Oligosaccharide chains of newly synthesized membrane receptors are trimmed and modified to optimize their trafficking and/or signalling before delivery to the cell surface. For most membrane receptors, the functional significance of oligosaccharide chain modification is unknown. During the maturation of Rh1 rhodopsin, a Drosophila light receptor, the oligosaccharide chain is trimmed extensively. Neither the functional significance of this modification nor the enzymes mediating this process are known. Here, we identify a dmppe (Drosophila metallophosphoesterase) mutant with incomplete deglycosylation of Rh1, and show that the retained oligosaccharide chain does not affect Rh1 localization or signalling. The incomplete deglycosylation, however, renders Rh1 more sensitive to endocytic degradation, and causes morphological and functional defects in photoreceptors of aged dmppe flies. We further demonstrate that the dMPPE protein functions as an Mn 2 þ /Zn 2 þ -dependent phosphoesterase and mediates in vivo dephosphorylation of a-Man-II. Most importantly, the dephosphorylated a-Man-II is required for the removal of the Rh1 oligosaccharide chain. These observations suggest that the glycosylation status of membrane proteins is controlled through phosphorylation/dephosphorylation, and that MPPE acts as the phosphoesterase in this regulation.

Background: Appropriate termination of photoresponse is critical for photoreceptors to achieve high temporal resolution and to prevent excessive Ca 2ϩ -induced cell toxicity. Results: We isolated a novel G␣ q mutant allele and revealed that metarhodopsin/G q interaction affects Arr2-Rh1 binding. Conclusion: G q modulates the termination of phototransduction and prevents retinal degeneration. Significance: Our study revealed the novel role of G q in phototransduction deactivation and in retinal degeneration.Appropriate termination of the phototransduction cascade is critical for photoreceptors to achieve high temporal resolution and to prevent excessive Ca 2؉ -induced cell toxicity. Using a genetic screen to identify defective photoresponse mutants in Drosophila, we isolated and identified a novel G␣ q mutant allele, which has defects in both activation and deactivation. We revealed that G q modulates the termination of the light response and that metarhodopsin/G q interaction affects subsequent arrestin-rhodopsin (Arr2-Rh1) binding, which mediates the deactivation of metarhodopsin. We further showed that the G␣ q mutant undergoes light-dependent retinal degeneration, which is due to the slow accumulation of stable Arr2-Rh1 complexes. Our study revealed the roles of G q in mediating photoresponse termination and in preventing retinal degeneration. This pathway may represent a general rapid feedback regulation of G protein-coupled receptor signaling.Heterotrimeric G proteins play pivotal roles in mediating extracellular signals from hormones, neurotransmitters, peptides, as well as sensory stimuli to intracellular signaling pathways (1, 2). In Drosophila photoreceptors, G proteins are essential for the activation of the phototransduction cascade (3, 4). Photon absorption leads to the photoisomerization of chromophores, resulting in the formation of activated metarhodopsin. In turn, metarhodopsin activates heterotrimeric G proteins and PLC.2 The activation of PLC leads to transient receptor potential and transient receptor potential-like channels opening and extracellular Ca 2ϩ influx (5-8). It is also critical for each step of the phototransduction cascade to be terminated appropriately, which is essential for the high temporal resolution of fly vision (9, 10). The most important step in phototransduction termination is the deactivation of metarhodopsin. During this step, arrestin (Arr2) plays an important role by displacing the G q ␣ subunit and allowing it to bind with rhodopsin (Rh1) (11,12). Unlike other G proteincoupled receptors (GPCRs), the phosphorylation of fly rhodopsin is not required for its deactivation (13) but is essential for its endocytosis (11). In contrast, the dephosphorylation of rhodopsin by retinal degeneration C (RDGC) is essential for receptor deactivation (14). Ca 2ϩ also plays critical roles in regulating the termination of the photoresponse in Drosophila (8,15,16). Several proteins that mediate this Ca 2ϩ -regulated termination have been identified, such as eye-specific protein kinase C...

Heterotrimeric G proteins play central roles in many signaling pathways, including the phototransduction cascade in animals. However, the degree of involvement of the G protein subunit Gαq is not clear since animals with previously reported strong loss-of-function mutations remain responsive to light stimuli. We recovered a new allele of Gαq in Drosophila that abolishes light response in a conventional electroretinogram assay, and reduces sensitivity in whole-cell recordings of dissociated cells by at least five orders of magnitude. In addition, mutant eyes demonstrate a rapid rate of degeneration in the presence of light. Our new allele is likely the strongest hypomorph described to date. Interestingly, the mutant protein is produced in the eyes but carries a single amino acid change of a conserved hydrophobic residue that has been assigned to the interface of interaction between Gαq and its downstream effector, PLC. Our study has thus uncovered possibly the first point mutation that specifically affects this interaction in vivo.