A proline-rich peptide originating from decomposing mangrove leaves is one natural metamorphic cue of the tropical jellyfish Cassiopea xamachana 'Lehrstuhl fiir Spezielle Zoologie, Ruhr-Universitit. D-44780 Bochum. Germany 'Institute of Ecology, University of Georgia, Athens. Georgia 30602-2602. USA 3Complex Carbohydrate Research Center, University of Georgia, Athens. Georgia 30602-4712, USA ABSTRACT: Planula larvae of the scyphozoan Cassiopea xamachana settle and n~etamorphose on degrading mangrove leaves of Rhizophora mangle that lie submerged In shallow water mangrove ecosystems. Our prior study (Fleck & Fitt 1999; J Exp Mar Biol Ecol 234:83-94) indicated that marine bacteria are involved in the release of at least 1 peptidic compound from such leaves. The goal of our present study was to isolate and purify at least 1 natural peptidic cue originating from deteriorating leaves by means of ultrafiltration, gel filtration and reversed phase HPLC and subsequently obtain characteristic data of this cue. The ultrafiltrate (110 kD) of the homogenate of decaying mangrove leaves was subjected to gel filtration on a Sephadex G 25 column, resulting in 3 fractions which were tested for their capacity to induce metamorphosis of planula larvae in bioassays performed in the laboratory. Fraction I ( 2 5 kD) was most effective in inducing metamorphosis of 75 % of planulae at 1 mg freeze-dried material ml-' seawater within 24 h. Fractions I1 and I11 (both 25 kD) resulted In metamorphosis of only 1 % of larvae or less within 72 h when applied at 5 mg rnl-' Isochratic HPLC separation of Fraction I with 24 % methanol yielded 2 biologically active fractions. One fraction (A/B), which induced 41 % of the larvae to metamorphose at 0.9 mg lyophilized material ml-' seawater within 24 h, consisted of a mixture of at least 2 subfractions and was not further analyzed. The other fraction (C) effected metamorphosis of 85 % of larvae at a concentration of 0.5 mg rnl-' within 24 h. Matrix-assisted laser desorption ionization (MALDI) mass spectrometry of this fraction revealed a molecular weight of approximately 5.8 kD. Automated amino acid analysis showed that Fraction C was rich in proline (ca 44%) and glycine residues (ca 16%), corresponding to characteristic proline-rich cell wall proteins of plants. Automated sequencing of the natural inducer failed due to a blocked amino terminus. The results of our present study suggest that metamorphic inducers for C. xamachana may emerge nonspecifically as a byproduct of bacterial degradation of deteriorating, proteinaceous plant tissue in their habitat.
Larvae-like vegetative buds of the scyphozoan Cassiopea andromeda metamorphose into polyps in the presence of oligopeptides that have a well-defined primary structure. Buds were incubated with the hexapeptide 14C-dansyl-GPGGPA, a representative inducer. Autoradiography of longitudinal sections of these buds revealed rapid internalization of peptide by the buds. Silver grain density was highest in the pre-pedal disc region (or aboral knob) of metamorphosing buds. Larvae and buds sporadically explore their habitat with this aboral knob, searching for a suitable solid substrate to which irreversible attachment will be made. Buds were incubated for 3, 8, or 16 h with 14C-dansyl-GPGGPA, then homogenized and the supernatants analyzed to determine the chemical fate of the inducer. The signal molecule was shown to be partly degraded to 14C-dansyl-GP, partly to 14C-dansyl-G, and in part still present in its original structure. These cleavage products were also found in the surrounding medium after an incubation time of 8 h with 14C-dansyl-GPGGPA, but did not induce metamorphosis. This study suggests that exposure of metamorphosis-inducing peptides to buds of Cassiopea andromeda results in signal termination.
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