Oral–aboral axis specification in the sea urchin embryo

Abstract

In sea urchin embryos, specification of the secondary (oral-aboral) axis occurs via nodal, expression of which is entirely zygotic and localized to prospective oral ectoderm at blastula stage. The initial source of this spatial anisotropy is not known. Previous studies have shown that oral-aboral (OA) polarity correlates with a mitochondrial gradient, and that nodal activity is dependent both on mitochondrial respiration and p38 stress activated protein kinase. Here we show that the spatial pattern of nodal activity also correlates with the mitochondrial gradient, and that the latter correlates with inhomogeneous levels of intracellular reactive oxygen species. To test whether mitochondrial H2O2 functions as a redox signal to activate nodal, zygotes were injected with mRNA encoding either mitochondrially-targeted catalase, which quenches mitochondrial H2O2 and down-regulates p38, or superoxide dismutase, which augments mitochondrial H2O2 and up-regulates p38. Whereas the former treatment inhibits the initial activation of nodal and entrains OA polarity toward aboral when confined to half of the embryo via 2-cell stage blastomere injections, the latter does not produce the opposite effects. We conclude that mitochondrial H2O2 is rate-limiting for the initial activation of nodal, but that additional rate-limiting factors, likely also involving mitochondria, contribute to the asymmetry in nodal expression.

Oxygen, pH, and oral-aboral axis specification in the sea urchin embryo

“…3). Both mitochondrial and cytoplasmic H 2 O 2 were reduced by this treatment, with a decrease in mitochondrial H 2 O 2 comparable in magnitude to that produced by a mitochondrially-targeted catalase (Mt-Cat) shown previously to both inhibit nodal activation and entrain oral-aboral polarity (Coffman et al 2009) (Fig. 1B).…”

“…The coverslip treatments were carried out as described in Coffman et al (2004). Preparation and microinjection of mRNA and image analyses were performed as described in Coffman et al (2009). Error bars depict the standard deviation; ANOVA followed by Dunnett's test was used to determine the indicated significance values (JMP version 8.0; Cary, NC).…”

“…The oral-aboral axis of the sea urchin embryo is specified via the redox-regulated activation of nodal (Coffman et al, 2009, and references therein). Embryos cultured at high density under cover glass develop a radialized phenotype owing to inhibition of nodal expression (Coffman et al 2004), an effect that we attributed to hypoxia.…”

“…This is probably attributable at least in part to decreased H 2 O 2 levels, as the window of vulnerability corresponds to the initial H 2 O 2 -dependent activation phase of nodal expression (Supplemental Figure 1; Coffman et al, 2009). Extracellular acidification appears to play an ancillary role, perhaps weakening ligand-receptor interactions and thus by dampening the feedback amplification of nodal expression.…”

Opposing Nodal/Vg1 and BMP signals mediate axial patterning in embryos of the basal chordate amphioxus

“…In sea urchins, dispersal of a concentration of mitochondria localized on the oral side of the embryo, where Nodal is normally expressed, disrupts oral/aboral polarity. Moreover, quenching mitochondrial H 2 O 2 also inhibits Nodal activation, which is required for specification of oral identity, although increasing H 2 O 2 does not activate Nodal (Coffman et al 2009). To clarify a possible relationship between mitochondria, Nodal and Vg1 mRNAs in amphioxus it would be useful in the future to determine if Nodal and Vg1 proteins co-localize with their respective mRNAs and with their receptors and how localization of these proteins correlates with localization of mitochondria.…”