Parsons, Michael John; (1997) Functional analysis of Sox3: An X-linked Sry related gene. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Sox genes comprise a substantial family of genes, each encoding a transcription factor. These proteins all possess an HMG box DNA binding domain with significant homology to that of the mammalian testes determining gene, Sry. Sox3 is an X-linked member of the family with an HMG box that is most homologous to that of SRY. In order to ascertain the expression of Sox3 during mammalian embryogenesis, whole mount in situ hybridisations were performed with antisense cRNA probes. Results indicate that in streak stage embryos, Sox3 is expressed strongly in extra-embryonic ectoderm of the developing chorion and at a lower level in the embryonic ectoderm. Later in development, expression is restricted to the neuroepithelium where Sox3 is detected throughout embryogenesis. To investigate the function of Sox3 during mouse embryogenesis, this X-linked gene was disrupted by homologous recombination in male (XY) embryonic stem cells. Results were obtained from the use of two targeting constructs: one inserted a selectable marker into the HMG box of Sox3 and the second replaced 75% of the coding region of Sox3 with a selectable marker. ES cell lines made with both of these constructs were tested for their ability to function in chimaeric mice and showed identical phenotypes. The resulting chimaeras were shown to die at about 10.5dpc. At a gross morphological level, the phenotype of the mutant embryos showed a failure of mesodermal derivatives to be properly organised; namely a failure of the allantois to fuse with the chorion, aberrant somitogenesis, a shortened posterior axis and head defects. Surprisingly, the mutant ES cells only had to contribute to 30% of the embryo to cause the mutant phenotypes which were observed. Differentiation of ES cells in vitro showed that the mutant cells could form derivatives of all three germ layers. These studies suggest that Sox3 plays an essential role in mouse embryogenesis. What this role may be and methods to further investigate SOX3 function are discussed.

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