InteractiveFly: GeneBrief

dysbindin: Biological Overview | References

The molecular mechanisms that achieve homeostatic stabilization of neural function remain largely unknown. To better understand how neural function is stabilized during development and throughout life, an electrophysiology-based forward genetic screen was used, and the function of more than 250 neuronally expressed genes was assessed for a role in the homeostatic modulation of synaptic transmission in Drosophila. This screen ruled out the involvement of numerous synaptic proteins and identified a critical function for dysbindin, a gene linked to schizophrenia in humans. dysbindin was found to be required presynaptically for the retrograde, homeostatic modulation of neurotransmission, and functions in a dose-dependent manner downstream or independently of calcium influx. Thus, dysbindin is essential for adaptive neural plasticity and may link altered homeostatic signaling with a complex neurological disease (Dickman, 2009).

At glutamatergic synapses of species ranging from Drosophila to human, disruption of postsynaptic neurotransmitter receptor function can be precisely offset by an increase in presynaptic neurotransmitter release to homeostatically maintain normal postsynaptic excitation. The Drosophila neuromuscular junction (NMJ) is a glutamatergic synapse that is used as a model for this form of homeostatic signaling in the nervous system. Efficient homeostatic modulation of presynaptic release at the Drosophila NMJ can occur in ten min following bath application of philanthotoxin-433 (PhTx; a polyamine toxin present in the venom sac of the solitary digger wasp Philanthus triangulum), which persistently and specifically inhibits postsynaptic glutamate receptors (Dickman, 2009).

This study has systematically screened for mutations that block the rapid, PhTx-dependent induction of synaptic homeostasis. Mutations in 276 genes were screened electrophysiologically. For each mutant, an average value was calculated for the amplitude of both the spontaneous miniature excitatory junctional potential (mEJP) and evoked excitatory junctional potential (EJP) following treatment of the dissected neuromuscular preparation with PhTx for 10 min. 14 mutants were isolated with average EJP amplitudes more than two standard deviations smaller than the distribution mean. From these candidates, 7 mutants were identified that block synaptic homeostasis without an obvious effect on NMJ morphology or baseline synaptic transmission. It is concluded that the molecular mechanisms of synaptic homeostasis can be genetically separated from the mechanisms responsible for normal neuromuscular development and baseline synaptic transmission (Dickman, 2009).

A fraction of the mutants assayed (19.5%) are previously published genetic lesions. This allows ruling out of the involvement of numerous genes and associated biochemical processes. Mutations that disrupt RNA-interference/micro-RNA processing, retrograde trans-synaptic signaling, synaptic transmission, active zone assembly, synaptic vesicle endocytosis and mitochondria all showed reliable homeostatic compensation. Therefore, synaptic homeostasis is a robust phenomenon, unperturbed by a broad spectrum of synaptic mutations. In addition, significant homeostatic compensation in synaptojanin and endophilin mutants argues against the involvement of synaptic vesicle endocytosis and indicates that the size of the recycling synaptic vesicle pool is not a limiting factor for synaptic homeostasis. These data also emphasize the importance and specificity of those identified mutations that do block synaptic homeostasis. These include four ion channels, two of which are of unknown function, and two calcium-binding proteins of unknown function. Thus, homeostatic signaling at the NMJ may include previously unexplored mechanisms of synaptic modulation (Dickman, 2009).

One mutation that was identified with a specific defect in homeostatic compensation is a transposon insertion that resides in the Drosophila homologue of dysbindin (CG6856). The DTNBP1 (dysbindin) locus is linked with schizophrenia in humans (Ross, 2006). A transposon insertion was identified within the dysbindin locus (pBac^e01028, referred to as dysb¹, that showed a complete absence of homeostatic compensation following application of PhTx. A similar effect was observed when dysb¹ was placed in trans to a deficiency that uncovers the dysb locus, indicating that the dysb¹ mutant was a strong loss of function or null mutation. No significant change in baseline synaptic transmission was observed in dysb¹ mutant animals (0.5 mM extracellular calcium). Thus, under these recording conditions, this mutation disrupted synaptic homeostasis without altering baseline neurotransmission. As a control, synaptic homeostasis was normal in animals in which the pBac^e01028 transposon was precisely excised (Dickman, 2009).

The dysb gene is ubiquitously expressed in Drosophila embryos. Therefore, a dysbindin transgene was generated and expressed in the dysb¹ mutant. Presynaptic expression of dysb fully restored homeostatic compensation in the dysb¹ mutant background, whereas muscle-specific expression of dysb did not. Thus, Dysbindin is necessary presynaptically for the rapid induction of synaptic homeostasis (Dickman, 2009).

It was next asked whether Dysbindin is also required for the sustained expression of synaptic homeostasis. Double mutant animals were generated harboring both the dysb¹ mutation and a mutation in a gene encoding a postsynaptic glutamate receptor (GluRIIA). GluRIIA mutant animals normally show robust homeostatic compensation. However, homeostatic compensation was blocked in GluRIIA; dysb¹ double mutant animals. Thus, dysbindin was also necessary for the sustained expression of synaptic homeostasis over several days of larval development (Dickman, 2009).

Synapse morphology was qualitatively normal in dysb mutants including both the shape of the presynaptic nerve terminal and the levels, localization and organization of synaptic markers including futsch-positive microtubules, synapsin and synaptotagmin. Bouton number and active zone density are also normal in dysb mutants. Thus, the disruption of synaptic homeostasis in dysb¹ mutants is not a secondary consequence of altered or impaired NMJ development (Dickman, 2009).

In the vertebrate nervous system, Dysbindin is associated with synaptic vesicles (Talbot, 2006). The localization was examined of a Venus-tagged dysb transgene (ven-dysb) that rescues the dysb¹ mutant. Ven-Dysb showed extensive overlap with synaptic vesicle associated proteins when expressed in neurons. Thus, Dysbindin functions presynaptically, potentially at or near the synaptic vesicle pool (Dickman, 2009).

To further define the function of Dysbindin, baseline synaptic transmission in the dysb mutant was investigated in greater detail. At 0.5 mM extracellular calcium, synaptic transmission in dysb¹ mutant animals was indistinguishable from wild type. However, when extracellular calcium was reduced, baseline synaptic transmission was significantly impaired in dysb compared to wild type and this defect was rescued by presynaptic expression of dysb. Thus, there is an alteration of the calcium dependence of synaptic transmission in the dysb mutant. Indeed, at reduced extracellular calcium, both paired-pulse facilitation and facilitation that occurs during a prolonged stimulus train were increased in dysb mutants (Dickman, 2009).

In vertebrates, the levels of dysb expression correlate with parallel changes in extracellular glutamate concentration (Numakawa, 2004). Therefore, whether dysb overexpression might increase presynaptic release was tested. In wild-type animals overexpressing dysb in neurons, synaptic transmission is normal at low extracellular calcium (0.2 and 0.3 mM Ca²⁺) but was enhanced at relatively higher extracellular calcium (0.5 mM Ca²⁺). The complementary effects of dysb loss-of-function and overexpression confirm that Dysbindin has an important influence on calcium-dependent vesicle release (Dickman, 2009).

The presynaptic Ca_V2.1 calcium channel, encoded by cacophony (cac), is required for synaptic vesicle release at the Drosophila NMJ. cac mutations decrease presynaptic calcium influx and also block synaptic homeostasis. Genetic interaction between dysb and cac was tested during synaptic homeostasis. Because homozygous cac and dysb mutations individually block synaptic homeostasis, analysis of double mutant combinations would not be informative. An analysis of heterozygous mutant combinations and gene overexpression were examined. Synaptic homeostasis was suppressed by a heterozygous mutation in cac. However, this suppression was not enhanced by the presence of a heterozygous mutation in dysb. In addition, neuronal overexpression of cac did not restore homeostatic compensation in dysb mutant animals and the enhancement of presynaptic release caused by neuronal dysb overexpression still occurs in a heterozygous cac mutant background. Thus, Dysbindin may function downstream or independently of Cac during synaptic homeostasis (Dickman, 2009).

To further explore the relationship between Dysbindin and Cac, it was asked whether dysb mutations might directly influence presynaptic calcium influx. The spatially averaged calcium signal in dysb¹ was indistinguishable from wild type, indicating no difference in presynaptic calcium influx. Thus, Dysbindin appears to function downstream or independently of calcium influx to control synaptic homeostasis (Dickman, 2009).

Through a systematic electrophysiological analysis of more than 250 mutants this study could rule out the involvement of numerous synaptic proteins and biochemical processes in the mechanisms of synaptic homeostasis and demonstrate that this phenomenon is separable from the molecular mechanisms that specify structural and functional synapse development. Dysbindin is therefore identified as an essential presynaptic component within a homeostatic signaling system that regulates and stabilizes synaptic efficacy. Dysbindin functions downstream or independently of the presynaptic Ca_V2.1 calcium channel in the mechanisms of synaptic homeostasis (Dickman, 2009).

Emerging lines of evidence suggest that glutamate hypofunction could be related to the etiology of schizophrenia. Likewise, reduced levels of dysbindin expression were associated with schizophrenia (Weickert, 3008; Talbot, 2004). The sandy mouse, which lacks Dysbindin, has a decreased rate of vesicle release (~30% decrease), a correlated decrease in vesicle pool size and an increased thickness of the postsynaptic density (Chen, 2008). This study confirms a modest, facilitatory function for Dysbindin during baseline transmission. However, numerous mutations with similar or more severe defects in baseline transmission show normal synaptic homeostasis. By contrast, loss of Dysbindin completely blocks the adaptive, homeostatic modulation of vesicle release, suggesting that the potential contribution of dysbindin mutations to schizophrenia may be derived from altered homeostatic plasticity as opposed to decreased baseline glutamatergic transmission (Dickman, 2009).

The schizophrenia susceptibility factor Dysbindin and its associated complex sort cargoes from cell bodies to the synapse

Dysbindin assembles into the biogenesis of lysosome related organelles complex 1 (BLOC-1), which interacts with the adaptor protein complex 3 (AP-3) mediating a common endosome trafficking route. Deficiencies in AP-3 and BLOC-1 affect synaptic vesicle composition. However, whether AP-3-BLOC-1-dependent sorting events that control synapse membrane protein content take place in cell bodies, upstream nerve terminals, remains unknown. This study tested this hypothesis analyzing the targeting of phosphatidylinositol-4-kinase type II α (PI4KIIα), a membrane protein present in pre and postsynaptic compartments. PI4KIIα co-purified with BLOC-1 and AP-3 in neuronal cells. These interactions translated into a decreased PI4KIIα content in the dentate gyrus of dysbindin-null BLOC-1 deficiency, and AP-3-null mice. Reduction of PI4KIIα in the dentate reflects a failure to traffic from the cell body. PI4KIIα was targeted to processes in wild type primary cultured cortical neurons and PC12 cells, but failed to reach neurites in cells lacking either AP-3 or BLOC-1. Similarly, disruption of an AP-3 sorting motif in PI4KIIα impaired its sorting into processes of PC12 and primary cultured cortical neuronal cells. These findings indicate a novel vesicle transport mechanism requiring BLOC-1 and AP-3 complexes for cargo sorting from neuronal cell bodies to neurites and nerve terminals (Larimore, 2011).

Nucleocytoplasmic shuttling of dysbindin-1, a schizophrenia-related protein, regulates synapsin I expression

Dysbindin-1 is a 50-kDa coiled-coil-containing protein encoded by the gene DTNBP1 (dystrobrevin-binding protein 1), a candidate genetic factor for schizophrenia. Genetic variations in this gene confer a susceptibility to schizophrenia through a decreased expression of dysbindin-1. It was reported that dysbindin-1 regulates the expression of presynaptic proteins and the release of neurotransmitters. However, the precise functions of dysbindin-1 are largely unknown. This study shows that dysbindin-1 is a novel nucleocytoplasmic shuttling protein and translocated to the nucleus upon treatment with leptomycin B, an inhibitor of exportin-1/CRM1-mediated nuclear export. Dysbindin-1 harbors a functional nuclear export signal necessary for its nuclear export, and the nucleocytoplasmic shuttling of dysbindin-1 affects its regulation of synapsin I expression. In brains of sandy mice, a dysbindin-1-null strain that displays abnormal behaviors related to schizophrenia, the protein and mRNA levels of synapsin I are decreased. These findings demonstrate that the nucleocytoplasmic shuttling of dysbindin-1 regulates synapsin I expression and thus may be involved in the pathogenesis of schizophrenia (Fei, 2010).

Search PubMed for articles about Drosophila Dysbindin

Chen, X. W., et al. (2008). DTNBP1, a schizophrenia susceptibility gene, affects kinetics of transmitter release. J. Cell Biol. 181(5): 791-801. PubMed Citation: 18504299

Dickman, D. K. and Davis, G. W. (2009). The schizophrenia susceptibility gene dysbindin controls synaptic homeostasis. Science 326(5956): 1127-30. PubMed Citation: 19965435

Fei, E., et al. (2010). Nucleocytoplasmic shuttling of dysbindin-1, a schizophrenia-related protein, regulates synapsin I expression. J. Biol. Chem. 285(49): 38630-40. PubMed Citation: 20921223

Larimore, J., et al. (2011). The schizophrenia susceptibility factor Dysbindin and its associated complex sort cargoes from cell bodies to the Synapse. Mol. Biol. Cell [Epub ahead of print]. PubMed Citation: 21998198

Numakawa, T., et al. (2004). Evidence of novel neuronal functions of dysbindin, a susceptibility gene for schizophrenia. Hum. Mol. Genet. 13(21): 2699-708. PubMed Citation: 15345706

Ross, C. A., et al. (2006). Neurobiology of schizophrenia. Neuron 52(1): 139-53. PubMed Citation: 17015232

Talbot, K., et al. (2004). Dysbindin-1 is reduced in intrinsic, glutamatergic terminals of the hippocampal formation in schizophrenia. J. Clin. Invest. 113(9): 1353-63. PubMed Citation: 15124027

Talbot, K., et al. (2006). Dysbindin-1 is a synaptic and microtubular protein that binds brain snapin. Hum. Mol. Genet. 15(20): 3041-54. PubMed Citation: 16980328

Weickert, C. S., et al. (2008). Reduced DTNBP1 (dysbindin-1) mRNA in the hippocampal formation of schizophrenia patients. Schizophr. Res. 98(1-3): 105-10. PubMed Citation: 17961984

date revised: 30 October 2011

Home page: The Interactive Fly © 2011 Thomas Brody, Ph.D.