Characterization and Endocytic Internalization of Epith-2 Cell Surface Glycoprotein during the Epithelial-to-Mesenchymal Transition in Sea Urchin Embryos

Abstract

The epithelial cells of the sea urchin Hemicentrotus pulcherrimus embryo express an Epith-2, uncharacterized glycoprotein, on the lateral surface. Here, we describe internalization of Epith-2 during mesenchyme formation through the epithelial-to-mesenchymal transition (EMT). Epith-2 was first expressed on the entire egg surface soon after fertilization and on the blastomeres until the 4-cell stage, but was localized to the lateral surface of epithelial cells at and after the 16-cell stage throughout the later developmental period. However, primary mesenchyme cells (PMC) and secondary mesenchyme cells (SMC) that ingress by EMT lost Epith-2 from their cell surface by endocytosis during dissociation from the epithelium, which was associated with the appearance of cytoplasmic Epith-2 dots. The cytoplasmic Epith-2 retained a similar relative molecular mass to that of the cell surface immediately after ingression through the early period of the spreading to single cells. Then, Epith-2 was completely lost from the cytoplasm. Tyrosine residues of Epith-2 were phosphorylated. The endocytic retraction of Epith-2 was inhibited by herbimycin A (HA), a protein tyrosine kinase (PTK) inhibitor, and suramin, a growth factor receptor (GFR) inhibitor, suggesting the involvement of the GFR/PTK (GP) signaling pathway. These two GP inhibitors also inhibited PMC and SMC spreading to individual cells after ingression, but the dissociation of PMC and SMC from the epithelium was not inhibited. In suramin-treated embryos, dissociated mesenchyme cells migrated partially by retaining their epithelial morphology. In HA-treated embryos, no mesenchyme cells migrated. Thus, the EMT occurs in relation to internalization of Epith-2 from presumptive PMC and SMC.

Mechanisms of the epithelial-to-mesenchymal transition in sea urchin embryos

“…Herbimycin A, a protein tyrosine kinase (PTK) inhibitor inhibits the retraction of the Epith-2 protein from the PMC surface, and these cells remain half embedded in the vegetal ectoderm. 61 Thus, the epitope of the anti-Epith-2 mAb is involved in epithelial cell adhesion. However, its molecular mechanism remains to be elucidated.…”

“…The spatiotemporal change in the expression pattern of Epith-1 and -2 from the very early period of cleavage to the total loss from the cell surface during migration in the blastocoel 4,8,61 strongly suggests the importance of the involvement of substances on the cell surface in the EMT signaling mechanism. Because the gene structures of Epith-1 and -2 have not been elucidated, further genomic analysis will enable the evaluation of the molecular regulatory mechanism of the morphogenetic role of these cell surface proteins.…”

“…Then the "puncta" are lost when PMCs begin to migrate, 10 which resembles the fates of Epith-1 8 and Epith-2. 61 In L. variegatus embryos, LvG-cadherin expression is suppressed by injection of a paired-class micromere anti-repressor (Pmar1) that is upstream of the TCF ets1. Ets1 is necessary for PMC formation.…”

“…Epith proteins remain in cytoplasmic vesicles and lose their immunoreactivity by the onset of migration. 8,10,61 Conversely, PMC surface-specific Msp130 appears in the cytoplasm and is transferred to the cell surface along with NgCAM in S. purpuratus. 10 However, the timing of the loss and gain of these cell surface proteins during ingression is slightly different among sea urchin species.…”

Posttranslational Modification of Intercellular Messenger Systems

“…This Research Topic is aimed to assemble a series of review articles and original research papers on structural analysis or functional significance of posttranslational modification of/by intercellular messengers, including hormonal messengers and gaseous messengers, in vertebrates and invertebrates: posttranslational modification of peptide hormones such as proopiomelanocortin (1, 2), ghrelin (3)(4)(5), and hormonal receptors and effectors (6)(7)(8). Review articles on gaseous messengers such as hydrogen sulfide (9) and nitric oxide (10) are also included.…”