Greenhill et al. (2011) developed a gene regulatory network of the main genes and interactions known to play a role in melanocyte biology, and generated a mathematical model to describe the behaviour of this complex network using semi quantitative data (ISH expression data). In this project we sought to collect expression data from four genes of the melanocyte GRN (sox10, kit, mitfa and dct) to develop a quantitative model that is able to describe the data more accurately. Moreover, we intended to identify more genes that are part of the melanocyte development process to be incorporated to the GRN.We analysed microarray data that compared differentially expressed genes between sox10 mutant and wild type embryos and validated five genes with a key role in melanocyte biology as downregulated in mitfa mutant embryos, which are downstream of mitfa in the GRN. We suggest that kit plays the role of factor Y in the Greenhill et al. (2011) GRN: Mitfa drives kit expression, and kit expression is transiently driven by Sox10 at early stages of development. As part of the feedback loop, kit seems to drive and maintain mitfa expression, however this needs to be validated. Finally we developed an experimental set up to obtain an estimate of gene expression per melanocyte from sox10, kit, mitfa and dct, using both qPCR and ISH cell count measurements. With this estimate we performed a parameter optimisation procedure, and found a set of parameters for the mathematical model that predicted the experimental data very accurately. The new model suggests that low expression values of sox10 are sufficient to drive mitfa expression in high levels. It also predicts that high expression of sox9b is needed to achieve the high expression levels of dct seen in the data, although these predictions need to be experimentally tested.This study represents the first attempt to obtain fine-scale gene expression data from melanocytes for the development of a quantitative mathematical model in zebrafish.
|Date of Award||10 May 2016|
|Supervisor||Robert Kelsh (Supervisor) & Hartmut Schwetlick (Supervisor)|
- Neural Crest
- Gene Regulatory Networks
- mathematical modeling