The light-activated TRP channel: the founding member of the TRP channel superfamily.

The light-activated Transient Receptor Potential (TRP) channel is the founding member of a large and diverse family of channel proteins. The TRP (dTRP) channel, which generates the electrical response to light has been investigated in a great detail two decades before the first mammalian TRP channel was discovered. Thus, dTRP is unique among members of the TRP channel superfamily because its physiological role and the enzymatic cascade underlying its activation are established.

PDA (prolonged depolarizing afterpotential)-defective mutants: the story of nina's and ina's--pinta and santa maria, too.

Our objective is to present a comprehensive view of the PDA (prolonged depolarizing afterpotential)-defective Drosophila mutants, nina's and ina's, from the discussion of the PDA and the PDA-based mutant screening strategy to summaries of the knowledge gained through the studies of mutants generated using the strategy. The PDA is a component of the light-evoked photoreceptor potential that is generated when a substantial fraction of rhodopsin is photoconverted to its active form, metarhodopsin. The PDA-based mutant screening strategy was adopted to enhance the efficiency and efficacy of ERG (electroretinogram)-based screening for identifying phototransduction-defective mutants.

The nonsense-mediated decay pathway maintains synapse architecture and synaptic vesicle cycle efficacy.

A systematic Drosophila forward genetic screen for photoreceptor synaptic transmission mutants identified no-on-and-no-off transient C (nonC) based on loss of retinal synaptic responses to light stimulation. The cloned gene encodes phosphatidylinositol-3-kinase-like kinase (PIKK) Smg1, a regulatory kinase of the nonsense-mediated decay (NMD) pathway. The Smg proteins act in an mRNA quality control surveillance mechanism to selectively degrade transcripts containing premature stop codons, thereby preventing the translation of truncated proteins with dominant-negative or deleterious gain-of-function activities.

Why Drosophila to study phototransduction?

This review recounts the early history of Drosophila phototransduction genetics, covering the period between approximately 1966 to 1979. Early in this period, the author felt that there was an urgent need for a new approach in phototransduction research. Through inputs from a number of colleagues, he was led to consider isolating Drosophila mutants that are defective in the electroretinogram.

A spinosyn-sensitive Drosophila melanogaster nicotinic acetylcholine receptor identified through chemically induced target site resistance, resistance gene identification, and heterologous expression.

Strains of Drosophila melanogaster with resistance to the insecticides spinosyn A, spinosad, and spinetoram were produced by chemical mutagenesis. These spinosyn-resistant strains were not cross-resistant to other insecticides. The two strains that were initially characterized were subsequently found to have mutations in the gene encoding the nicotinic acetylcholine receptor (nAChR) subunit Dalpha6.

Heterozygous mutation of Drosophila Opa1 causes the development of multiple organ abnormalities in an age-dependent and organ-specific manner.

Optic Atrophy 1 (OPA1) is a ubiquitously expressed dynamin-like GTPase in the inner mitochondrial membrane. It plays important roles in mitochondrial fusion, apoptosis, reactive oxygen species (ROS) and ATP production. Mutations of OPA1 result in autosomal dominant optic atrophy (DOA).

Role of Ca2+/calmodulin-dependent protein kinase II in Drosophila photoreceptors.

Ca(2+) modulates the visual response in both vertebrates and invertebrates. In Drosophila photoreceptors, an increase of cytoplasmic Ca(2+) mimics light adaptation. Little is known regarding the mechanism, however.

DAG lipase activity is necessary for TRP channel regulation in Drosophila photoreceptors.

In Drosophila, a phospholipase C-mediated signaling cascade links photoexcitation of rhodopsin to the opening of the TRP/TRPL channels. A lipid product of the cascade, diacylglycerol (DAG) and its metabolite(s), polyunsaturated fatty acids (PUFAs), have both been proposed as potential excitatory messengers. A crucial enzyme in the understanding of this process is likely to be DAG lipase (DAGL).

Presynaptic calcium channel localization and calcium-dependent synaptic vesicle exocytosis regulated by the Fuseless protein.

A systematic forward genetic Drosophila screen for electroretinogram mutants lacking synaptic transients identified the fuseless (fusl) gene, which encodes a predicted eight-pass transmembrane protein in the presynaptic membrane. Null fusl mutants display >75% reduction in evoked synaptic transmission but, conversely, an approximately threefold increase in the frequency and amplitude of spontaneous synaptic vesicle fusion events. These neurotransmission defects are rescued by a wild-type fusl transgene targeted only to the presynaptic cell, demonstrating a strictly presynaptic requirement for Fusl function.

Role of protein phosphatase 2A in regulating the visual signaling in Drosophila.

Drosophila visual signaling, a G-protein-coupled phospholipase Cbeta (PLCbeta)-mediated mechanism, is regulated by eye-protein kinase C (PKC) that promotes light adaptation and fast deactivation, most likely via phosphorylation of inactivation no afterpotential D (INAD) and TRP (transient receptor potential). To reveal the critical phosphatases that dephosphorylate INAD, we used several biochemical analyses and identified protein phosphatase 2A (PP2A) as a candidate. Importantly, the catalytic subunit of PP2A, microtubule star (MTS), is copurified with INAD, and an elevated phosphorylation of INAD by eye-PKC was observed in three mts heterozygotes.

Effects of a mutation in the Drosophila porin gene encoding mitochondrial voltage-dependent anion channel protein on phototransduction.

Mitochondrial porins, also know as VDACs (voltage-dependent anion channels), play an important role in regulating energy metabolism, apoptosis, and the transport of metabolites across the mitochondrial outer membrane. So far three distinct isoforms of VDAC (VDAC1-3) have been reported in vertebrates, but their functions remain unknown. The annotation database of the Drosophila melanogaster genome sequence has identified four genes (porin, CG17137, CG17139, and CG17140) encoding different isoforms of VDACs.

Complete RNAi rescue of neuronal degeneration in a constitutively active Drosophila TRP channel mutant.

RNA interference has been widely used to reduce the quantity of the proteins encoded by the targeted genes. A constitutively active, dominant allele of trp, TrpP365, causes massive degeneration of photoreceptors through a persistent and excessive Ca2+ influx. Here we show that a substantial reduction of the TRP channel protein by RNAi in TrpP365 heterozygotes completely rescues the neuronal degeneration and significantly improves the light-elicited responses of the eye.

Specific molecular alterations in the norpA-encoded phospholipase C of Drosophila and their effects on electrophysiological responses in vivo.

A large number of mutants in the norpA gene, which encodes the phospholipase C (PLC) involved in Drosophila phototransduction, is available for the investigation of the effects of specific amino acid substitutions in PLC on biochemical and electrophysiological properties of these mutants. Of the 47 norpA mutants screened for PLC protein content, all but one (H43) displayed drastically decreased amounts of the protein suggesting that almost any mutational alteration has a deleterious effect on the integrity of the protein. Three new amino acids were identified in the catalytic domains X and Y that are important for PLC catalytic activity and the generation of photoreceptor responses (ERG).

Genetic approaches to visual transduction in Drosophila melanogaster.

Because almost everything we know about Drosophila phototransduction has come from studies based on genetic approaches, this review begins with a discussion of genetic approaches. We then present a brief overview of Drosophila phototransduction (section on Drosophila phototransduction: an overview) followed by a more detailed treatment of individual components of the transduction machinery (section on Components of the phototransduction machinery). Discussion of transduction mechanisms is presented under three headings: Mechanism(s) of channel excitation, Organization of the transduction proteins, and Regulatory mechanisms in phototransduction.

Altered drug resistance and recovery from paralysis in Drosophila melanogaster with a deficient histamine-gated chloride channel.

The recent identification and characterization of two genes, encoding histamine-gated chloride channel subunits from Drosophila melanogaster, has confirmed that histamine is a major neurotransmitter in the fruitfly. One of the cloned genes, hclA (synonyms: HisCl-alpha1; HisCl2), corresponds to ort (ora transientless), mutationsin which affect synaptic transmission in the Drosophila visual system. We identified a mutational change (a null mutation) in the genomic and RNA copies of hclA derived from mutants carrying the ort(1) allele.

Photoreceptor degeneration and Ca2+ influx through light-activated channels of Drosophila.

We discuss in this chapter the role of Ca2+ homeostasis in maintaining the structural integrity of photoreceptor cells in Drosophila. Both insufficient and excessive amounts of Ca2+ in photoreceptor cells appear to lead to cell degeneration. Because one of the two classes of light-sensitive channels in Drosophila photoreceptors is highly Ca2+-permeable, how well this class of channels functions can profoundly affect Ca2+ homeostasis.

The target of Drosophila photoreceptor synaptic transmission is a histamine-gated chloride channel encoded by ort (hclA).

By screening Drosophila mutants that are potentially defective in synaptic transmission between photoreceptors and their target laminar neurons, L1/L2, (lack of electroretinogram on/off transients), we identified ort as a candidate gene encoding a histamine receptor subunit on L1/L2. We provide evidence that the ort gene corresponds to CG7411 (referred to as hclA), identified in the Drosophila genome data base, by P-element-mediated germ line rescue of the ort phenotype using cloned hclA cDNA and by showing that several ort mutants exhibit alterations in hclA regulatory or coding sequences and/or allele-dependent reductions in hclA transcript levels. Other workers have shown that hclA, when expressed in Xenopus oocytes, forms histamine-sensitive chloride channels.

Single amino acid change in the fifth transmembrane segment of the TRP Ca2+ channel causes massive degeneration of photoreceptors.

The trp gene encodes subunits of a highly Ca(2+)-permeable class of light-activated channels of Drosophila photoreceptors. The recently characterized mutation in this gene, Trp(P365), is semidominant and causes massive degeneration of photoreceptors by making the TRP channel constitutively active. We show that a single amino acid change, Phe-550 to Ile, near the beginning of the fifth transmembrane domain of TRP channel subunits is necessary to induce, and sufficient to closely mimic, the original mutant phenotypes of Trp(P365).