Abstract
The study of the neural crest is fundamental in answering questions that arise in both Stem Cell Biology and Developmental Biology. This project focuses on the development of a neural crest derivative, the melanocyte. In zebrafish, melanocyte specification depends upon the Sry-related High Mobility Group box gene (sox) sox10 and Wnt signalling to drive the transcription of mitfa (microphthalmia-associated transcription factor). However, the mechanisms involved in stable melanocyte differentiation are not fully understood. In vivo data suggested that sox10 formed part of a Feed-Forward Loop repressing melanocyte differentiation and is relieved by Mitfa-dependent repression of sox10. Aspects of the melanocyte gene regulatory network derived from experimental data and mathematical modelling were tested with the aim of improving the understanding of melanocyte biology.Modelling predicted an unknown factor, named Factor Y, was involved as a limiting factor for the maintenance of mitfa expression in differentiating melanocytes. Wnt signalling was tested as a candidate for this factor by investigating the effects of small molecule activators of the pathway during melanocyte differentiation. Resultantly, we observed increased cell dendricity, disruption of cell organisation pattern in the head of embryos and activation of mitfa expression. However, no other pronounced changes in cell differentiation were seen. Therefore, we suggest that maintenance of mitfa and of melanocyte differentiation would depend on a complex regulatory ensemble which includes factors other than Wnt signalling.
sox10 and mitfa expression levels were characterised through time using the Real time Quantitative Polymerase Chain Reaction to quantify the genetic variability occurring during melanocyte derivation. The previously described decrease of sox10 expression in melanocyte lineage was confirmed. Unexpectedly, the decrease of mitfa expression was observed suggesting that mitfa is maintained at low levels in melanocytes where stochastic effects could be important. This could also suggests the existence of a new unknown factor involved downstream of mitfa for melanocyte differentiation.
The role of histone deacetylase (Hdac(s)) as a candidate repressor of sox10 expression during differentiation was investigated in melanocytes. Treating embryos with Hdac(s) inhibitor caused the regression of melanocyte differentiation and the persistence of sox10 expression in melanocytes. These results suggested a potential role for Hdac in Mitfa-dependent downregulation of sox10.
| Date of Award | 1 Sept 2013 |
|---|---|
| Original language | English |
| Awarding Institution |
|
| Supervisor | Robert Kelsh (Supervisor) |