Sox2 regulatory sequences direct expression of a beta-geo transgene to telencephalic neural stem cells and precursors of the mouse embryo, revealing regionalization of gene expression in CNS stem cells

Sox2 is one of the earliest known transcription factors expressed in the de veloping neural tube. Although it is expressed throughout the early neuroep ithelium, we show that its later expression must depend on the activity of more than one regionally restricted enhancer element. Thus, by using transg enic assays and by homologous recombination-mediated deletion, we identify a region upstream of Sox2 (-5.7 to -3.3 kb) which can not only drive expres sion of a beta-geo transgene to the developing dorsal telencephalon, but wh ich is required to do so in the context of the endogenous gene. The critica l enhancer can be further delimited to an 800 bp fragment of DNA surroundin g a nuclease hypersensitive site within this region, as this is sufficient to confer telencephalic expression to a 3.3 kb fragment including the Sox2 promoter, which is otherwise inactive in the CNS. Expression of the 5.7 kb Sox2 beta-geo transgene localizes to the neural pl ate and later to the telencephalic ventricular zone. We show by in vitro cl onogenic assays, that transgene-expressing (and thus G418-resistant) ventri cular zone cells include cells displaying functional properties of stem cel ls, i.e. self-renewal and multipotentiality, We further show that the major ity of telencephalic stem cells express the transgene, and this expression is largely maintained over two months in culture (more than 40 cell divisio ns) in the absence of G418 selective pressure. In contrast, stem cells grow n in parallel from the spinal cord never express the transgene, and die in G418, Expression of endogenous telencephalic genes was similarly observed i n long-term cultures derived from the dorsal telencephalon, but not in spin al cord-derived cultures. Thus, neural stem cells of the midgestation embryo are endowed with region- specific gene expression (at least with respect to some networks of transcr iption factors, such as that driving telencephalic expression of the Sox2 t ransgene), which can be inherited through multiple divisions outside the em bryonic environment.