BACTERIORHODOPSIN IS INVOLVED IN HALOBACTERIAL PHOTORECEPTION

The bacterio-opsin gene was introduced into a ''blind'' Halobacterium salinarium mutant that (i) lacked an the four retinal proteins [bacter iorhodopsin (BR), halorhodopsin, and sensory rhodopsins (SRs) I and II ] and the transducer protein for SRI and (ii) showed neither attractan t response to long wavelength light nor repellent response to short wa velength light. The resulting transformed cells acquired the capabilit y to sense light stimuli. The cells accumulated in a light spot, demon strating the BR-mediated orientation in spatial light gradients. As in wild-type cells, a decrease in the intensity of long wavelength light caused a repellent response by inducing reversals of swimming directi on, but, in contrast to wild-type cells, a decrease in the intensity o f short wavelength light also repelled the cells. An increase in fight intensity evoked an attractant response (i.e., a transient suppressio n of spontaneous reversals). Signal processing times and adaptation ki netics were similar to the SRI-mediated reactions. However, compared t o SR-mediated photoresponses, higher light intensities were necessary to induce the BR-mediated responses. The light sensitivity of the tran sformant was increased by adding 1 mM cyanide and decreased by the add ition of arginine, agents that respectively reduce and increase the li ght-independent generation of the electrochemical potential difference of H+ ions (DELTAmu(H+)BAR). A decrease in irradiance to an intensity that was still high enough to saturate BR-initiated DELTAmu(H+)BAR, c hanges failed to induce the repellent effect, but the addition of a pr otonophorous uncoupler sensitized the cell to these light stimuli. The BR D96N mutant (Asp-96 is replaced by Asn) with decreased proton pump activity showed strongly reduced BR-mediated responses. Azide, which increases this mutant's H+ pump efficiency, increased the photosensiti vity of the mutant cells. Moreover, azide diminished (i) the membrane potential decreasing and (ii) repellent effects of blue light added to the orange background illumination in this mutant. We conclude that t he BR-mediated photoreception is due to the light-dependent generation of DELTAmu(H+)BAR. Our data are consistent with the assumption that t he H. salinarium cell monitors the membrane energization level with a ''protometer'' system measuring total DELTAmu(H+)BAR changes or its el ectric potential difference component.