Drosophila Rhodopsin 7 can partially replace the structural role of Rhodopsin 1, but not its physiological function

Journal of Comparative Physiology A, May 2017

Rhodopsin 7 (Rh7), a new invertebrate Rhodopsin gene, was discovered in the genome of Drosophila melanogaster in 2000 and thought to encode for a functional Rhodopsin protein. Indeed, Rh7 exhibits most hallmarks of the known Rhodopsins, except for the G-protein-activating QAKK motif in the third cytoplasmic loop that is absent in Rh7. Here, we show that Rh7 can partially substitute Rh1 in the outer receptor cells (R1–6) for rhabdomere maintenance, but that it cannot activate the phototransduction cascade in these cells. This speaks against a role of Rh7 as photopigment in R1–6, but does not exclude that it works in the inner photoreceptor cells.

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Drosophila Rhodopsin 7 can partially replace the structural role of Rhodopsin 1, but not its physiological function

Drosophila Rhodopsin 7 can partially replace the structural role of Rhodopsin 1, but not its physiological function Rudi Grebler 0 1 2 3 Christa Kistenpfennig 0 1 2 3 Dirk Rieger 0 1 2 3 Joachim Bentrop 0 1 2 3 Stephan Schneuwly 0 1 2 3 Pingkalai R. Senthilan 0 1 2 3 Charlotte Helfrich‑Förster 0 1 2 3 0 Developmental Biology, Institute of Zoology, University of Regensburg , Regensburg , Germany 1 Cell- and Neurobiology, Zoological Institute, Karlsruhe Institute of Technology (KIT) , Karlsruhe , Germany 2 Neurobiology and Genetics, Biocenter, Theodor Boveri Institute, University of Würzburg , 97074 Würzburg , Germany 3 Present Address: Oxitec Ltd , 71 Innovation Drive, Milton Park, Oxford OX14 4RQ , UK Rhodopsin 7 (Rh7), a new invertebrate Rhodopsin gene, was discovered in the genome of Drosophila melanogaster in 2000 and thought to encode for a functional Rhodopsin protein. Indeed, Rh7 exhibits most hallmarks of the known Rhodopsins, except for the G-protein-activating QAKK motif in the third cytoplasmic loop that is absent in Rh7. Here, we show that Rh7 can partially substitute Rh1 in the outer receptor cells (R1-6) for rhabdomere maintenance, but that it cannot activate the phototransduction cascade in these cells. This speaks against a role of Rh7 as photopigment in R1-6, but does not exclude that it works in the inner photoreceptor cells. Phototransduction; Electroretinograms; Rhodopsins; Compound eyes; Drosophila melanogaster - Vision begins with the absorption of photons by visual pigment molecules. Rhodopsins are membrane-bound G-protein-coupled receptors that absorb photons, undergo conformational changes, and activate a G-protein to initiate visual signal transduction. Animal genomes typically contain multiple Rhodopsin genes coding for Rhodopsins with different spectral properties providing the basis for color vision. Each photoreceptor cell usually expresses a single rhodopsin, but exceptions are known in both vertebrates and invertebrates (Applebury et al. 2000; Hu et al. 2011, 2014; Mazzoni et al. 2004; Stavenga and Arikawa 2008). The fruit fly Drosophila melanogaster possesses six different well-characterized Rhodopsin molecules, Rh1 to Rh6. With the exception of Rh2, all Rhodopsins are found in the receptor cells of the compound eyes: Rh1 is expressed in the six outer receptor cells (R1–6) of each eye unit and Rh3 to Rh6 are expressed in the two inner receptor cells (R7, R8) (reviewed in Rister et al. 2013; Behnia and Desplan 2015). A seventh Rhodopsin, Rh7, of still unknown location and function was predicted from the genome in 2000 (Adams et al. 2000; Terakita 2005). qPCR studies showed that Rh7 is expressed at low levels in the compound eyes, suggesting that it may be co-expressed with one or several of the other Rhodopsins (Posnien et al. 2012; Senthilan and Helfrich-Förster 2016). The aim of the present study was to investigate a potential function of Rh7 in the compound eyes. A suited method to reveal the properties of an unknown Rhodopsin is to express it in R1–6 instead of Rh1 (Feiler et al. 1988; 1992; Townson et al. 1998; Salcedo et al. 1999; Knox et al. 2003; Hu et al. 2014). Rh1 is required for proper rhabdomere morphogenesis and maintenance, in addition to its role as photopigment (O’Tousa et al. 1985; Kumar and Ready 1995; Kumar et al. 1997; Zuker et al. 1985). Thus, loss of Rh1 (in ninaE17 mutants) leads to the collapse of rhabdomeric microvilli inside the photoreceptor cytoplasm (Ahmad et al. 2007; Bentrop 1998; Kurada and O’Tousa 1995; Leonard et al. 1992). This can be prevented by expressing other functional Rhodopsins in R1–6 of ninaE17 mutants (Kumar et al. 1997). Here, we expressed Rh7 instead of Rh1 under the Rh1 promotor (Rh1–Rh7;ninaE17 flies) and investigated whether Rh7 can (1) rescue the retinal degradation provoked by the ninaE 17 mutation and (2) lead to normal electroretinogram (ERG) responses. We also expressed Rh7 in addition to Rh1 (Rh1–Rh7 flies) to see whether this increases the ERG responses. Materials and methods Wild-type CantonS (WTCS) as well as flies with yellow body color and white eyes (yellow− white1118 = y− w1118) served as control for semi-thin sections and immunocytochemistry. For qPCR, deep pseudopupil, and ERG measurements, only flies in the y− w1118 background were used. In addition, ninaE17 and sevLY3 mutants were in the y− w1118 background. ninaE (=neither inactivation nor afterpotential E) codes for Rh1 and y− w1118; ninaE17 mutants are Rh1 null mutants (O’Tousa et al. 1985). sev (sevenless) codes for a tyrosin kinase that is critical for the development of the photoreceptor cell R7 (Basler and Hafen 1988) and y− w1118 sevLY3 mutants lack the inner photoreceptor cell R7 (Harris et al. 1976). In the following, we will omit “y− w1118” and simply use ninaE17 and sevLY3. Generation of Rh1–Rh7 transgenic flies To generate flies expressing the Rh7 coding region under control of the Rh1 promotor, the full-length Rh7 CDS was amplified by (...truncated)


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Rudi Grebler, Christa Kistenpfennig, Dirk Rieger, Joachim Bentrop, Stephan Schneuwly, Pingkalai R. Senthilan, Charlotte Helfrich-Förster. Drosophila Rhodopsin 7 can partially replace the structural role of Rhodopsin 1, but not its physiological function, Journal of Comparative Physiology A, 2017, pp. 1-11, DOI: 10.1007/s00359-017-1182-8