Antimicrobial peptides in sea urchins. Isolation, characterization and expression

Echinoderms are interesting animals for biodiscovery/bioprospecting as sources for novel compounds that have interesting activities. These compounds might be developed into potential drug candidates. In this work, two novel families of short proteins, referred to as antimicrobial peptides (AMPs) were isolated and characterized from the sea urchin, Strongylocentrotus droebachiensis. One of the functions of AMPs is that they act as antibiotics and thus have the ability to inhibit the growth of bacteria and other microorganisms. Even though sea urchins are evolutionary old animals, they have spikes and a hard shell that protect the internal organs suspended in coelomic fluid (blood). However, little is known about the functions and importance of the AMPs in the defence system of cells in adult sea urchins and larva. The fist group of AMPs that were detected were the cysteine-rich peptides, named strongylocins, isolated from extracts of the coelomocytes (blood cells). They have a novel cysteine arrangement pattern when compared to other peptides. The mature strongylocins have activity against both Gram-positive and Gram-negative bacteria. The amino acid sequences for the precursor strongylocins contain a signal peptide, a prosequence and a native region. Their gene sequences indicate that strongylocins (except strongylocin 1b) have three introns and four exons. Similar genes were found in the sister species, S. purpuratus. Recombinant products of these genes inhibited growth of bacteria by a nonlytic, but presumably intracellular mechanism. The other group of AMPs, named centrocins, are composed of two chains, one heavy chain and one light chain. The native peptides were very potent against bacteria. The gene sequences of centrocins also code for a signal peptide, two prosequences and a native region and contain one intron and two exons. A synthetic heavy chain had anti-fungi and anti-yeast activities in addition to being active against bacteria. Of the four types of coelomocytes, strongylocin 1 was detected in phagocytes and the vibratile/colourless spherule cells, strongylocin 2 was found in phagocytes and red spherule cells while centrocin 1 was found only in phagocytes and was located in the granular vesicles of the cells. Both the peptide families were detected in the important pluteus developmental stage (mid and late stage) of the sea urchin larvae. The distribution of centrocin 1 in the secondary mesenchyme cells around coelomic pouches and in some arms implies the importance of these molecules for immunity. All together, this work gives novel insight to molecules that are believed to be important for protecting the animals against microbial infections. These molecules are regarded as important players in the host defence systems of both adult animals and larva