Pax6 regulates the formation of the habenular nuclei by controlling the temporospatial expression of Shhin the diencephalon in vertebrates

Abstract

BackgroundThe habenula and the thalamus are two critical nodes in the forebrain circuitry and they connect the midbrain and the cerebral cortex in vertebrates. The habenula is derived from the epithalamus and rests dorsally to the thalamus. Both epithalamus and thalamus arise from a single diencephalon segment called prosomere (p)2. Shh is expressed in the ventral midline of the neural tube and in the mid-diencephalic organizer (MDO) at the zona limitans intrathalamica between thalamus and prethalamus. Acting as a morphogen, Shh plays an important role in regulating cell proliferation and survival in the diencephalon and thalamic patterning. The molecular regulation of the MDO Shh expression and the potential role of Shh in development of the habenula remain largely unclear.ResultsWe show that deleting paired-box and homeobox-containing gene Pax6 results in precocious and expanded expression of Shh in the prospective MDO in fish and mice, whereas gain-of-function of pax6 inhibits MDO shh expression in fish. Using gene expression and genetic fate mapping, we have characterized the expression of molecular markers that demarcate the progenitors and precursors of habenular neurons. We show that the thalamic domain is shifted dorsally and the epithalamus is missing in the alar plate of p2 in the Pax6 mutant mouse. Conversely, the epithalamus is expanded ventrally at the expense of the thalamus in mouse embryos with reduced Shh activity. Significantly, attenuating Shh signaling largely rescues the patterning of p2 and restores the epithalamus in Pax6 mouse mutants, suggesting that Shh acts downstream of Pax6 in controlling the formation of the habenula. Similar to that found in the mouse, we show that pax6 controls the formation of the epithalamus mostly via the regulation of MDO shh expression in zebrafish.ConclusionsOur findings demonstrate that Pax6 has an evolutionarily conserved function in establishing the temporospatial expression of Shh in the MDO in vertebrates. Furthermore, Shh mediates Pax6 function in regulating the partition of the p2 domain into the epithalamus and thalamus.

Heterozygous defects in PAX6 gene and congenital hypopituitarism

“…The known control that PAX6 exerts over the temporospatial expression of Sonic hedhehog (SHH) (16), the critical importance of PAX6 on pituitary development through SHH (9), and the critical importance of SHH on palate development (17) strongly support the argument for PAX6 defects being pathogenic events responsible for the phenotype. Previously, several authors described submicroscopic deletions downstream of PAX6, with the coding region of PAX6 being unaffected (18,19,20,21,22).…”

Distinct requirements for Wntless in habenular development

“…To determine the relationship between Wnt responsive cells and habenular development, we examined expression of the developing brain homeobox 1b ( dbx1b ) gene, whose transcripts are highly enriched in the developing habenulae and thalamic progenitors of the mouse (Chatterjee et al, 2014; Quina et al, 2009; Vue et al, 2007) and in proliferating cells of the presumptive dorsal habenulae of zebrafish (Dean et al, 2014). In WT embryos, GFP labeling from the Wnt reporter partially overlaps with dbx1b expression at 27 and 35 hpf (Fig.…”

Convergence of signaling pathways underlying habenular formation and axonal outgrowth in zebrafish

“…In zebrafish embryos homozygous for a mutation in the smo gene, which encodes a G protein-coupled receptor essential for Hh signaling, expression of cxcr4b was reported as absent in the epithalamic region in one study (Halluin et al, 2016) and expanded in another (Chatterjee et al, 2014). In our experiments, neither cxcr4b nor dbx1b transcripts were detected in this region of the smo mutant brain.…”

“…Recent studies have also implicated Hh signaling as an early regulator of habenular development (Chatterjee et al, 2014;Halluin et al, 2016). In zebrafish embryos homozygous for a mutation in the smo gene, which encodes a G protein-coupled receptor essential for Hh signaling, expression of cxcr4b was reported as absent in the epithalamic region in one study (Halluin et al, 2016) and expanded in another (Chatterjee et al, 2014).…”

“…However, there are conflicting reports on whether Hh signaling antagonizes or promotes formation of the dHb (Chatterjee et al, 2014;Halluin et al, 2016). Mutations in the zebrafish wntless (wls) gene, which encodes a protein essential for Wnt secretion, or in the fibroblast growth factor 8a ( fgf8a) gene, result in small dHb (Regan et al, 2009;Kuan et al, 2015).…”