Cytochemical investigations on tunic morphogenesis in the sea peach Halocynthia papillosa (Tunicata, Ascidiacea) 2: demonstration of proteins.

In a previous paper, cellulose fibres were demonstrated in the larval, the metamorphosing, and the juvenile tunics. In this paper we used cytochemical methods and X-ray microanalysis to obtain additional information on tunic morphogenesis in Halocynthia papillosa. The chemical composition of the tunic evolves with its structural complexity. The larval and juvenile fibres are shown to be structurally and chemically different. While neither proteins nor glycosaminoglycans seem to be associated with the larval fibres, the juvenile fibres consist of a cellulose core wrapped in a sheath of tannophilic proteins. Patches of glycosaminoglycans line their longitudinal axes. In the course of metamorphosis, the cuticle undergoes profound modifications in regions of spine morphogenesis. Granular material that was previously called fibro-granular material (Lübbering et al., 1993) is essential to the formation of cuticular plates and spines. During metamorphosis, this material accumulates in epidermal granules and is discharged into the tunic. It crosses the fundamental layer of the tunic and reaches the cuticle. Our results strongly suggest that this material consists of proteins rich in cysteine and hydrophobic amino acids.

Cytochemical investigations on tunic morphogenesis in the sea peach Halocynthia papillosa (Tunicata, Ascidiacea). 1: demonstration of polysaccharides.

The distribution of carbohydrates was demonstrated in the embryonic, larval, and juvenile tunics of Halocynthia papillosa. An enzyme-gold marker (cellobiohydrolase-Au) was used to identify cellulose on ultrathin sections. This is the first time this biopolymer has been detected in the embryonic or larval tunic of an ascidian. Cellulose is present from the initial tail-bud stage onwards, as soon as the outer compartment of the tunic appears. Both compartments of the larval tunic also contain non-cellulosic polysaccharides, as demonstrated by the periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) method. Our observations point to two types of cellulose synthesis. One occurs during the embryonic and larval stages, when glycogen-like material is stored in epidermal intracellular lacunae and discharged into the tunic where it is presumably used to synthesize cellulose throughout the depth of the tunic. The second occurs from the onset of metamorphosis onwards, just above the apical plasmalemma of epidermal cells, like cellulose biogenesis in plants.