A search for new-type photoreceptors in unicellular flagellate algae and true slime mold.

点击翻译按钮获取中文摘要

In general, an attempt to clone homologs of plant or bacterial photoreceptors in fungi or in unicellular flagellates is unsuccessful because of their genetical diversity. We therefore combined several new PCR strategies in order to search for true archaeal-type rhodopsin phototaxis receptor(s) that had been physiologically well-established but had not been detected in molecular level.Expressed Sequence Tag (EST) database of Chlamydomonas reinhardtii publicized in March 2000 from Kazusa DNA Research Institute contained a partial cDNA sequence that showed a little similarity in amino acid level to those of F-G helices of bacterial opsins. Using temperature-asymmetric PCR in combination with a highly degenerated primer, we have extended this sequence toward N-terminus using Chlamydomonas genomic DNA as a template. We found a reasonable similarity in the PCR product to DNA sequences corresponding to C-G helices of bacterial opsins. The complete DNA sequence was obtained and deposited in GenBank (accession numbers: AB058890 and AB58891). Using indirect immunofluorescence we also demonstrated the localization of one of these new archaeal-type opsins of Chlamydomonas in a cell-surface region close to the eyespot apparatus by which Chlamydomonas phototaxis is mediated. A homology modeling successfully indicated an archaeal-type folding of the proteins, accounting for previously established stereospecificity of retinal isomers in the phototaxis receptor(s).