Structure, regulation, and function of micro1 in the sea urchin Hemicentrotus pulcherrimus

Abstract

The animal-vegetal axis of sea urchin embryos is morphologically apparent at the 16-cell stage, when the mesomeres, macromeres, and micromeres align along it. At this stage, the micromere is the only autonomously specified blastomere that functions as a signaling center. We used a subtraction PCR survey to identify the homeobox gene micro1 as a micromere-specific gene. The micro1 gene is a representative of a novel family of paired-like class homeobox genes, along with PlHbox12 from Paracentrotus lividus and pmar1 from Strongylocentrotus purpuratus. In the present study, we showed that micro1 is a multicopy gene with six or more polymorphic loci, at least three of which are clustered in a 30-kb region of the genome. The micro1 gene is transiently expressed during early cleavage stages in the micromere. Recently, nuclear beta-catenin was shown to be essential for the specification of vegetal cell fates, including micromeres, and the temporal and spatial coincidence of micro1 expression with the nuclear entry of beta-catenin is highly suggestive. We demonstrated that micro1 is a direct target of beta-catenin. In addition, we showed that micro1 is necessary and sufficient for micromere specification. These observations on the structure, regulation, and function of micro1 lead to the conclusion that micro1 and pmar1 (and potentially PlHbox12) are orthologous.

Activation of the skeletogenic gene regulatory network in the early sea urchin embryo

“…Considerable evidence indicates that pmar1 is a pivotal regulator of the micromere-PMC GRN. Misexpression of pmar1 is sufficient to activate the skeletogenic GRN in non-micromere-derived cells (Oliveri et al, 2002;Oliveri et al, 2003;Nishimura et al, 2004;Yamazaki et al, 2005;Yamazaki et al, 2009) (this study). It has been more difficult to test directly the function of pmar1 in the large micromere territory (where the gene is ordinarily expressed) owing to the difficulty in blocking the expression of multiple tandem pmar1 genes using MOs.…”

Par6 regulates skeletogenesis and gut differentiation in sea urchin larvae

“…Modified pBluescript RN3 (pBluescript RN3'; Nishimura et al 2004) was used to make expression constructs. For rescue experiments, a truncated construct lacking the 21 bp following the initiation methionine codon was generated by inverse-PCR, using Hp-par6 in pBluescript RN3' (Shiomi and Yamaguchi 2008) and KOD FX DNA polymerase (TOYOBO).…”

Lessons from a gene regulatory network: echinoderm skeletogenesis provides insights into evolution, plasticity and morphogenesis

“…One very unusual feature of the pmar1 gene may be related to its relatively recent recruitment into the PMC GRN; in all three species of euechinoid urchins that have been examined, the pmar1 locus consists of several (at least ten) nearly identical, tandem copies of the gene (Nishimura et al, 2004;Ettensohn et al, 2007;Sea Urchin Genome Sequencing Consortium, 2006). This suggests that recent duplications of the pmar1 gene might have been associated with its shift in developmental function.…”

“…It was shown previously that activation of the network depends upon maternally derived components of the canonical Wnt signaling pathway, a pathway that plays a key role in early axis specification in diverse animal phyla (reviewed by Ettensohn, 2006). β-Catenin is an essential early activator of the micromere-PMC GRN and is likely to act exclusively and directly via activation of pmar1 (paired-class micromere anti-repressor 1), which encodes a transcriptional repressor of the homeodomain family (Kitamura et al, 2002;Oliveri et al, 2002;Oliveri et al, 2003;Nishimura et al, 2004;Yamazaki et al, 2005). Although β-catenin protein is present throughout the vegetal region of the embryo during early cleavage, it activates pmar1 only in the micromeres, by mechanisms that are not understood.…”