Spectral characteristics of invertebrate rhodopsins

2.4.1 Spectral absorption of rhodopsins and formation of photointermediates

Invertebrates exhibit a considerable variation in the types of different rhodopsins expressed by a species. Analysis of the genome of the soil-living nematode worm Caenorhabditis elegans suggests that it may not express a rhodopsin at all, while the visual system of a crustacean, the mantis shrimp Haptosquilla, might harbour of up to 16 distinct rhodopsins [5]. Rhodopsins with different absorption characteristics are generally found in invertebrates with eyes capable of colour vision. The compound eye of Drosophila, which expresses in total five rhodopsins, provides such a multi-input system, as do the eyes of bees, ants, butterflies, moths, and many other insects. Another rhodopsin, Rh2, is expressed in ocelli located at the vortex of the head. In Drosophila, the expression pattern of rhodopsins is correlated with a highly regular positioning of the photoreceptor cells within each ommatidium of the eye. The outer photoreceptor cells Rl-6 express the same rhodopsin (Rhl). The two central cells, R7 and R8, of the ommatidium join their individual rhabdomeres to form a rhabdomere located centrally in the open rhabdom (Figure 6). R7 and R8 cells appear to be specialized for colour vision. The central photoreceptors are functionally subdivided into two pairs which either express the combination Rh3/Rh5 or the combination Rh4/Rh6 (Figure 6). The developmental mechanisms underlying the type of rhodopsin patterning realized here is a topic of current research [100,101].

Figure 6. Expression pattern of rhodopsins in the Drosophila compound eye. The central panel indicates the expression pattern of the visual pigments in photoreceptor cells of the ommatidium. The absorption spectra of rhodopsins are shown in red, those of metarhodopsins in blue.

Figure 6. Expression pattern of rhodopsins in the Drosophila compound eye. The central panel indicates the expression pattern of the visual pigments in photoreceptor cells of the ommatidium. The absorption spectra of rhodopsins are shown in red, those of metarhodopsins in blue.

It has been already pointed out that the photointermediate sequence initiated by absorption of a photon by an invertebrate rhodopsin molecule terminates in a relatively long-lived metarhodopsin state. For the stable M-state of fly Rhl [102], as well as for other stable invertebrate metarhodopsins, this state exists in a pH-dependent equilibrium with an alkaline form of metarhodopsin [63,64], The shift in maximum absorption of acid M (565 nm) to alkaline M (about 380 nm) suggests that the retinylidene Schiff base becomes depro-tonated. However, none of the numerous microspectrophotometric measurements performed with different insect species provided any evidence for the in vivo formation of the alkaline metarhodopsin form. The intracellular pH, ion composition, protein-protein interactions, etc. apparently favour the formation of stable acid metarhodopsin. Investigations of the transition states, which are assumed by an invertebrate rhodopsin after photon absorption, suggest that primary events in the formation of the photointermediate bathorhodopsin are similar but not identical to those occurring in vertebrate rhodopsins [103]. At later steps the homology in transition states is even less obvious. In cephalopods, a spectrally distinct late intermediate, named mesorhodopsin, is formed prior to the formation of stable acid metarhodopsin; however, the complete sequence of transition states that finally leads to the formation of long-lived metarhodopsin has not been determined. Analysis of time-resolved transient grating signals in Octopus, which were shown to represent chromo-phore-independent protein dynamics, has revealed the transition of the late intermediate mesorhodopsin into a transient form of acid metarhodopsin. Transient acid M itself is transformed into stable acid metarhodopsin in a spectrally silent transition with a time constant of 180 (is [104]. It is assumed that in cephalopods this transient acid M represents the G-protein activating state of rhodopsin [62].

As a result of the formation of stable metarhodopsin, irradiation of invertebrate photoreceptors always establishes an equilibrium between rhodopsin and metarhodopsin. Accordingly, invertebrate rhodopsins are characterized sufficiently only by indicating the absorption of rhodopsin as well as the absorption of its stable M-state. Both rhodopsin states constitute a photoconvertible system in which the amount of rhodopsin/metarhodopsin present is determined by the wavelength of incident light and the individual spectral characteristics of both states [17,63,64], The absorption properties of the rhodopsin/metarhodopsin systems of Drosophila (Figure 7) reveal some principles that hold also for other invertebrate rhodopsins. The absorbance coefficient of metarhodopsin is higher than that of the corresponding rhodopsin by a factor of up to 1.8 [82], The absorption maxima of metarhodopsins derived from rhodopsins absorbing below 500 nm are bathochromically shifted in relation to the absorption of the corresponing rhodopsins (Rhl, Rh3, Rh4, Rh5). Rhodopsins absorbing maximally at wavelengths above 500 nm, like Rh6, are converted into metarhodopsins with hypsochromically shifted absorption maxima [17]. In some cases, for example in cephalopods, the absorption spectra of rhodopsin and metarhodopsin more or less overlap [66]. Accordingly, two classes of rhodopsin/metarhodopsin systems exist, one in which metarhodopsin can be photoconverted into rhodopsin by 100% and one in which this is not the case. Whether this has consequences for the rhodopsin renewal mechanisms, in which the rhodopsin content of a photoreceptor membrane is maximized light-dependently, is not yet known. There is, however, evidence that the hypsochromically shifted M-state of Drosophila Rh6 does not accumulate in an amount comparable to that of the other types of rhodopsin expressed in the Drosophila eye [24]. This might indicate that, in the case of Rh6, less efficient photoregeneration is compensated by a more efficient mechanism of rhodopsin turnover or by chemical regeneration of rhodopsin.

2.4.2 Spectral tuning of rhodopsin and metarhodopsin

The rhodopsin/metarhodopsin systems of the compound eye of Drosophila and of the ocelli, cover a wavelength range of 350 nm from the red to the UV spectral range (Figure 7). The mechanism underlying spectral tuning of rhodopsins is a highly active research field of functional genomics. So far, this topic has been investigated and discussed primarily on the basis of information on the spectral absorption of vertebrate rhodopsins [29,105]. A primary mode to modulate spectral properties of rhodopsins is seen in the interaction of a

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