RESUMO
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. At the neuromuscular junction (NMJ) synapse, an extended allelic series of Smg1 mutants show impaired structural architecture, with decreased terminal arbor size, branching and synaptic bouton number. Functionally, loss of Smg1 results in a ~50% reduction in basal neurotransmission strength, as well as progressive transmission fatigue and greatly impaired synaptic vesicle recycling during high-frequency stimulation. Mutation of other NMD pathways genes (Upf2 and Smg6) similarly impairs neurotransmission and synaptic vesicle cycling. These findings suggest that the NMD pathway acts to regulate proper mRNA translation to safeguard synapse morphology and maintain the efficacy of synaptic function.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/fisiologia , Células Fotorreceptoras de Invertebrados/metabolismo , Terminações Pré-Sinápticas/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Vesículas Sinápticas/metabolismo , Animais , Proteínas de Drosophila/genética , Teste de Complementação Genética , Testes Genéticos , Transdução de Sinal Luminoso/genética , Morfogênese/genética , Junção Neuromuscular/fisiologia , Células Fotorreceptoras de Invertebrados/patologia , Terminações Pré-Sinápticas/metabolismo , Proteínas Serina-Treonina Quinases/genética , Retina/crescimento & desenvolvimento , Retina/patologia , Deleção de Sequência/genética , Transmissão Sináptica/genética , Vesículas Sinápticas/genética , Vesículas Sinápticas/patologiaRESUMO
This is the first of two reviews that include some of the studies that we, members of the Pak lab and collaborators, carried out from 1998 to 2010 on the functional and physical interactions among several Drosophila phototransduction components. The report includes our studies on the regulations and/or the functions of arrestin II (Arr2), norpA (PLC), inactivation no afterpotential D (INAD), transient receptor potential (TRP), TRP-like (TRPL), inactivation no afterpotential E (INAE), and Porin.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/fisiologia , Proteínas do Olho/metabolismo , Transdução de Sinal Luminoso/fisiologia , Animais , Arrestinas/genética , Arrestinas/metabolismo , Proteínas de Drosophila/genética , Proteínas do Olho/genética , Regulação da Expressão Gênica , Canais Iônicos , Canal de Cátion TRPA1 , Canais de Cátion TRPC/genética , Canais de Cátion TRPC/metabolismo , Canais de Ânion Dependentes de Voltagem/genética , Canais de Ânion Dependentes de Voltagem/metabolismoRESUMO
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. Using this strategy, two classes of PDA-defective mutants were identified and isolated, nina and ina, each comprising multiple complementation groups. The nina mutants are characterized by allele-dependent reduction in the major rhodopsin, Rh1, whereas the ina mutants display defects in some aspects of functions related to the transduction channel, TRP (transient receptor potential). The signaling proteins that have been identified and elucidated through the studies of nina mutants include the Drosophila opsin protein (NINAE), the chaperone protein for nascent opsin (NINAA), and the multifunctional protein, NINAC, required in multiple steps of the Drosophila phototransduction cascade. Also identified by the nina mutants are some of the key enzymes involved in the biogenesis of the rhodopsin chromophore. As for the ina mutants, they led to the discovery of the scaffold protein, INAD, responsible for the nucleation of the supramolecular signaling complex. Also identified by the ina mutants is one of the key members of the signaling complex, INAC (ePKC), and two other proteins that are likely to be important, though their roles in the signaling cascade have not yet been fully elucidated. In most of these cases, the protein identified is the first member of its class to be so recognized.
Assuntos
Proteínas de Drosophila/deficiência , Proteínas do Olho/metabolismo , Mutação/genética , Células Fotorreceptoras de Invertebrados/fisiologia , Proteínas de Ligação ao Retinol/metabolismo , Animais , Animais Geneticamente Modificados , Drosophila/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Eletrorretinografia , Proteínas do Olho/genética , Testes Genéticos , Proteínas de Ligação ao Retinol/deficiência , Transdução de Sinais/genéticaRESUMO
This is the second of two reviews that include some of the studies we, members of the Pak laboratory and collaborators, did from 2000 to 2010 on the mutants that affect synaptic transmission in the Drosophila visual system. Of the five mutants we discuss, two turned out to also play roles in the larval neuromuscular junction. This review complements the one on phototransduction to give a fairly complete account of what we focused on during the 10-year period, although we also did some studies on photoreceptor degeneration in the early part of the decade. Besides showing the power of using a genetic approach to the study of synaptic transmission, the review contains some unexpected results that illustrate the serendipitous nature of research.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/fisiologia , Proteínas do Olho/metabolismo , Transdução de Sinal Luminoso/fisiologia , Sinapses/fisiologia , Vias Visuais/fisiologia , Animais , Canais de Cloreto , Proteínas de Drosophila/genética , Proteínas de Drosophila/história , Proteínas do Olho/genética , Proteínas do Olho/história , Ribonucleoproteínas Nucleares Heterogêneas Grupo F-H , História do Século XX , História do Século XXI , Canais Iônicos , Transdução de Sinal Luminoso/genética , Mutação , Fosfatidilinositol 3-Quinases , Sinapses/genética , Transmissão Sináptica/genética , Transmissão Sináptica/fisiologia , Canal de Cátion TRPA1 , Canais de Cátion TRPCRESUMO
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). However, DAGLs that might fulfill this role have not been previously identified in any organism. In this work, the Drosophila DAGL gene, inaE, has been identified from mutants that are defective in photoreceptor responses to light. The inaE-encoded protein isoforms show high sequence similarity to known mammalian DAG lipases, exhibit DAG lipase activity in vitro, and are highly expressed in photoreceptors. Analyses of norpA inaE double mutants and severe inaE mutants show that normal DAGL activity is required for the generation of physiologically meaningful photoreceptor responses.