Evolutionary convergence in Otx expression in the pentameral adult rudiment in direct-developing sea urchins.

Convergence is a significant evolutionary phenomenon. Arrival at similar morphologies from different starting points indicates a strong role for natural selection in shaping morphological phenotypes. There is no evidence yet of convergence in the developmental mechanisms that underlie the evolution of convergent developmental phenotypes. Here we report the expression domains in sea urchins of two important developmental regulatory genes ( Orthodenticle and Runt), and show evidence of molecular convergence in the evolution of direct-developing sea urchins. Indirect development is ancestral in sea urchins. Evolutionary loss of the feeding pluteus stage and precocious formation of the radially symmetric juvenile has evolved independently in numerous sea urchin lineages, thus direct development is an evolutionary convergence. Indirect-developing species do not express Otx during the formation of their five primordial tube feet, the ancestral condition. However, each direct-developing urchin examined does express Otx in the tube feet. Otx expression in the radial arms of direct-developing sea urchins is thus convergent, and may indicate a specific need for Otx use in direct development, a constraint that would make direct development less able to evolve than if there were multiple molecular means for it to evolve. In contrast, Runt is expressed in tube feet in both direct- and indirect-developing species. Because echinoderms are closely related to chordates and postdate the protostome/deuterostome divergence, they must have evolved from bilaterally symmetrical ancestors. Arthropods and chordates use Otx in patterning their anterior axis, and Runt has multiple roles including embryonic patterning in arthropods, and blood and bone cell differentiation in vertebrates. Runt has apparently been co-opted in echinoderms for patterning of pentamery, and Otx in pentameral patterning among direct-developing echinoids. The surprisingly dynamic nature of Otx evolution reinvigorates debate on the role of natural selection vs shared ancestry in the evolution of novel features.

Epithelial cell wedging and neural trough formation are induced planarly in Xenopus, without persistent vertical interactions with mesoderm.

In this study we investigate the induction of the cell behaviors underlying neurulation in the frog, Xenopus laevis. Although planar signals from the organizer can induce convergent extension movements of the posterior neural tissue in explants, the remaining morphogenic processes of neurulation do not appear to occur in absence of vertical interactions with the organizer (R. Keller et al. , 1992, Dev. Dyn. 193, 218-234). These processes include: (1) cell elongation perpendicular to the plane of the epithelium, forming the neural plate; (2) cell wedging, which rolls the neural plate into a trough; (3) intercalation of two layers of neural plate cells to form one layer; and (4) fusion of the neural folds. To allow planar signaling between all the inducing tissues of the involuting marginal zone and the responding prospective ectoderm, we have designed a "giant sandwich" explant. In these explants, cell elongation and wedging are induced in the superficial neural layer by planar signals without persistent vertical interactions with underlying, involuted mesoderm. A neural trough forms, and neural folds form and approach one another. However, the neural folds do not fuse with one another, and the deep cells of these explants do not undergo their normal behaviors of elongation, wedging, and intercalation between the superficial neural cells, even when planar signals are supplemented with vertical signaling until the late midgastrula (stage 11.5). Vertical interactions with mesoderm during and beyond the late gastrula stage were required for expression of these deep cell behaviors and for neural fold fusion. These explants offer a way to regulate deep and superficial cell behaviors and thus make possible the analysis of the relative roles of these behaviors in closing the neural tube.

Surface mesoderm in Xenopus: a revision of the stage 10 fate map.

We have used two complementary cell labeling techniques to investigate dorsal mesoderm formation in Xenopus laevis and Hymenochirus boettgeri. Epithelial grafts from fluorescently labeled donors into unlabeled hosts demonstrate that in Xenopus, as previously shown for Hymenochirus, superficial cells of the dorsal marginal zone have the ability to invade the notochord and somite and participate in their normal morphogenesis, in a stage-specific and region-specific manner. A new method for superficial fate mapping using cell surface biotinylation confirms this result for Hymenochirus and demonstrates that in Xenopus as well, even in normal development in the absence of surgical disruption, notochord and the most posterior somitic mesoderm originate partly in the superficial epithelial layer. This finding is contrary to the widespread belief that Xenopus mesoderm originates solely in the deep mesenchymal layer. In Xenopus (but not in Hymenochirus), the amount of superficial contribution to mesoderm varies, such that in some spawnings it appears not to be present, while in others it is evident in all or most embryos.

Dorsal mesoderm has a dual origin and forms by a novel mechanism in Hymenochirus, a relative of Xenopus.

The dorsal mesoderm in the frog Hymenochirus forms by a mechanism not previously described in any other vertebrate. Unlike its close relative, Xenopus laevis, in which the mesoderm derives entirely from the deep mesenchymal cells of the marginal zone, Hymenochirus has "surface mesoderm" originating in the involuting marginal zone epithelium. Fluorescently labeled grafts show stage-specific invasion of deep axial tissue by cells originally located in the surface layer. These cells participate in normal mesoderm development. In video recordings, the labeled surface area shrinks as surface cells invade the deep layer. Furthermore, the mechanism of surface mesoderm morphogenesis differs from that described in other amphibians. Scanning electron microscopy at several neurula stages indicates that prospective somite cells do not individually detach from their epithelial neighbors to ingress into the deep layer, as seen in other amphibians; instead, their basal ends adhere to the somitic mesoderm as a coherent layer, taking on somitic morphology while still a part of the archenteron lining. This novel morphogenetic process we dub "relamination." Prospective notochord cells individually spread on the ventral surface of the notochord, gradually ingressing from their epithelial neighbors, but by a mechansism involving active pulling and spreading by their invasive basal ends rather than depending on apical constriction as do the corresponding "bottle cells" in other amphibians. Lateral endoderm migrates dorsally, replacing the relaminating surface mesoderm and fusing at the dorsal midline of the archenteron. These processes demonstrate the diversity of morphogenesis at the cellular, and presumably the molecular, level and shed light on the evolution of morphogenetic mechanisms.

Notochord morphogenesis in Xenopus laevis: simulation of cell behavior underlying tissue convergence and extension.

Cell intercalation and cell shape changes drive notochord morphogenesis in the African frog, Xenopus laevis. Experimental observations show that cells elongate mediolaterally and intercalate between one another, causing the notochord to lengthen and narrow. Descriptive observations provide few clues as to the mechanisms that coordinate and drive these cell movements. It is possible that a few rules governing cell behavior could orchestrate the shaping of the entire tissue. We test this hypothesis by constructing a computer model of the tissue to investigate how rules governing cell motility and cell-cell interactions can account for the major features of notochord morphogenesis. These rules are drawn from the literature on in vitro cell studies and experimental observations of notochord cell behavior. The following types of motility rules are investigated: (1) refractory tissue boundaries that inhibit cell motility, (2) statistical persistence of motion, (3) contact inhibition of protrusion between cells, and (4) polarized and nonpolarized protrusive activity. We show that only the combination of refractory boundaries, contact inhibition and polarized protrusive activity reproduces normal notochord development. Guided by these rules, cells spontaneously align into a parallel array of elongating cells. Self alignment optimizes the geometric conditions for polarized protrusive activity by progressively minimizing contact inhibition between cells. Cell polarization, initiated at refractory tissue boundaries, spreads along successive cell rows into the tissue interior as cells restrict and constrain their neighbors' directional bias. The model demonstrates that several experimentally observed intrinsic cell behaviors, operating simultaneously, may underlie the generation of coordinated cell movements within the developing notochord.