ИСТИНА

Phototaxis (oriented movement towards or away from the light source) in Chlamydomonas reinhardtii involves an all-trans retinal-containing photoreceptor (chlamyrhodopsin). Its photoexcitation gives rise to a cascade of electrical currents across the cell membrane, which results in an increase in intracellular Ca2+ concentration and alteration of flagellar beating. Chlamyrhodopsin-mediated photocurrents and phototactic orientation were measured in suspensions of carotenoid-deficient Chlamydomonas mutants upon reconstitution with all-trans 9-desmethyl-retinal (9-dmr) or retinal. In parallel, photoelectric responses of individual reconstituted cells were studied by the suction pipette technique in a hybrid strain that carried deficiencies in both retinal biosynthesis and cell wall structure. The addition of 9-dmr resulted in a complete reconstitution of the photoreceptor protein with similar time course and concentration dependence, as that of retinal. However, the amplitude of the photoreceptor current (PC) measured in 9-dmr-reconstituted cells was about two times smaller and the rise time longer, as compared to cells reconstituted with retinal, indicating a different signal transduction mechanism for 9-dmr-chlamyrhodopsin. Restoration of PC measured in double-flash experiments was faster in 9-dmr-reconstituted cells than in those reconstituted with retinal, which showed that substitution of 9-dmr instead of the native chromophore did not lead to an extension of the chlamyrhodopsin photocycle to hundreds of milliseconds, as it had been found in arhaebacterial sensory rhodopsins. In contrast to the wild type, mutant cells reconstituted by either 9-dmr or retinal swam away from the light source at low fluence rates of the actinic light (negative phototaxis) and switched to positive phototaxis, when the fluence rate exceeded a certain threshold. The reversal of the phototaxis direction in 9-dmr-reconstituted cells was observed at higher stimulus intensity than that in cells reconstituted with retinal, which could be explained by the decreased PC found in the former cells.