The fate of larval flagellated cells during metamorphosis of the sponge Halisarca dujardini

Sponge larval flagellated cells have been known to form the external layer of larva, but their subsequent fate and morphogenetic role are still unclear. It is actually impossible to follow flagellated cell developmental fate unless a specific marker is found. We used percoll density gradient fractionation to separate different larval cell types of Halisarca dujardini (Demospongiae, Halisarcida). A total of 5 fractions were obtained which together contained all cell types. Fraction 1 contained about 100% FC and its polypeptide composition was very different to that of the other fractions. Of all larval cell types, flagellated cells displayed the lowest in vitro aggregation capacity. We raised a polyclonal antibody against a 68 kDa protein expressed by larval flagellated cells. Its specificity was tested on total protein extract from adult sponges by Western blotting and proved to be suitable for immunofluorescence. By means of double immunofluorescence using both this polyclonal antibody and commercial anti-tubulin antibodies, we studied the distribution of the 68 kDa protein in larval flagellated cells and its fate at successive stages of metamorphosis. In juvenile sponges just after metamorphosis the choanocytes and the upper pinacoderm were labelled with both antibodies. In larval flagellated cells, the 68 kDa protein was found all over the cytoplasm appearing as granules, while in adult sponges, it was present in the apical part of choanocytes in the vicinity of collars. Direct participation of the larval flagellated cells in the development of definitive structures was demonstrated.