Academic journal article Genetics

Overexpression Screen in Drosophila Identifies Neuronal Roles of GSK-3[beta]/shaggy as a Regulator of AP-1-Dependent Developmental Plasticity

Academic journal article Genetics

Overexpression Screen in Drosophila Identifies Neuronal Roles of GSK-3[beta]/shaggy as a Regulator of AP-1-Dependent Developmental Plasticity

Article excerpt

ABSTRACT

AP-1, an immediate-early transcription factor comprising heterodimers of the Fos and Jun proteins, has been shown in several animal models, including Drosophila, to control neuronal development and plasticity. In spite of this important role, very little is known about additional proteins that regulate, cooperate with, or are downstream targets of AP-1 in neurons. Here, we outline results from an overexpression/misexpression screen in Drosophila to identify potential regulators of AP-1 function at third instar larval neuromuscular junction (NMJ) synapses. First, we utilize >4000 enhancer and promoter (EP) and EPgy2 lines to screen a large subset of Drosophila genes for their ability to modify an AP-1-dependent eye-growth phenotype. Of 303 initially identified genes, we use a set of selection criteria to arrive at 25 prioritized genes from the resulting collection of putative interactors. Of these, perturbations in 13 genes result in synaptic phenotypes. Finally, we show that one candidate, the GSK-3β-kinase homolog, shaggy, negatively influences AP-1-dependent synaptic growth, by modulating the Jun-N-terminal kinase pathway, and also regulates presynaptic neurotransmitter release at the larval neuromuscular junction. Other candidates identified in this screen provide a useful starting point to investigate genes that interact with AP-1 in vivo to regulate neuronal development and plasticity.

THE ability of an animal to adjust to its environment relies significantly on the capacity of neuronal circuits to change in response to stimuli. This intrinsic plasticity of neurons results in persistent modifications that are orchestrated by the synthesis of new proteins, either through translation of preexisting mRNA or activation of nuclear transcription (Nestler et al. 2001; West et al. 2001; Lonze and Ginty 2002; Klann et al. 2004). Several transcription factors have thus been implicated in the regulation of neuronal plasticity both during development and during acute changes such as those required for memory formation. Of these, a fundamentally important candidate is the immediateearly transcription factor AP-1 comprising heterodimers of Fos and Jun (Kaminska et al. 1994; Hiroi et al. 1998; Morris et al. 2000; Kaczmarek et al. 2002; Sanyal et al. 2002; Etter et al. 2005). Experimental demonstrations in diverse systems have verified the central role of AP-1 in controlling the development, growth, survival, and plasticity of neurons. These include learning behavior in rodents, developmental plasticity in insect models, and cocaine addiction in mammals, to name a few (Sanyal et al. 2002; McClung and Nestler 2003; Yasoshima et al. 2006). Despite this obvious centrality of AP-1 function in neurons, additional proteins that interact with and modulate AP-1 function in neurons remain largely unknown.

Neural roles for Drosophila Fos and Jun, encoded by the genes kayak and jra, respectively, have been identi- fied on the basis of experiments that explore AP-1 function in the developing compound eye and at the third instar larval neuromuscular junction (NMJ). For instance, Fos regulates G2/M transition in dividing ommatidia and also controls cell survival and growth (Ciapponi and Bohmann 2002; Hyun et al. 2006). More pertinently, studies using the motor synapse demonstrate that AP-1 positively regulates growth and presynaptic neurotransmitter release (Sanyal et al. 2002; Collins et al. 2006). Inhibiting AP-1 activity in differentiated postmitotic motor neurons results in smaller synapses with reduced evoked release of neurotransmitter. Conversely, coexpression of wild-type Fos and Jun proteins, but not either protein alone, leads to expanded synapses and enhanced transmitter release. These phenotypes suggest significant roles for the AP- 1 protein in mediating changes in the structure and function of neuronal synapses that are likely to be relevant in the context of neuronal plasticity. Recent experiments also indicate a requirement for AP-1 in long-term olfactory conditioning in flies (A. …

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