles have been mostly established in vitro using dominant negative or constitutively active constructs. Since these approaches are often not entirely specific for single pathways, this work used the Cre/loxP system to genetically delete an individual member of the Rho family, RhoA, to examine its role following a loss-of-function approach. Specifically, we examined a mouse line where part of the RhoA gene has been deleted by means of the Emx1::Cre mouse line. This idea is based on previous experiences with the deletion of Cdc42 in the developing neocortex, which leads to a loss of apical progenitors. RhoA often works as a functional antagonist to Cdc42. Using immunofluorescence, we could detect a loss of RhoA at embryonic day 12 (E12) in Emx1::Cre-positive offspring carrying the floxed RhoA-construct in both alleles (cKO). At E14, we detected an increase in mitotic cells to 160% (±25%, p

Abstract Considering the globally increasing rate of incidence, Type 2 diabetes mellitus belongs to the most frequent endocrine metabolic disorders today. In addition to insulin resistance of peripheral tissues, this disease is the result of dysfunction of the endocrine pancreas and in particular of the functional failure of β-cells. The progress of therapeutical strategies is based on the research of the underlying mechanisms. The aim of the present dissertation was the analysis of new molecular regulators which might improve our underdstanding of the differentiation of pancreatic islets and the functional maintenance of adult β-cells. The first part of this work concerned the role of the transcription factor Pax6 and especially the role of its transactivation domain (TA) and of its two DNA binding domains, the paired domain (PD) and the homeodomain (HD), in differentiation of pancreatic endocrine cells. By analyzing four different mouse lines with specific mutations in one of these three domains, we found that the PD of Pax6 is essential for differentiation of glucagon producing α-cells. Inactivation of this domain resulted in a phenotype similar to that of Pax6 knockout mice (Pax6-/-) with a near complete absence of glucagon positive α-cells, a markedly reduced number of insulin producing β-cells, and a disorganized islet structure. Mutations of HD or TA showed a less severe pancreatic phenotype. Islets either exhibited no morphological changes or they showed a reduction of α- and β-cells. Intraperitoneal glucose tolerance tests demonstrated the utmost importance of the transcription factor Pax6 for maintenance of normal pancreatic endocrine function in adult animals. In the second part of this study we identified new genes and proteins, respectively, which could play a regulatory role in normal function of β-cells. In particular it was possible to show that Eny2, hitherto a protein only described in yeast or invertebrates like drosophila, is involved in the regulation of insulin secreting vertebrate cells. si-RNA mediated knockdown of Eny2 resulted in markedly increased glucose and incretin-induced insulin secretion. This could be at least in part attributed to a higher glucose-dependent cellular metabolism and an enhanced signal transduction via protein kinase A and is accompanied by elevated levels of intracellular calcium. Taken together, these results indicate that Eny2 functions as a negative regulator of glucose-stimulated and incretin-mediated insulin secretion, at least in vitro. However, a gap of knowledge still remains between the established nuclear functions of Eny2 and the cellular phenotype we observed upon its suppression. Nevertheless, the effects of an Eny2-knockdown are glucose dependent and additive to the incretin signaling. This feature makes this model attractive to obtain new insights in how insulin secretion of β- cells proceeds and how to find new therapeutical strategies to treat type 2 diabetes mellitus.

The vast majority of neurons in the murine brain are generated during embryonic neurogenesis. However, at least two neurogenic niches continue to produce specific types of neurons throughout life. The adult dentate gyrus harbours stem cells that generate dentate granule neurons and the subependymal zone produces distinct types of olfactory interneurons. The adult neurogenic subependymal zone is derived from the embryonic dorsal and ventral subventricular zone of the telencephalon, i. e. progenitor domains which generate both the ventral and dorsal glutamatergic and GABAergic neurons, respectively. While a cascade of transcription factors beginning with Pax6 governs the generation of glutamatergic cortical neurons, transcription factors of the Dlx family are crucial for the embryonic neurogenesis of GABAergic neurons. Notably, Pax6 and Dlx transcription factors factors are expressed in the adult subependymal zone. In this study I investigated the regionalization of the adult subependymal neurogenic niche in regard to Pax6 and Dlx and I examined the role of these factors in neuronal subtype specification. Consistent with their embryonic origin progenitors in the adult brain express Dlx1 and Dlx2 in the lateral, but not the dorsal subependymal zone. Using retroviral vectors I demonstrated that Dlx2 is necessary for neurogenesis of virtually all olfactory interneurons arising from the lateral subependymal zone. Beyond its function in generic neurogenesis, Dlx2 plays a crucial role in neuronal subtype specification in the adult olfactory bulb promoting specification of dopaminergic interneurons. Strikingly, Dlx2 requires interaction with Pax6, as Pax6 deletion blocks Dlx2 mediated neuronal specification. Of note, however, Pax6 protein is expressed in a gradient being especially abundant in dorsal regions of the adult subpenedymal zone. While playing obviously a role in the genesis of GABAergic interneurons, I also investigated whether the dorsal subependymal zone could give rise to glutamatergic neurons which have so far been overlooked. Surprisingly, progenitors located mainly in dorso-rostral regions of the subependymal zone express transcription factors previously linked to glutamatergic neurogenesis like Pax6 → Neurogenin2 → Tbr2 → Tbr1. These neurons migrate along the rostral migratory stream and integrate into the glomerular layer of the olfactory bulb. Finally, I provide evidence that these Tbr2-positive cells could become recruited following cortical lesions where callosal projection neurons are depleted.

The predominant precursor cell type during cortical neurogenesis are radial glia cells, which receive extrinsic and intrinsic signals that might influence cell proliferation and neurogenesis. These radial glia cells have direct contact to the growth factor rich basement membrane throughout cell division. However, it is not known, how the signals received from the basal cell attachment influence the behavior of radial glia cells in regard to the regulation of cell proliferation and neurogenesis. Therefore, I examined the lamininγ1 (LNγ1) mutant, lacking the contact of radial glial endfeet to the basement membrane, and the α6 integrin-/- with a disturbed assembly of the basement membrane. The analysis of the LNγ1 mutant and the α6 integrin-/- showed no defects in the radial glia progeny, cell proliferation or their orientation of cell division. Thus, these results strongly suggest that the direct contact of radial glia cells to the basement membrane is not required for these aspects. Radial glia cells of the dorsal telencephalon are also known to be specified by the expression of the transcription factor Pax6, which plays a pivotal role in the regulation of cell proliferation, neurogenesis and regionalisation during development of the telencephalon. In order to understand how Pax6 coordinates these diverse functions at the molecular level, the roles of the different DNA-binding domains of Pax6, the paired domain (PD), the splice variant of the paired domain (PD5a) and the homeodomain (HD) were analyzed in loss- and gain-of-function approaches. The analysis of the specific paired domain mutant Pax6Aey18-/-, that lacks large parts of the paired domain, but contains an intact homeodomain and transactivating domain (TAD), showed that the paired domain is required for the regulation of neurogenesis, cell proliferation and regionalisation in the developing telencephalon and eye. The homeodomain plays only a minor role during telencephalic development, in contrast to its function in the eye, as shown by the analysis of Pax64Neu-/- mice, which have a point mutation in the DNA-binding domain of the homeodomain, while paired domain and transactivating domain are still functional. Moreover retrovirus-mediated overexpression of Pax6 and Pax6(5a) in cortical cells showed that splicing of the paired domain regulates between a Pax6 form that affects neurogenesis, and cell proliferation, while the other Pax6 form, containing exon5a, regulates exclusively cell proliferation.

The molecular signals specifying neuronal, glial or multipotent precursors in the developing and adult central nervous system (CNS) are largely unknown. Radial glial cells have recently been discovered as major precursor population generating neurons or glial cells in separate lineages in the developing CNS. Towards this aim to identify the key molecular determinants for neurogenic versus gliogenic fate in radial glial cells I used a novel method to selectively enrich neurogenic or non-neurogenic radial glia and compared their gene expression to adult subependymal zone (SEZ) precursors in vitro and in vivo. The expression profile of the transcription factors Olig2 and Pax6 were particularly intriguing. Olig2 was 67 fold higher in multipotent compared to neuronal or glial precursors, while Pax6 showed strongest expression in neuronal precursors. I therefore focused further on the functional analysis of Pax6 and Olig2 in neural stem cell in vitro and in vivo. Interference with Olig2 in neural stem cells in vitro revealed its role in self-renewal. In adult neural stem cells of the SEZ overexpression of Olig2 promoted oligodendrocyte formation. Pax6, in contrast, proved to be a very potent neurogenic determinant since neurogenesis is not only Pax6-dependent in neural stem cells in vitro, but also in adult neural stem cell in vivo. Taken together, these results demonstrate a pathway combining transcription factors of dorsal and ventral regions that is activated in a specific lineage progression of adult neural stem cells in vitro and in vivo. Most importantly also in regard to therapeutic approaches, this work revealed the molecular mechanisms to direct adult neural stem cells towards a specific cell fate, neuronal or oligodendroglial.

The cells of the mammalian central nervous system (CNS) arise from multipotential precursor cells. The mechanisms that drive precursor cells toward a distinct cell fate are not well understood. Since transcription factors are known to control fate decisions, I attempted to determine the role of transcription factors Emx1, Emx2 and Pax6 that are particularly interesting since they specify area identities in the mouse telencephalon. To analyze their roles in precursor cells I chose gain-of-function experiments. Overexpression of these transcription factors showed that Emx2, Emx1 and Pax6 affect precursor cells in a region-specific manner. Emx2 transduction increases proliferation by promoting symmetric cell divisions, whereas blockade of endogenous Emx2 by antisense Emx2 mRNA limits the number and fate of progenitors generated by an individual cortical precursor cell. In the Emx2-/- asymmetrical cell divisions are increased in the cerebral cortex in vivo. In contrast to Emx2 Pax6 decreases proliferation. Pax6 deficient cells show more symmetrical cell divisions while Pax6 promotes asymmetric cell divisions in vitro. Emx2 endows in vitro cortical precursor cells with the capacity to generate multiple cell types, including neurons, astrocytes and oligodendrocytes. Emx1 keeps cells in an undifferentiated cell type, while Pax6 increases the proportion of neurons and can also convert astrocytes to neurons. The bHLH transcription factors Olig2 and Mash1 are up-regulated upon Emx2-transduction whereas Pax6 negatively influences those transcription factors and specifically up-regulates Ngn2. Thus, Emx2 is the first cell-intrinsic determinant able to instruct CNS precursors towards a multipotential fate. These results demonstrated an important role of Pax6 as intrinsic fate determinant of the neurogenic potential of glial cells. Taken together, Emx2 and Pax6 have opposing roles in cell proliferation, mode of cell division and cell fate.

The forebrain is generated by distinct sets of precursor cells that express specific transcription factors as well as secreted signaling factors in a time- and region-dependent manner. The distribution of these factors is similar in the avian and mammalian forebrains. In this work I aimed to examine the molecular mechanisms regulating telencephalic patterning. Therefore, I first compared the expression pattern of homeobox transcription factors, known to play crucial roles in regionalization of the forebrain, such as Emx1 and Emx2. This analysis showed particularly intriguing domains in the developing telencephalon expressing either only Emx2, such as the dorso-ventricular ridge (DVR) and the cortical hem, both genes, such as the hippocampus and the pallium or none, such as the subpallium and the choroid plexus (ChP). Taken together with other expression patterns I could conclude that the DVR, the nature of which was debated for a long period of time, displays a dorsal nature and that the pallial/subpallial boundary is located between DVR and subpallium. Next, I aimed to examine the role of Emx1 and Emx2 in the specification of these distinct regions. Therefore, I used a misexpression approach targeting Emx1 and Emx2 into the anlage of the choroid plexus (ChP) where these transcription factors are normally not expressed. In this region normal development was disturbed. The normally non-neuronal, thin morphology of the ChP with a low rate of proliferation and the characteristic expression of Otx2 and Bmp7 was lost. Instead, the rate of proliferation and the thickness of the tissue were increased and rather displayed “hem-like” properties. Instead, the Otx2-positive region of the ChP was shifted beside the region of ectopic Emx1/2-expression and exhibited intermediate properties, with features of ChP-tissue like Otx2 and Bmp7-expression, but also features of the cortical hem with a higher rate of proliferation and increased thickness of tissue. Thus, Emx1 and Emx2 play a key role in instructing dorsal neuroepithelium to proliferate. The misexpression of these genes is sufficient to convert non-neuronal ChP-tissue into neuroepithelium. Ectopic expression of Emx1/2 in the dorsal pallium, its normal region of expression, also displayed alterations. Ectopic Emx-expression blocked the expression of the neurogenic transcription factor Pax6 and suppressed neuronal differentiation. This change of neuronal differentiation could be caused by reduction of Pax6. Taken together, Emx1 and Emx2 are two potent factors that can change regional identity, enhance proliferation and block neuronal differentiation.

Diese Arbeit befasst sich mit der Embryonalentwicklung des Vorderhirns bei der Maus. Es werden die zellulären und molekularen Mechanismen untersucht, die eine distinkte Entwicklung von zwei benachbarten Regionen im Telencephalon, dem zerebralen Cortex und dem Striatum, ermöglichen. Es wird gezeigt, dass Zellen, die im Cortex entstehen, innerhalb des Cortex wandern, aber nicht über die Grenze in den Striatum hinein wandern können. Auf der anderen Seite können Zellen aus dem Striatum in den Cortex hinein wandern. Die Untersuchung dieser Zellwanderung in Mausmutanten zeigt, dass die Transkriptionsfaktoren Ngn2 und Pax6, die nur von den corticalen und Grenz-Zellen exprimiert werden, notwendig sind für die Restriktion der Zellen innerhalb des Cortex. Pax6 muss auch anwesend sein, um auch die Wanderung der striatalen Zellen gering zu halten. Weiterhin wird gezeigt, dass die interzelluläre Kommunikation via Gap-Junctions an der Grenzregion zwischen Cortex und Striatum unterbrochen wird. Somit weist die cortico-striatale Grenze die gleichen Merkmale wie andere Grenzen in der Embryonalentwicklung von Vertebraten oder auch von Insekten: Eine distinkte Genexpression, die Restriktion der Zellwanderung, und die Unterbrechung der interzellulären Kommunikation.