Identification of Reissner's fiber-like glycoproteins in two species of freshwater planarians (Tricladida), by use of specific polyclonal and monoclonal antibodies

Abstract

By using one polyclonal antiserum raised against bovine Reissner's fiber and seven monoclonal antibodies raised against bovine Reissner's fiber and against immunopurified bovine subcommissural organ glycoproteins, we have investigated two freshwater planarian species (Girardia tigrina, Schmidtea mediterranea) by light- and electron-microscopic immunocytochemistry. ELISA probes showed that the monoclonal antibodies recognized different, nonoverlapping, unrepeated, proteinaceous epitopes present in the same compounds of bovine Reissner's fiber. Cells immunoreactive to the polyclonal and monoclonal antibodies were found in the dorsal and ventral integument of both planarian species. Labeled cuboid epidermal cells bore cilia and displayed several types of secretory granules; they were covered by a film of immunoreactive material. Studies on adjacent thin and semithin sections revealed coexistence of label in the same regions and in the same cells when two different monoclonal antibodies were used. These results indicate that a secretory substance immunologically similar to the secretion of the vertebrate subcommissural organ is present in primitive tripoblasts such as planarians, suggesting that these secretions are ancient and well conserved in phylogeny.

The central nervous system of sea cucumbers (Echinodermata: Holothuroidea) shows positive immunostaining for a chordate glial secretion

“…Our data suggest that the RS-like material can possibly carry out a variety of functions within a single organism, since it is present in such different tissues as the epidermis and the nervous tissue. These findings coincide with other reports demonstrating the presence of RS and its major component, SCO-spondin, both in the nervous system (e.g., [ 11 , 19 ]) and in non-neural tissues [ 36 , 45 ]. Sequence analysis suggests that SCO-spondin is a multidomain protein with a very complex structure [ 23 , 41 ], which can be involved in a wide range of processes mediated via protein/protein interactions.…”

“…The important issue in any immunocytochemical study is the specificity of recognition of the epitope by the primary antibody. The two polyclonal rabbit antisera used in the present study were raised against Reissner's fiber isolated from the bovine spinal cord and a wealth of subsequent studies has convincingly demonstrated that both antisera specifically recognized Reissner's substance (RS) in a wide range of animal species from mammals to planarians [ 11 , 19 , 20 , 31 , 36 , 39 , 40 ]. Besides some minor differences in immunolabelling in the hyponeural part of the radial nerve cord, both antibodies used in the present study show similar staining patterns with different visualization techniques employed (peroxidase-anti-peroxidase approach, immunofluorescence, immunoelectron microscopy).…”

“…These findings, in turn, imply that the RS-like material does play some important role(s) both in the echinoderm nervous systems and the epidermis. The function of RS in the epidermis is completely unknown [ 36 ]. As to the nervous system, RS has been shown to be involved in axonal guidance, as well as in promoting neuronal survival, differentiation, and aggregation in vertebrates [ 24 , 30 , 41 ].…”

“…In planarians, for instance, strongly immunoreactive secretory cells are located in the epidermis. Certain vertebrate epidermal cells are also reported to be positively labeled with anti-RS antibodies [ 36 ]. Although the ectoneural cords of the sea star belong to the central nervous system, they are actually still a part of the epidermis.…”