The tight junction scaffolding protein cingulin regulates neural crest cell migration

Abstract

Neural crest cells give rise to a diverse range of structures during vertebrate development. These cells initially exist in the dorsal neuroepithelium and subsequently acquire the capacity to migrate. Although studies have documented the importance of adherens junctions in regulating neural crest cell migration, little attention has been paid to tight junctions during this process. We now identify the tight junction protein cingulin as a key regulator of neural crest migration. Cingulin knock-down increases the migratory neural crest cell domain, which is correlated with a disruption of the neural tube basal lamina. Overexpression of cingulin also augments neural crest cell migration and is associated with similar basal lamina changes and an expansion of the premigratory neural crest population. Cingulin overexpression causes aberrant ventrolateral neuroepithelial cell delamination, which is linked to laminin loss and a decrease in RhoA. Together, our results highlight a novel function for cingulin in the neural crest.

The tight junction protein claudin-1 influences cranial neural crest cell emigration

“…To uncover possible effects on molecular markers associated with EMT, we inspected serial transverse sections of treated embryos for changes in Cad6-B and laminin. We found no significant difference in the distribution and level of these proteins upon claudin-1 overexpression (Supplementary Fig 1c,d), similar to what we have observed previously (Wu et al, 2011). In summary, claudin-1 appears to be a critical component in the tight junctions linking premigratory neural crest cells, since loss of claudin-1 leads to changes in gene expression that point to premature neural crest cell emigration and an increase in the number of migratory neural crest cells, while overexpression of claudin-1 results in the opposite phenotype.…”

“…To elucidate potential effects on molecular markers associated with EMT, we examined serial transverse sections of treated embryos for changes in the premigratory neural crest cell marker Cadherin6B (Cad6-B) and laminin, which labels the basal lamina. We found no significant difference in the distribution and level of these proteins upon claudin-1 depletion (Supplemental Fig 1a,b), comparable (for Cad6-B) with our prior studies on cingulin, another tight junction component (Wu et al, 2011). …”

“…HNK-1, a protein found on the surface of migratory neural crest cells (Bronner-Fraser, 1986), was increased on the electroporated side of 3/4 embryos electroporated with Claudin-1 MO but not in 5/5 controls (Fig 2i,j; yellow arrowhead in i). We also observed no change in cell proliferation via phospho-histone H3 (PH3) immunostaining (Fig 2k,l, 3/3 embryos)(Coles et al, 2007; Jhingory et al, 2010; Wu et al, 2011) or in cell death through a TUNEL assay (Fig 2m,n, 4/4 embryos)(Coles et al, 2007; Jhingory et al, 2010; Wu et al, 2011) or caspase 3 immunostaining (data not shown) in the neural tube and migratory neural crest cell population upon depletion of claudin-1.…”

“…It is known that tight junctions must be dismantled prior to neural crest cell emigration (Sauka-Spengler and Bronner-Fraser, 2008) and that occludin, a tight junction transmembrane protein, is down-regulated in the neuroepithelium during neural tube closure (Aaku-Saraste et al, 1996). It was also recently shown that the tight junction scaffolding protein cingulin plays a critical role in regulating neural crest cell emigration in the developing chick embryo (Wu et al, 2011). We now characterize the expression of claudin-1 in early chick embryos and suggest a functional role for claudin-1 during neural crest cell emigration.…”

Pard3 regulates contact between neural crest cells and the timing of Schwann cell differentiation but is not essential for neural crest migration or myelination

“…Down-regulation of some tight junction components coincides with neural tube closure and neural crest delamination (Sauka-Spengler and Bronner-Fraser, 2008) and knock-down of the tight junction protein Cingulin enlarged the migratory neural crest population (Wu et al, 2011). We therefore considered the possibility that Pard3 similarly influences neural crest delamination.…”