Drosophila NinaB and NinaD Act Outside of Retina to Produce Rhodopsin Chromophore

Abstract

The Drosophila ninaB gene encodes a ␤,␤-carotene-15,15 -oxygenase responsible for the centric cleavage of ␤-carotene that produces the retinal chromophore of rhodopsin. The ninaD gene encodes a membrane receptor required for efficient use of ␤-carotene. Despite their importance to the synthesis of visual pigment, we show that these genes are not active in the retina. Mosaic analysis shows that ninaB and ninaD are not required in the retina, and exclusive retinal expression of either gene, or both genes simultaneously, does not support rhodopsin biogenesis. In contrast, neuron-specific expression of ninaB and ninaD allows for rhodopsin biogenesis. Additional directed expression studies failed to identify other tissues supporting ninaB activity in rhodopsin biogenesis. These results show that nonretinal sites of NinaB ␤,␤-carotene-15,15 -oxygenase activity, likely neurons of the central nervous system, are essential for production of the visual chromophore. Retinal or another C 20 retinoid, not members of the ␤-carotene family of C 40 carotenoids, are supplied to photoreceptors for rhodopsin biogenesis.Mutants in eight Drosophila genes, designated ninaA through ninaH, are characterized by reduced rhodopsin levels in photoreceptors and altered electroretinograms (1). The opsin protein component of rhodopsin is coded by the ninaE gene (2, 3). The low rhodopsin phenotypes observed in other nina mutants are caused by deficits in the post-translational rhodopsin maturation process. For example, ninaA encodes a molecular chaperone required for movement of newly synthesized rhodopsin from the endoplasmic reticulum to the photosensitive rhabdomeric membranes (4, 5). In the case of ninaB and ninaD, defective rhodopsin production is the result of a failure to generate the chromophore of rhodopsin, 3-OH retinal (6).Animal species usually obtain retinals in their food. Plants and microorganisms produce C 40 carotenoids such as ␤-carotene and zeaxanthin, which animals metabolize to C 20 retinoids. In Drosophila, the ninaD gene encodes a membrane "scavenger" receptor proposed to mediate the cellular uptake of carotenoids (7). The ninaB gene encodes a ␤,␤-carotene-15,15Ј-oxygenase (BCO) 1 responsible for the centric cleavage of ␤-carotene to form retinal (8). This enzyme was originally named as a ␤-carotene dioxygenase. It has now been renamed BCO (9) in light of recent data showing related enzymes act as monooxygenases (10).In vertebrates, vitamin A is essential for development and differentiation processes as well as its role in vision. The availability of vitamin A for metabolic processes is governed by multiple factors, including dietary absorption, transport, metabolism, and storage (11). A human BCO is expressed in the retinal pigment epithelium and also in the kidney, intestine, liver, brain, stomach, and testis (12, 13), suggesting that the processing of dietary carotenoids occurs in a variety of vertebrate tissues. Additional studies show that centric cleavage of ␤-carotene plays a major role in the processing of carote...

Phototransduction and retinal degeneration in Drosophila

“…The question arises as to the identity of the scavenger receptor that may function in concert with NINAB. Although NINAD has been suggested to play such a role [80], the different expression patterns of ninaB and ninaD make this hypothesis unlikely [78]. Rather, santa maria and ninaB are expressed and function in the same extraretinal glia and neuronal cells in fly heads.…”

The Drosophila ninaG Oxidoreductase Acts in Visual Pigment Chromophore Production

“…The gel was blotted onto Hybond-XL nylon transfer membrane (Amersham Biosciences) using the manufacturer's instructions, probed with Rh1 cDNA, random-labeled with [␣-32 P]dCTP using the Megaprime DNA labeling system (Amersham Biosciences) and autoradiographed to visualize Rh1 RNA levels. FRT/ FLP (flippase recombinase/flippase recombinase target) genetic mosaics were constructed and analyzed as described previously (8).…”

“…The ninaB encodes an oxygenase enzyme (7) responsible for cleaving ␤-carotene to generate retinal. Both of these gene products act outside of the retina to carry out their roles in vitamin A metabolism (8).…”

Rhodopsin Formation inDrosophilaIs Dependent on the PINTA Retinoid-Binding Protein

“…1 B) and ninaB (one allele). The ninaE locus encodes the structural gene for the Rh1 opsin (O'Tousa et al, 1985;Zuker et al, 1985), and ninaB encodes a ␤,␤-carotene-15,15Ј-dioxgenase required external to the retina for biogenesis of the chromophore (von Lintig et al, 2001;Gu et al, 2004). The remaining mutation, referred to as pinta (Fig.…”

“…One enzyme that functions in the transformation of the retinal to the chromophore is an oxidoreductase encoded by the neither inactivation nor afterpotential G (ninaG) locus (Sarfare et al, 2005). In addition, two genes (ninaB and ninaD) function outside the retina for production of vitamin A (Stephenson et al, 1983;Gu et al, 2004) and encode a ␤,␤-carotene-15,15Ј-dioxgenase and a class B scavenger receptor, respectively (von Lintig et al, 2001;Kiefer et al, 2002). However, the RBPs required for Drosophila vision have not been identified.…”