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- Baumgardt, Magnus, 1976-
(författare)
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Genetic mechanisms behind cell specification in the Drosophila CNS
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The human central nervous system (CNS) contains a daunting number of cells and tremendous cellular diversity. A fundamental challenge of developmental neurobiology is to address the questions of how so many different types of neurons and glia can be generated at the precise time and place, making precisely the right connections. Resolving this issue involves dissecting the elaborate genetic networks that act within neurons and glia, as well as in the neural progenitor cells that generates them, to specify their identities.My PhD project has involved addressing a number of unresolved issues pertaining to how neural progenitor cells are specified to generate different types of neurons and glial cells in different temporal and spatial domains, and also how these early temporal and spatial cues are integrated to activate late cell fate determinants, which act in post-mitotic neural cells to activate distinct batteries of terminal differentiation genes.Analyzing the development of a specific Drosophila melanogaster (Drosophila) CNS stem cell – the neuroblast 5-6 (NB5-6) – we have identified several novel mechanisms of cell fate specification in the Drosophila CNS. We find that, within this lineage, the differential specification of a group of sequentially generated neurons – the Ap cluster neurons – is critically dependent upon the simultaneous triggering of two opposing feed-forward loops (FFLs) within the neuroblast. The first FFL involves cell fate determinants and progresses within the post-mitotic neurons to establish a highly specific combinatorial code of regulators, which activates a distinct battery of terminal differentiation genes. The second loop, which progresses in the neuroblast, involves temporal and sub-temporal genes that together oppose the progression of the first FFL. This leads to the establishment of an alternative code of regulators in late-born Ap cluster neurons, whereby alternative cell fates are specified. Furthermore, we find that the generation and specification of the Ap cluster neurons is modulated along the neuraxis by two different mechanisms. In abdominal segments, Hox genes of the Bithorax cluster integrates with Pbx/Meis factors to instruct NB5-6 to leave the cell cycle before the Ap cluster neurons are generated. In brain segments, Ap cluster neuron equivalents are generated, but improperly specified due to the absence of the proper Hox and temporal code. Additionally, in thoracic segments we find that the specification of the Ap cluster neurons is critically dependent upon the integration of the Hox, Pbx/Meis, and the temporal genes, in the activation of the critical cell fate determinant FFL.We speculate that the developmental principles of (i) feed-forward combinatorial coding; (ii) simultaneously triggered yet opposing feed-forward loops; and (iii) integration of different Hox, Pbx/Meis, and temporal factors, at different axial levels to control inter-segmental differences in lineage progression and specification; might be used widely throughout the animal kingdom to generate cell type diversity in the CNS.
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| 2. |
- O'Farrell, Fergal
(författare)
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A screen for mutations affecting PNS development in Drosophila identifies the trim gene, dappled
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The peripheral nervous system of Drosophila melanogaster contains a variety of sense organs, ranging from the relatively simple four celled bristle organ to the more complex compound eye. The development of each organ type is well described, providing a useful backdrop for functional studies of genes acting in one or more of the many processes involved in organogenesis. We have used the bristle organ to screen for genes affecting PNS development. Two of the candidates recovered via this approach, string (stg, Drosophila cdc25, the universal regulator of the G2 to M phase mitotic transition), and dappled (dpld, a poorly described gene implicated in tumor suppression) were selected for further study. Examination of stg mis-expression phenotypes in the adult bristle organ revealed cell fate transformations corresponding to the generation of two pIIa structural precursor cells at the expense of a neural precursor cell. This transformation most reasonably resulted from an abnormally short G2 arrest, indicating that the time spent in the G2 phase is crucial to correct cell fate determination. dpld is a member of the Tripartite Motif (TRIM) superfamily, members of which are involved in diverse biological processes e.g. proliferation, apoptosis and immune response. dpld belongs to a subgroup of NHL domain containing TRIM proteins, that are known to be involved in tumor suppression. Phylogenetic analysis placed dpld in the lin-41 sub-clade of the TRIM superfamily. A combination of insilico, genetic and cell culture assay approaches showed dpld to be susceptible to miRNA regulation. As homologous genes are also miRNA regulated this regulatory mechanism may be conserved throughout this sub-clade, between vertebrates and invertebrates. Pre-existing loss of function dpld alleles were characterized, however, subsequent complementation studies revealed that characteristic aspects of the described dpld phenotype, in fact mapped outside the dpld locus, and were caused by mutations of nearby genes. The tumor-causing locus was mapped to the Cytb5 gene (mutated in both pre-existing dpld alleles), while the embryonic lethality and PNS phenotype was mapped to the scraps locus. scraps encodes for Drosophila Anillin, known to be required during cytokinesis. We provide the first characterization of scraps null alleles and detail a biased requirement for scraps within neural precursor cells of the embryonic PNS. A novel loss of function dpld allele was recovered. This mutation is lethal, however it does not have an associated tumor phenotype. This finding, together with our complementation study indicates that the existing classification of dpld as a tumor suppressor is inaccurate. Subsequent studies detail dpld requirements in the developing fly retina. There, dpld mutation resulted in excessive proliferation, while conversely, mis-expression caused a reduction. Additionally, and perhaps consequently, cell differentiation was affected. Thus, regulation of proliferation by NHL-TRIM genes seems a conserved feature. We additionally identified a novel Drosophila TRIM gene of the same class as dpld, which we have dubbed another bbox affiliate (abba), bringing the number of NHL containing TRIM genes in Drosophila to four.
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