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. 2011 Oct 9;479(7371):108-12.
doi: 10.1038/nature10451.

Feedback From Rhodopsin Controls Rhodopsin Exclusion in Drosophila Photoreceptors

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Free PMC article

Feedback From Rhodopsin Controls Rhodopsin Exclusion in Drosophila Photoreceptors

Daniel Vasiliauskas et al. Nature. .
Free PMC article

Abstract

Sensory systems with high discriminatory power use neurons that express only one of several alternative sensory receptor proteins. This exclusive receptor gene expression restricts the sensitivity spectrum of neurons and is coordinated with the choice of their synaptic targets. However, little is known about how it is maintained throughout the life of a neuron. Here we show that the green-light sensing receptor rhodopsin 6 (Rh6) acts to exclude an alternative blue-sensitive rhodopsin 5 (Rh5) from a subset of Drosophila R8 photoreceptor neurons. Loss of Rh6 leads to a gradual expansion of Rh5 expression into all R8 photoreceptors of the ageing adult retina. The Rh6 feedback signal results in repression of the rh5 promoter and can be mimicked by other Drosophila rhodopsins; it is partly dependent on activation of rhodopsin by light, and relies on G(αq) activity, but not on the subsequent steps of the phototransduction cascade. Our observations reveal a thus far unappreciated spectral plasticity of R8 photoreceptors, and identify rhodopsin feedback as an exclusion mechanism.

Figures

Figure 1
Figure 1. Rh6 acts to repress Rh5 expression in yR8 PRs
a: Genomic rh6 locus. The promoter region sufficient to drive rh6 expression in yR8 is in blue, exons are in green and mutations in red. In rh6fs mutants, 58bp of the promoter are deleted. In rh61 mutants, 21bp at the first exon-intron junction are replaced with AA, leading to an immediate truncation of the ORF. b: Percentage of R8 PRs expressing Rh5 as a function of time (days post-eclosion) in wild-type (blue) and rh61 mutants (red). Error bars represent 84% Confidence Intervals. c-g: Whole mount retina stained with specific antibodies for Rh5 (blue) and Rh6 (red). c, c’: Normal expression of Rh5 and Rh6 in 2 week old flies. c’ Shows Rh5 alone. d, e: In rh61 mutants, Rh5 is gradually de-repressed. At eclosion, retinas have a normal number of Rh5-expressing R8s (d). By 2 weeks post-eclosion, most R8s express Rh5 (e). f, g: rh6fs promoter mutation leads to loss of detectable Rh6 expression in almost all yR8s. As in rh61 mutants (d,e), at eclosion rh6fs retinas have a normal number of Rh5-expressing R8s (f), but by 2 weeks post-eclosion, most R8 express Rh5 (g).
Figure 2
Figure 2. Rh6 represses transcription of the rh5 gene
a-d: rh5 mRNA, detected by in situ hybridization in transverse cryo-sections of 3 week old fly eyes. Many more cells are expressing rh5 mRNA in the R8 layer of rh6 mutants (b) as compared to wild-type flies (a). In sev mutants, very few cells express rh5 (c). However, in sev; rh6 double mutants, rh5 is extensively de-repressed in R8 PRs (d). e, f: In 3 week old control flies, a rh5 reporter (rh5>GFP) (green) is expressed in pR8s that also express Rh5 protein (blue), but not in yR8 cells which express Rh6 (red) (e). In rh6 mutants, rh5>GFP is de-repressed in most yR8 cells (f).
Figure 3
Figure 3. Mutation of rh6 does not lead to change in yR8 cell identity
a, b: A rh6-lacZ reporter (red) is expressed normally in rh6 mutants. It is induced in a pattern complementary to the expression of Rh5 (blue) in young flies (a). In 2 week old rh6 mutants, Rh5 expression expands into the lacZ positive, yR8 cells (b). c-f: Z-projections of confocal stacks encompassing nuclei and Rh-containing rhabdomeres of R8 PRs. c, d: Expression of the nuclear pR8 marker melt-nlacZ (green) does not change in rh6 mutants. It is normally expressed together with Rh5 (blue) in pR8 and never in Rh6-expressing yR8 cells (red) (c). In 5 week old rh6 mutants, melt-nlacZ is not de-repressed along with Rh5 and remains restricted to pR8 (d). e, f: Expression of nuclear yR8 marker wts-nlacZ does not change in rh6 mutants. It is normally expressed together with Rh6 (red) in yR8 and never in Rh5-expressing pR8 cells (blue) (e). In rh6 mutants, wts-nlacZ remains in yR8 of 4 week old flies as Rh5 is de-repressed (f).
Figure 4
Figure 4. Part of the phototransduction pathway is required to maintain repression of Rh5
a, b: Forced expression of Rh4 (red) in yR8 with rh6-Gal4 in rh61 mutants prevents Rh5 (blue) de-repression (b) observed in rh61 mutant flies (a). c: Forced expression of Rh5 (blue) in yR8 with rh6-Gal4 in rh61 mutants prevents rh5>GFP (green) de-repression observed in rh61 mutant flies (compare to Fig. 2f and Supplementary Fig. 7f). d-e: Dark-reared flies partially de-repress Rh5 in yR8 PRs. In the light, wt flies do not de-repress Rh5 (blue) in Rh6-expressing yR8s (red) (d). After 2-3 weeks in complete darkness, a significant number of yR8s of wt flies express low levels of Rh5 in addition to Rh6 (arrowheads, e). d, e show close ups of dorsal retinas, just dorsal to the equator. f, g: Gαq is required to maintain repression of Rh5 in yR8. In 2-3 week old (g), but not in just eclosed (f) Gαq1 mutants, Rh5 (blue) is expressed in yR8 and thus is co-expressed (arrowheads) with Rh6 (red). b’, d’-g’: Rh5 expression alone as in b, d-g.

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