Hedgehog signaling regulates cell motility and optic fissure and stalk formation during vertebrate eye morphogenesis

Abstract

Establishment of precise three-dimensional tissue structure is vital for organ function. In the visual system, optic fissure and stalk morphogenesis is a crucial yet poorly understood process, disruptions of which can lead to coloboma, a birth defect causing visual impairment. Here, we use four-dimensional imaging, cell tracking, and molecular genetics in zebrafish to define the cell movements underlying normal optic fissure and stalk formation. We determine how these events are disrupted in a coloboma model in which the Hedgehog (Hh) receptor ptch2 is lost, resulting in overactive Hh signaling. In the ptch2 mutant, cells exhibit defective motile behaviors and morphology. Cells that should contribute to the fissure do not arrive at their correct position, and instead contribute to an ectopically large optic stalk. Our results suggest that overactive Hh signaling, through overexpression of downstream transcriptional targets, impairs cell motility underlying optic fissure and stalk formation, via non-cell-autonomous and cell-autonomous mechanisms. More broadly, our cell motility and morphology analyses provide a new framework for studying other coloboma-causing mutations that disrupt optic fissure or stalk formation.

Optic cup morphogenesis requires neural crest-mediated basement membrane assembly

“…Several signaling pathways crucial for eye development are regulated in part by POM, including Hh, TGFβ and Wnt (Fuhrmann et al, 2000;Grocott et al, 2011;Sedykh et al, 2017), thus neural crest cells could drive optic cup morphogenesis by modulating morphogen signaling. However, in zebrafish tfap2a;foxd3 mutants, Pax2a expression is not expanded in a manner consistent with aberrant Hh signaling (Gordon et al, 2018;Lee et al, 2008;Sedykh et al, 2017), nor is TGFβ signaling altered. The remaining mesodermal mesenchyme may be sufficient for proper Hh and TGFβ signaling in zebrafish; additional studies will be necessary to dissect specific roles of mesodermal mesenchyme during optic cup morphogenesis.…”

“…To determine how optic cup morphogenesis might be influenced by neural crest, we first asked when and where these cells interact with the developing eye. To visualize both the eye and neural crest, we used double-transgenic Tg(bactin2:EGFP-CAAX) z200 ;Tg(sox10:memRFP) vu234 zebrafish in which GFP labels all cell membranes and RFP labels neural crest cell membranes (Gordon et al, 2018;Kirby et al, 2006). We performed four-dimensional time-lapse imaging of optic cup morphogenesis (12.5-24.5 hpf; Fig.…”

“…We first examined movements of nasal retina and RPE cells because we saw gross morphological defects in the nasal hemisphere of tfap2a;foxd3 double-mutant eyes. We selected these cells from their final positions at 24 hpf in double-mutant and wild-type data sets (Gordon et al, 2018), and tracked these cells retrospectively to establish their origins and movements from the optic vesicle stage; a subset of cells and trajectories is shown (Fig. 2).…”

“…Transgenic alleles used were: Tg(sox10:memRFP) vu234 (Kirby et al, 2006), Tg(sox10:GFP) ba4 (Dutton et al, 2008), Tg(bactin2:EGFP-CAAX) z200 (Gordon et al, 2018), Tg(hsp70:lyn-mCherry-2A-WT-Nid1a) z202 and Tg(hsp70:lyn-mCherry-2A-DI-Nid1a) z203 .…”

Primary cilia deficiency in neural crest cells causes Anterior Segment Dysgenesis

“…The Hh signal transduction pathway is intimately linked to the primary cilium [23,24], and its dosage is critical to ensure normal eye development [38,82,83]. In this study, we have shown that at an early stage of the AS development virtually all the POM cells are actively responding to Hh signaling.…”