ИСТИНА

In modern developmental biology, the organizer is defined as “an embryonic region capable of inducing a secondary axis when transplanted to an ectopic site”. It is very common for embryonic organizers to be associated with the blastopore region. In general, blastopore-associated organizer is a complex signaling center that is responsible to maintain BMP and Wnt signaling gradients regulating the establishment of primary body axes. The obvious question is whether we can trace back the blastoporal organiser governing axial patterning to the more basal metazoan phyla. We hope that the study of the blastoporal organizer of a representative of the basal animal phylum Cnidaria, Nematostella vectensis will help us to shed light on the evolutionary origin of the embryonic organizers. To determine the precise localization of the zone having the organizing capacity, we have performed several types of transplantation experiments at the gastrula stage. Small tissue fragments taken from different regions of the vitally stained gastrulae have been transplanted into the blastocoel roof of recipient embryos. We found that only the blastopore lip fragment was able to efficiently induce the outgrowth of a secondary axis in a host embryo (Kraus et al., 2007).To what extent do host tissues contribute to the secondary axial structures? Transplantation of fluorescently labeled donor tissues helps us to answer this question. Secondary body column consists from the both donor and host tissues. The grafted blastopore lip fragment always obtained an apical position in the outgrowing secondary axis. Positive outcomes of transplantation experiments are very variable. We have obtained the formation of secondary structures upon transplantation only in 60% of cases. Many of these embryos formed an elongated secondary body contained more or less complete set of internal and external morphological structures. However, some embryos formed very short secondary body (just an outgrowth of the host body) with only few secondary structures. It is possible that differences in the inductive capacities of the grafted tissue may be based on the unequal distribution of determinants along the circumference of the Nematostella blastopore. Results of the transplantations may also depend on the changes in the inductive capacities of the organizer region in the course of development. To determine the time frame of inductive activity of the Nematostella organizer, we have performed three series of transplantations. We transplanted blastopore lip fragments taken from the embryos at different stages of gastrulation (early gastrula, mid-gastrula, late gastrula). We found that the inductive capacity of the blastopore lip changes in the course of gastrulation and that the result of transplantation depends on the age of the host. The highest inductive capacity is observed at mid-gastrula stage. We also performed ectopic activation of the Wnt pathway by incubating the embryos in the solution of azakenpaullone, which is a specific inhibitor of GSK-3β. The outcomes of these experiments strongly depended on the age of embryos subjected to treatment. The treatment beginning at the blastula stage resulted in the formation of an oversized mouth at the oral pole of the embryo and multiple mouth-like openings on its aboral surface. Such embryos never produced primary polyps. When late gastrulae/planulae were subjected to treatment, they produced a secondary body axis containing a complete set of morphological structures. CONCLUSIONS 1) We have shown that morphological variability of the transplantation outcomes may reflect the asymmetric distribution of inductive capacities along the circumference of the Nematostella blastopore. 2) We found that the inductive capacity of the Nematostella blastopore lip changes in the course of gastrulation. 3) Our results suggest that before the mid-gastrula stage the aboral area of Nematostella embryo is highly responsive to the signals from the organizer. Later in development this area becomes much more restricted in its developmental capacities.