Partial cloning of putative G-proteins modulating mechanotransduction in the ciliate stentor.
Signal transduction systems known to utilize G-proteins in higher eukaryotes undoubtedly evolved prior to the development of metazoa. Pharmacological evidence indicates that the ciliates Paramecium, Stentor, and Tetrahymena all utilize signaling systems similar to those found in mammals. However, there has been relatively little direct evidence for the existence of G-proteins in ciliates. Since highly conserved heterotrimeric G-proteins form the basis of receptor-coupled signal transduction systems in a wide variety of metazoa, it is of interest to know if these important signaling molecules were early to evolve and are present and functionally important in a wide variety of unicellular organisms. We have previously shown that mechanotransduction in Stentor is modulated by opiates in a manner that may involve pertussis toxin-sensitive G-proteins. Here we utilize drugs known to interact with G-proteins to further test for the involvement of these important signaling molecules in Stentor mechanotransduction. We present behavioral and electrophysiological data demonstrating that putative G-proteins in Stentor decrease mechanical sensitivity by modulating the mechanotransduction process. In addition, we report the partial cloning of 4 G-protein alpha-subunits from Stentor. We confirm that these clones are of Stentor origin and are transcribed. Furthermore, we employ antisense oligodeoxynucleotide-mediated knockout to demonstrate that these ciliate G-proteins exert a modulatory influence on Stentor behavior, and that a G1/G0-like clone mediates the inhibitory action of beta-endorphin on mechanotransduction.
Marino
M J
MJ
Department of Neuroscience, University of Pittsburgh, Pennsylvania 15260, USA.
Sherman
T G
TG
Wood
D C
DC
D000818:Animals; D016798:Ciliophora / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular / Q000379:methods*; D016054:DNA, Protozoan / Q000032:analysis; D004594:Electrophysiology; D019204:GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000031:analogs & derivatives* / Q000378:metabolism; D015151:Immunoblotting; D008465:Mechanoreceptors / Q000378:metabolism*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D015398:Signal Transduction
D016054:DNA, Protozoan; C034882:guanosine 5'-O-(1-thiotriphosphate); D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019204:GTP-Binding Proteins