Modena, Nicola (2008) Lo Zebrafish come strumento per lo studuio integrato di segnali endocrini e paracrini che regolano le prime fasi dello sviluppo tissutale. [Ph.D. thesis]
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Studies performed in this research are aimed to define how different cell types are specified during neural development of the model organism zebrafish (Danio rerio), a little teleost fish of Cyprinid family.
Investigations involved the analysis of Olig genes, a family of transcriptional factors important for development and determination of different cell types of CNS (Central Nervous System), such as interneurons, motoneurons, oligodendrocytes, astrocytes, and neural crest cells, a structure that is unique of vertebrates.
Phylogenetic comparisons (Bronchain et al., 2007) identified three Olig members in amniotes, while in fishes and amphibians four members were isolated: olig1, olig2, olig3 and olig4.
Moreover, this project dealt with a microarray analysis to identify molecular targets controlled by the transcriptional factor olig4. So, this led to the identification of a group of genes related to the inactivation of this transcriptional factor that offered a wide image on possible regulatory patterns involved in CNS development, and on possible olig4 function in limiting neural crest development.
The availability of an olig4 genetic mutant is now confirming results already observed during microarray analysis and further evidences are coming from in situ hybridization staining using a group of target genes.
An additional member of the olig family has been cloned in zebrafish and it has been named olig3 on the basis of phylogenetic comparisons (Bronchain et al., 2007). In zebrafish, olig3 in situ hybridization staining analysis revealed that expression patterns of olig3 and olig4 (which sequences show a high level of similarity), together correspond to the expression pattern of murine Olig3 (an homologue of olig4 in mouse has not yet been identified).
In this study, the zebrafish olig3 gene has been cloned and its expression pattern compared with that of some neural tube markers and other members of the olig family, to elucidate inter-reciprocal position of expression domains.
Analysis have been performed based on treatments with specific inhibitors of some paracrine signaling pathways involved in zebrafish CNS development. This analysis allowed focusing on regulatory pathways of olig family members controlled by morphogenetic gradients during early embryonic development.
Furthermore, this research project involved a study on zebrafish moonshine (mon) gene. This gene codifies for Trimm33, a protein implicated in adult hematopoiesis. Zebrafish mon gene product is related to different co-activators an co-repressors of the TGF-ß signaling pathway.
In this study, genetic moonshine mutants (montg234) were treated with an artificial inhibitor of TGF-ß signaling pathway that allows reproducing in vivo the phenotype of genetic mutants for this important signaling pathway. The effect obtained after the treatment showed in zebrafish a restored expression of haematopoietic precursor markers such as gata1 and this was confirmed through in situ hybridization staining. Results added insight on mon involvement upstream TGF-ß signals in zebrafish haematopoietic pathway.
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