A Raf/MEK/ERK signaling pathway is required for development of the sea urchin embryo micromere lineage through phosphorylation of the transcription factor Ets

Abstract

In the sea urchin embryo, the skeleton of the larva is built from a population of mesenchymal cells known as the primary mesenchyme cells (PMCs). These derive from the large micromeres that originate from the vegetal pole at fourth cleavage. At the blastula stage, the 32 cells of this lineage detach from the epithelium and ingress into the blastocoel by a process of epithelial-mesenchymal transition. We report that shortly before ingression,there is a transient and highly localized activation of the MAP-kinase ERK in the micromere lineage. We show that ingression of the PMCs requires the activity of ERK, MEK and Raf, and depends on the maternal Wnt/β-catenin pathway. Dissociation experiments and injection of mRNA encoding a dominant-negative form of Ras indicated that this activation is probably cell autonomous. We identified the transcription factors Ets1 and Alx1 as putative targets of the phosphorylation by ERK. Both proteins contain a single consensus site for phosphorylation by the MAP kinase ERK. In addition, the Ets1 protein sequence contains a putative ERK docking site. Overexpression of ets1 by injection of synthetic mRNA in the egg caused a dramatic increase in the number of cells becoming mesenchymal at the blastula stage. This effect could be largely inhibited by treating embryos with the MEK inhibitor U0126. Moreover, mutations in the consensus phosphorylation motif substituting threonine 107 by an aspartic or an alanine residue resulted respectively in a constitutively active form of Ets1 that could not be inhibited by U0126 or in an inactive form of Ets1. These results show that the MAP kinase pathway, working through phosphorylation of Ets1, is required for full specification of the PMCs and their subsequent transition from epithelial to mesenchymal state.

Growth factor-mediated mesodermal cell guidance and skeletogenesis during sea urchin gastrulation

“…Bradham et al (Bradham et al, 2004) showed that inhibiting PI3K blocks the elongation of skeletal rods, but not PMC specification, cell migration, the extension of filopodia, cell fusion or skeletal initiation. The MAPK pathway, by contrast, regulates early PMC specification, thereby affecting all aspects of PMC morphogenesis (Fernandez-Serra et al, 2004;Röttinger et al, 2004). It has been suggested that MAPK signaling may regulate skeletogenesis by two distinct pathways: a Ras-independent pathway necessary for PMC ingression and a Ras-dependent pathway required for PMC migration and skeletogenesis (Fernandez-Serra et al, 2004;Röttinger et al, 2004).…”

“…The MAPK pathway, by contrast, regulates early PMC specification, thereby affecting all aspects of PMC morphogenesis (Fernandez-Serra et al, 2004;Röttinger et al, 2004). It has been suggested that MAPK signaling may regulate skeletogenesis by two distinct pathways: a Ras-independent pathway necessary for PMC ingression and a Ras-dependent pathway required for PMC migration and skeletogenesis (Fernandez-Serra et al, 2004;Röttinger et al, 2004). As the VEGF pathway does not regulate PMC ingression, it might function by a Ras/MAPK-dependent mechanism.…”

Manganese: A New Emerging Contaminant in the Environment

“…It has been widely demonstrated that Extracellular signal-Regulated Kinase (ERK) MapK mediated signalling controls the expression of several regulatory genes which participate in the specification and differentiation of the sea urchin skeleton (Fernandez-Serra et al, 2004;Röttinger et al, 2004). Thus, we analyzed, by Western blotting, the activation of ERK in manganese exposed embryos during development.…”

Human lysyl-tRNA synthetase is secreted to trigger proinflammatory response

“…5 A and B). To determine the functional significance of ERK and p38 MAPK for the KRS signaling, we pretreated the cells with U0126 that inhibits MEK, the upstream kinase for ERK, and the SB compounds that specifically block p38 MAPK (37,38). The KRS-dependent activation of ERK and p38 MAPK was blocked with these chemical inhibitors (Fig.…”