Porcn is essential for growth and invagination of the mammalian optic cup

Abstract

Microphthalmia, anophthalmia, and coloboma (MAC) are congenital ocular malformations causing 25% of childhood blindness. The X-linked disorder Focal Dermal Hypoplasia (FDH) is frequently associated with MAC and results from mutations in Porcn, a membrane bound O-acyl transferase required for palmitoylation of Wnts to activate multiple Wnt-dependent pathways. Wnt/β-catenin signaling is suppressed in the anterior neural plate for initiation of eye formation and is subsequently required during differentiation of the retinal pigment epithelium (RPE). Non-canonical Wnts are critical for early eye formation in frog and zebrafish. However, it is unclear whether this also applies to mammals. We performed ubiquitous conditional inactivation of Porcn in mouse around the eye field stage. In PorcnCKO, optic vesicles (OV) arrest in growth and fail to form an optic cup. Ventral proliferation is significantly decreased in the mutant OV, with a concomitant increase in apoptotic cell death. While pan-ocular transcription factors such as PAX6, SIX3, LHX2, and PAX2 are present, indicative of maintenance of OV identity, regional expression of VSX2, MITF, OTX2, and NR2F2 is downregulated. Failure of RPE differentiation in PorcnCKO is consistent with downregulation of the Wnt/β-catenin effector LEF1, starting around 2.5 days after inactivation. This suggests that Porcn inactivation affects signaling later than a potential requirement for Wnts to promote eye field formation. Altogether, our data shows a novel requirement for Porcn in regulating growth and morphogenesis of the OV, likely by controlling proliferation and survival. In FDH patients with ocular manifestations, growth deficiency during early ocular morphogenesis may be the underlying cause for microphthalmia.

Optic cup morphogenesis across species and related inborn human eye defects

“…In mouse organoids, failed RPE specification prevents the initial steps of OV invagination ( Eiraku et al, 2011 ), although, once the OC is formed, the RPE seems to function as a shell, with high mechanical rigidity and a strong actomyosin activation that, if broken, does not impact on the convex NR shape ( Eiraku et al, 2011 ). OC malformations have also been reported after genetic manipulations of components of the Wnt and Fgf signalling pathways that interfere with the acquisition of RPE and CM identity and/or OV growth ( Balasubramanian et al, 2021 ; Bankhead et al, 2015 ; Carpenter et al, 2015 ; Fuhrmann et al, 2022 ). For example, failed secretion of lens-derived Wnt ligands perturbs proliferation of the peripheral region of the eye cup, with a more evident effect on cells of the RPE ( Carpenter et al, 2015 ).…”

“…This defect is associated with a poorly folded, ‘saucer’ shaped OC ( Balasubramanian et al, 2021 ; Carpenter et al, 2015 ). Similarly, preventing post-translational modifications, and thus secretion, of Wnt ligands in the eye field diminishes OV growth and impairs its folding ( Fuhrmann et al, 2022 ). In other words, RPE specification ( Balasubramanian et al, 2021 ; Hägglund et al, 2013 ; Martinez-Morales et al, 2001 ; Tang et al, 2010 ;) and growth ( Carpenter et al, 2015 ; Fuhrmann et al, 2022 ) are prerequisites for OC generation in vivo .…”

Signaling – transcription interactions in mouse retinal ganglion cells early axon pathfinding –a literature review

“…Among the multiple FGF ligands, FGF8 coming from the telencephalic vesicles has the main role in optic vesicle patterning, while the others are able to compensate in its absence, as shown in zebrafish ( 82 ). RPE differentiation requires the Wnt/beta-catenin pathway, including Porcn function ( 83 ).…”