The sex determining region Y-box 2 (sox2) gene is one of the most important
transcription factors during development, particularly the development of the central
nervous system (CNS). It is expressed in embryonic stem cells and later in neural stem
cells, where it modulates their maintenance and differentiation. In humans, heterozygous
mutations are associated with eye malformations, including anophthalmia and severe
microphthalmia. Also, a subset of patients has extra-ocular phenotypes, such as hearing
loss, seizures and pituitary hypoplasia. Although the roles of sox2 in embryonic stem
cells and eye development are well studied, the function of sox2 in brain development
and disease is still elusive. The aim of this project was to characterize a novel role for
sox2 in the development of zebrafish epithalamus, which was identified from an in silico
screen previously performed in our laboratory.
The zebrafish epithalamus, located in the dorsal diencephalon, consists of three
main structures: the pineal gland, the parapineal organ and the habenular nuclei. The
pineal gland, also known as epiphysis, is a photoreceptive (in zebrafish) and
neuroendocrine organ that detects light and rhythmically produces melatonin in order to
regulate the circadian rhythms. The parapineal organ is located to the left side of the
pineal gland and is important for the elaboration of the asymmetries observed between
the left and right habenular nuclei. Finally, the bilateral habenulae are part of the dorsal
diencephalic conduction system that links the forebrain with the mid- and hindbrain. The
left and right habenulae show both molecular and neuroanatomical asymmetries,
including differences in neuropil organization, in levels of gene expression and in the
morphology and connectivity of their neurons’ projections. The relatively simple
architecture of the pineal gland and the asymmetric character of the habenulae provide a
useful tool for studying cell-fate determination, cell migration and establishment of brain
asymmetries.
In this study, we used zebrafish as a model to dissect the novel functions of sox2
in the development of the epithalamus. We showed that sox2 works synergistically with
Notch pathway to negatively regulate neurogenesis within the pineal gland. The pineal
gland consists of only two cell types: the photoreceptors and the projection neurons.
Previous studies showed that the Notch and BMP pathways are important for the proper
specification of these cells. Here, we show that sox2 normally inhibits the photoreceptor
cell fate, whereas it has no effect on the number of projection neurons. Therefore, sox2
complements Notch and BMP pathways in cell-fate determination within the pineal
gland.
In addition, downregulation of sox2 results in abnormal parapineal organ
development and disruption of the asymmetric architecture of the habenulae. A subset of
sox2 morphant embryos develops right-sided parapineal organs, which is consistent with
abnormal bilateral expression of the Nodal gene, pitx2 (paired-like homeodomain
transcription factor 2). Also, timelapse experiments showed that migration of the
parapineal cells is defective, resulting in scattered cells. The aberrant parapineal
development leads to disorganization of the habenular nuclei, as shown by the abnormal
neuropil arrangement and the expression of the asymmetric marker kctd12.1 (potassium
channel tetramerisation domain containing 12.1)