Semaphorin 3 C and Netrin-1 , Are Chemorepellents in Tetrahymena Thermophila

During vertebrate development, neuronal growth is guided by chemical signals. Chemoattractants encourage neuronal growth cones to form, while chemorepellents cause growth cone collapse. Integration of these signals allows for proper neural positioning in the developing organism. Not all signaling pathways are clear-cut, however. Netrin-1, for example, can function as either a chemoattractant or a chemorepellent, depending upon the cell type involved, the signals the cell has previously received, and the concentration of the signal. Netrin-1 primarily signals through a G-protein mediated receptor via the adenylyl cyclase pathway. The semaphorins mainly serve as chemorepellents and as immune signals in vertebrates. However, signaling through the semaphorins is not well understood. Tetrahymena thermophila are free-living, ciliated eukaryotic organisms that are often used as a model for showing behavioral responses to both chemoattractants and chemorepellents. A number of polycationic peptides function as chemorepellents in this organism, including lysozyme, VIP, PACAP, nociceptin, substance P, and ACTH derivatives. In the current study, we used two polycationic peptides derived from the vertebrate neuronal chemorepellents, semaphorin 3C and netrin-1, in order to determine whether they were chemorepellents in Tetrahymena thermophila. Both peptides were chemorepellents in Tetrahymena thermophila. Semaphorin 3C peptide showed chemorepellent activity with an EC100 of approximately 10 μM. Netrin-1 peptide showed chemorepellent activity over a range of concentrations from micromolar to nanomolar. However, there was a great deal of variability in the response to netrin-1. Further characterization of the pathways involved in the signaling of these repellents will allow comparison between ciliate and vertebrate signaling, and may help us better understand vertebrate development. Introduction Materials and Methods Behavioral assays were conducted using a dissection microscope, a 3-well microtiter plate, and a modified Pasteur pipette as described in Mace et al., 2000, and as pictured below.