Sema6a and Plxna2 mediate spatially regulated repulsion within the developing eye to promote eye vesicle cohesion

Abstract

Organs are generated from collections of cells that coalesce and remain together as they undergo a series of choreographed movements to give the organ its final shape. We know little about the cellular and molecular mechanisms that regulate tissue cohesion during morphogenesis. Extensive cell movements underlie eye development, starting with the eye field separating to form bilateral vesicles that go on to evaginate from the forebrain. What keeps eye cells together as they undergo morphogenesis and extensive proliferation is unknown. Here, we show that plexina2 (Plxna2), a member of a receptor family best known for its roles in axon and cell guidance, is required alongside the repellent semaphorin 6a (Sema6a) to keep cells integrated within the zebrafish eye vesicle epithelium. sema6a is expressed throughout the eye vesicle, whereas plxna2 is restricted to the ventral vesicle. Knockdown of Plxna2 or Sema6a results in a loss of vesicle integrity, with time-lapse microscopy showing that eye progenitors either fail to enter the evaginating vesicles or delaminate from the eye epithelium. Explant experiments, and rescue of eye vesicle integrity with simultaneous knockdown of sema6a and plxna2, point to an eye-autonomous requirement for Sema6a/Plxna2. We propose a novel, tissue-autonomous mechanism of organ cohesion, with neutralization of repulsion suggested as a means to promote interactions between cells within a tissue domain.

Identification of target genes downstream of semaphorin6A/PlexinA2 signaling in zebrafish

“…Confirming our previous results (Ebert et al, ), morpholino antisense oligonucleotides (MOs) targeted to either sema6A or plxnA2 resulted in decreased proliferation and cohesion of RPCs within migrating optic vesicles compared with control embryos. To observe these phenotypes, we used the rx3 :GFP zebrafish transgenic line that expresses green fluorescent protein (GFP) in RPCs and the hypothalamus (Rembold et al, ).…”

“…We observed overlap between previously reported plxnA2 and sema6A expression (Ebert et al, ) and the candidate genes, supporting a relationship between Sema6A/PlxnA2 signaling and these genes. We generated cDNA from uninjected control, sema6A morphant, and plxnA2 morphant embryos at 18 somites and performed RT‐PCR (Fig.…”

PKC induces release of a functional ectodomain of the guidance cue semaphorin6A

“…However, the intracellular domain confers resistance to ectodomain release, as a truncated form of Sema6A lacking the intracellular domain increases sSema6A production. Finally, we show that sSema6A is functional as it promotes the cohesion of early zebrafish eye fields in cultured explants, a phenotype known to be induced by Sema6A‐FC . To our knowledge, these data constitute the first characterization of a natural and regulated release of an ectodomain for a Sema6 class protein that functions in the nervous system.…”

“…We have been studying the molecular mechanisms and developmental function of Sema6A‐PlxnA signaling. Previously, we reported that Sema6A is highly expressed in retinal precursor cells in the early eye field and is critical in maintaining early eye field cohesion . Given that our proteomics analyses identified Sema6A in CSF , we were prompted to ask if Sema6A might generate a naturally released ectodomain.…”

Adhesive/Repulsive Codes in Vertebrate Forebrain Morphogenesis

“…In addition, the BMP and the Wnt/βcatenin-independent pathways control not only eye fate specification, but also eye field integrity by promoting cell cohesion in this domain and controlling some of the same molecules controlled by Rx3 [82,119] (Figure 4). A very recent study has added a further player to this list of adhesion/repulsion mechanisms, by identifying the Semaphorin/Plexin ligand-receptor pair as a critical controller of optic vesicle integrity [120]. In their study, the authors show that these two molecules generate a code of "attractive" and "repulsive" regions crucial for the maintenance of the regionalization of the evaginating optic vesicle as morphogenesis proceeds, and for its overall integrity.…”