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The cell-cycle checkpoint protein p53 both directs terminal differentiation and protects embryos from DNA damage. To study invertebrate p53 during early development, we identified three differentially expressed p53 family members (p53, p97, p120) in the surf clam, Spisula solidissima. In these mollusks, p53 and p97 occur in both embryonic and adult tissue, whereas p120 is exclusively embryonic. We sequenced, cloned, and characterized p120 cDNA. The predicted protein, p120, resembles p53 across all evolutionarily conserved regions and contains a C-terminal extension with a sterile alpha motif (SAM) as in p63 and p73. These vertebrate forms of p53 are required for normal inflammatory, epithelial, and neuronal development. Unlike clam p53 and p97, p120 mRNA and protein levels are temporally expressed in embryos, with mRNA levels decreasing with increasing p120 protein (R(2) = 0.97). Highest surf clam p120 mRNA levels coincide with the onset of neuronal growth. In earlier work we have shown that neuronal development is altered by exposure to polychlorinated biphenyls (PCBs), a neurotoxic environmental contaminant. In this study we show that PCBs differentially affect expression of the three surf clam p53 family members. p120 mRNA and protein are reduced the most and earliest in development, p97 protein shows a smaller and later reduction, and p53 protein levels do not change. For the first time we report that unlike p53 and p97, p120 is specifically embryonic and expressed in a time-dependent manner. Furthermore, p120 responds to PCBs by 48 hr when PCB-induced suppression of the serotonergic nervous system occurs.

Polychlorinated biphenyls (PCBs) are ubiquitous environmental pollutants that accumulate to toxic levels in the food chain. Using Spisula solidissima (surf clam) embryos as a developmental model, it was shown that Aroclor 1254 specifically targets two neuronal structures during embryonic development. Embryos were exposed to 1, 10), or 100 ppm Aroclor 1254 or an acetone vehicle control posthatching for 24, 48, and 72 h. Embryos labeled with a serotonin antibody or a neural antigen antibody and a rhodamine-conjugated secondary antibody were viewed by confocal microscopy. The cerebropleural ganglion showed a decrease both in serotonin production and in the size of the serotonin-synthesizing region upon exposure to 10 and 100 ppm Aroclor 1254. These decreases were detectable as early as 48 h postfertilization. When exposed to 100 ppm Aroclor 1254, the primitive neural plexus, which coordinates the movements of the mouth and velum, showed a delay in onset and cessation of expression of a molluscan-specific neural antigen. Exposure to Aroclor 1254 did not affect the overall growth and morphology of the embryos. In addition, analyses of total protein profiles and heat-shock protein 70 levels showed that exposure to Aroclor 1254 did not trigger protein degradation or cause a stress or shock response. These results show that exposure of Spisula embryos to Aroclor 1254 specifically targets neurogenesis while having no effect on the overall growth of the embryo.

Polychlorinated biphenyls (PCBs) are ubiquitous environmental contaminants that exert neurotoxic effects during embryonic development. The present study demonstrates that early embryonic exposure to a mixture of PCBs (Aroclor 1254) results in a decrease in serotonergic cell growth. Using a novel, marine invertebrate embryo model, Spisula solidissima, immunocytochemistry, and confocal microscopy techniques, a dose-dependent decrease in serotonergic cell number was quantified within 24 h of exposure. This effect was seen with doses as low as 1 ppm Aroclor 1254. These findings demonstrate that environmentally relevant doses of Aroclor 1254 impair development of the serotonergic nervous system.

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