Expression of Pigment Cell-Specific Genes in the Ontogenesis of the Sea Urchin Strongylocentrotus intermedius

Abstract

One of the polyketide compounds, the naphthoquinone pigment echinochrome, is synthesized in sea urchin pigment cells. We analyzed polyketide synthase (pks) and sulfotransferase (sult) gene expression in embryos and larvae of the sea urchin Strongylocentrotus intermedius from various stages of development and in specific tissues of the adults. We observed the highest level of expression of the pks and sult genes at the gastrula stage. In unfertilized eggs, only trace amounts of the pks and sult transcripts were detected, whereas no transcripts of these genes were observed in spermatozoids. The addition of shikimic acid, a precursor of naphthoquinone pigments, to zygotes and embryos increased the expression of the pks and sult genes. Our findings, including the development of specific conditions to promote pigment cell differentiation of embryonic sea urchin cells in culture, represent a definitive study on the molecular signaling pathways that are involved in the biosynthesis of pigments during sea urchin development.

Figure 1

The structure of the naphthoquinone pigment echinochrome (a). One of the quinone biosynthesis pathways (the formation of chimaphilin from a shikimic acid through the mevalonic acid biosynthetic pathway) in accordance with [9] (b).

Figure 2

Expression of the Sipks (a) and Sisult (b) genes in spermatozoids (Sp), unfertilized eggs (Egg), coelomocytes (Cel), ambulacra (Ambl), embryos, and larvae of the sea urchin S. intermedius at various stages of development: blastula, 14 hpf (Bl), gastrula, 24 hpf (Gl), prism, 34 hpf (Pr), and pluteus, 72 hpf (Pl). *P < .05; **P < .01.

Figure 3

Effect of shikimic acid (ShA) on the expression of the Sipks (a) and Sisult (b) genes in zygotes, blastula, and gastrula cells of S. intermedius. The time of incubation with ShA is 8 days (all embryos of the control group (C) were at the pluteus stage). ShA concentrations tested: 1—0.1 mM, 2— 0.5 mM. *P < .05.

Figure 4

Normal embryo development of the sea urchin S. intermedius: zygotes (20 min pf, Zy); blastula stage (14 hpf, Bl); gastrula stage (24 hpf, Gl); (d) prism stage (34 hpf, Pr); (e) pluteus (8 dpf, Pl). Nomarski's optics. Bar, 50 μm.

Figure 5

Effect of shikimic acid (ShA) on the larval morphology of the sea urchin S. intermedius. Disturbances in embryo development after 8 days of incubation with ShA. ShA was added to (a) zygotes; (b) embryos of the blastula stage; (c) embryos of the gastrula stage. ShA concentrations tested: 1–0.1 mM, 2–0.5 mM, and 3–2.0 mM. Nomarski's optics. Bar, 50 μm.

Figure 6

Embryonic pigment cells in a blastula-derived cell culture of the sea urchin S. intermedius. (a) Multilayer cell sheets (2-3 days of cultivation in seawater supplemented with 2% fetal bovine serum); (b) spread pigment cells cultivated in the coelomic fluid of control sea urchins for 3 days; (c) rounded pigment cells cultivated in the coelomic fluid of injured sea urchins for 3 days. Bar, 10 μm.