Mesoderm As a Primary Driver of Segmental Patterning in Leeches
In Chapter 4, segmentation was defined as the existence of metameric units in two or more germ layers, with the added requirement that the spatial frequency of the metamers be conserved across the germ layers. This definition raises the question of how spacing of the units in one germ layer is matched to that in the other germ layer(s). One obvious solution to this problem is to have the metameric repeats in one germ layer serve to organize those in the other layer(s).
During cleavage, the four bilateral pairs of ectoteloblasts (N, О/P, О/P, and Q) arise from a bilateral pair of precursor blastomeres, the left and right NOPQ cells. Moreover, as previously described, the segmental mesoderm on each side arises from a single ipsilateral stem cell, the M teloblast. Numerous experiments have documented the fact that leech embryos are essentially lacking in their capacity for regulative replacement of ablated lineages (e.g., Blair and Weisblat 1982, 1984; Stuart et al. 1987, 1989; Torrence et al. 1989). Thus, it was possible to ask if mesoderm is required for segmentation in ectoderm and vice versa by experiments in which individual cells were injected either with fluorescent lineage tracers or cell-restricted killing ablatants, e.g., cytotoxic enzymes such as DNAse I (Blair 1982). When an NOPQ proteloblast was killed on one side of the embryo, thereby removing the ectoderm from that side, two different outcomes were observed. In some embryos, the two m bandlets remained on their respective sides; in this case, the mesodermal bandlet that had no overlying ectodermal bandlets was severely disorganized and failed to generate segments, while the contralateral side developed normally. In other embryos, the m bandlet on the ablated side switched sides, so that both m bandlets were in contact w'ith the remaining ectoderm; in this case, both m bandlets formed somites, but the left and right sides were often out of register, indicating that this process occurs independently on the right and left sides.
When the converse experiments were carried out, killing the mesodermal precursor on one side, the ipsilateral ectodermal blast cells initially exhibited their normal, lineage-specific division patterns (Zackson 1984; Huang and Weisblat 1996) but then became disorganized and failed to generate recognizable segmental structures (Blair 1982). Thus, in Helobdella, ectoderm and mesoderm exhibit a mutual interdependence in generating segmental organization of those germ layers.
Other experiments have revealed a requirement for mesodermal signaling in patterning the endodermal midgut as well. Endodermal nuclei arising by cellu- larization of the SYC nuclei express lox3, a parahox gene of the xloxlpdx family (Wedeen and Shankland 1997). Prospective endodermal nuclei also express another homeobox gene loxlO (of the nk-2 gene family), although the exact timing of the onset of this expression relative to the cellularization of SYC nuclei to form definitive endoderm remains to be determined (Nardelli-Haefliger and Shankland 1993). After labeling the M lineage on one side of the embryo with a photosensitizing lineage tracer (fluorescein dextran), it was possible to photolesion nascent mesodermal somites. In lesioned segments, no lox3 expression was detected, and the endodermal cells were completely absent from precisely the regions from which the normally overlying mesoderm was absent (Wedeen and Shankland 1997). Thus, it appears that short-range signaling from visceral mesoderm is required for the migration, differentiation, and/or survival of segmentally patterned midgut endoderm.