In situ screening for genes expressed preferentially in secondary mesenchyme cells of sea urchin embryos

Vegetal plate specification was assessed in S. purpuratus embryos after micromere deletions at the 4th, 5th and 6th cleavages, by assaying expression of the early vegetal plate marker Endo 16, using whole-mount in situ hybridization. After 4th cleavage micromere deletions, the embryos typically displayed weak Endo16 expression in relatively few cells of the lineages that normally constitute the vegetal plate, while after 5th and 6th cleavage micromere deletions the embryos exhibited strong Endo16 expression in larger fractions of cells belonging to those lineages. When all four micromeres were deleted, the embryos were severely delayed in initiating gastrulation and sometimes failed to complete gastrulation. However, if only one micromere was allowed to remain in situ throughout development, the embryos exhibited strong Endo16 expression and gastrulation occurred normally, on schedule with controls. Additional measurements showed that these microsurgical manipulations do not alter cleavage rates or generally disrupt embryo organization. These results constitute direct evidence that the micromeres provide signals required by the macromere lineages for initiation of vegetal plate specification. The specification of the vegetal plate is completed in a normal manner only if micromere signaling is allowed to continue at least to the 6th cleavage stage.

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Lectin uptake and incorporation into the calcitic spicule of sea urchin embryos

Zygote ◽

10.1017/s0967199414000094 ◽

2014 ◽

Vol 23 (3) ◽

pp. 467-473 ◽

Cited By ~ 2

Author(s):

Nancy M. Mozingo

Keyword(s):

Sea Urchin ◽

Wheat Germ ◽

Matrix Proteins ◽

Fluorescent Label ◽

Sea Urchin Embryos ◽

Intracellular Vesicles ◽

Mesenchyme Cells ◽

Fluorescent Tag ◽

Larval Skeleton ◽

Membranous Material

SummaryPrimary mesenchyme cells (PMCs) are skeletogenenic cells that produce a calcareous endoskeleton in developing sea urchin larvae. The PMCs fuse to form a cavity in which spicule matrix proteins and calcium are secreted forming the mineralized spicule. In this study, living sea urchin embryos were stained with fluorescently conjugated wheat germ agglutinin, a lectin that preferentially binds to PMCs, and the redistribution of this fluorescent tag was examined during sea urchin development. Initially, fluorescence was associated primarily with the surface of PMCs. Subsequently, the fluorescent label redistributed to intracellular vesicles in the PMCs. As the larval skeleton developed, intracellular granular staining diminished and fluorescence appeared in the spicules. Spicules that were cleaned to remove membranous material associated with the surface exhibited bright fluorescence, which indicated that fluorescently labelled lectin had been incorporated into the spicule matrix. The results provide evidence for a cellular pathway in which material is taken up at the cell surface, sequestered in intracellular vesicles and then incorporated into the developing spicule.

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