Transient receptor potential (TRP) ion channels are common components of mechanosensing pathways, mainly described in mammals and other multicellular organisms. To gain insight into the evolutionary origins of eukaryotic mechanosensory proteins, we investigated the involvement of TRP channels in mechanosensing in a unicellular eukaryotic protist, the dinoflagellate Lingulodinium polyedra. BLASTP analysis of the protein sequences predicted from the L. polyedra transcriptome revealed six sequences with high similarity to human TRPM2, TRPM8, TRPML2, TRPP1, and TRPP2; and characteristic TRP domains were identified in all sequences. In a phylogenetic tree including all mammalian TRP subfamilies and TRP channel sequences from unicellular and multicellular organisms, the L. polyedra sequences grouped with the TRPM, TPPML, and TRPP clades. In pharmacological experiments, we used the intrinsic bioluminescence of L. polyedra as a reporter of mechanoresponsivity. Capsaicin and RN1734, agonists of mammalian TRPV, and arachidonic acid, an agonist of mammalian TRPV, TRPA, TRPM, and Drosophila TRP, all stimulated bioluminescence in L. polyedra. Mechanical stimulation of bioluminescence, but not capsaicin-stimulated bioluminescence, was inhibited by gadolinium (Gd3+), a general inhibitor of mechanosensitive ion channels, and the phospholipase C (PLC) inhibitor U73122. These pharmacological results are consistent with the involvement of TRP-like channels in mechanosensing by L. polyedra. The TRP channels do not appear to be mechanoreceptors but rather are components of the mechanotransduction signaling pathway and may be activated via a PLC-dependent mechanism. The presence and function of TRP channels in a dinoflagellate emphasize the evolutionary conservation of both the channel structures and their functions.
Keywords: AA, amino acids; AMTB hydrochloride, N-(3-Aminopropyl)-2-[(3-methylphenyl)methoxy]-N-(2-thienylmethyl)benzamide hydrochloride; Ce, Caenorhabditis elegans; Cr, Chlamydomonas reinhardtii; DMSO, dimethyl sulfoxide; Dm, Drosophila melanogaster; Dr, Danio rerio; FSW, filtered seawater; Gd3+, gadolinium; GsMTx4, Grammostola spatulata mechanotoxin 4; HC067047, 2-Methyl-1-[3-(4-morpholinyl)propyl]-5-phenyl-N-[3-(trifluoromethyl)phenyl]-1H-pyrrole-3-carboxamide; HMM, Hidden Markov Model; Hs, Homo sapiens; Lp, Lingulodinium polyedra; ML204, 4-Methyl-2-(1-piperidinyl)-quinoline; Mb, Monosiga brevicollis; ORF, open reading frame; PIP2, Phosphatidylinositol 4,5-bisphosphate; PLC, phospholipase C; Pt, Paramecium tetraurelia; RHC80267, O,O′-[1,6-Hexanediylbis(iminocarbonyl)]dioxime cyclohexanone; RN1734, 2,4-Dichloro-N-isopropyl-N-(2-isopropylaminoethyl)benzenesulfonamide; RN1747, 1-(4-Chloro-2-nitrophenyl)sulfonyl-4-benzylpiperazine; TMHMM, transmembrane helix prediction; TRP, transient receptor potential channel; U73122, 1-[6-[((17β)-3-Methoxyestra-1,3,5[10]-trien-17-yl)amino]hexyl]-1H-pyrrole-2,5-dione.