Subcellular targeting of DISC1 is dependent on a domain independent from the Nudel binding site.

Disrupted in schizophrenia 1 (DISC1) has been identified as a putative risk factor for schizophrenia and affective disorders through study of a Scottish family with a balanced (1;11) (q42.1;q14.3) translocation, which results in the disruption of the DISC1 locus and cosegregates with major psychiatric disease. Several other reports of genetic linkage and association between DISC1 and schizophrenia in a range of patient populations have added credibility to the DISC1-schizophrenia theory, but the function of the DISC1 protein is still poorly understood. Recent studies have suggested that DISC1 plays a role in neuronal outgrowth, possibly through reported interactions with the molecules Nudel and FEZ1. Here we have analyzed the DISC1 protein sequence to identify previously unknown regions that are important for the correct targeting of the protein and conducted imaging studies to identify DISC1 subcellular location. We have identified a central coiled-coil region and show it is critical for the subcellular targeting of DISC1. This domain is independent from the C-terminal Nudel binding domain highlighting the multidomain nature/functionality of the DISC1 protein. Furthermore, we have been able to provide the first direct evidence that DISC1 is localized to mitochondria in cultured cortical neurons that are dependent on an intact cytoskeleton. Surprisingly, Nudel is seen to differentially associate with mitochondrial markers in comparison to DISC1. Disruption of the cytoskeleton results in colocalization of Nudel and mitochondrial markers-the first observation of such a direct relationship. Mitochondrial dysfunction has been implicated to play a role in schizophrenia so we speculate that mutations in DISC1 or Nudel may impair mitochondrial transport or function, initiating a cascade of events culminating in psychiatric illness.

Expression of disrupted in schizophrenia 1 (DISC1) protein in the adult and developing mouse brain indicates its role in neurodevelopment.

Disrupted in Schizophrenia 1 (DISC1) was identified as a potential susceptibility gene for schizophrenia due to its disruption by a balanced t(1;11) (q42;q14) translocation, which has been shown to cosegregate with major psychiatric disease in a large Scottish family. We have recently presented evidence that DISC1 exists in a neurodevelopmentally regulated protein complex with Nudel. In this study, we report the protein expression profile of DISC1 in the adult and developing mouse brain utilizing immunohistochemistry and quantitative Western blot. In the adult mouse brain, DISC1 is expressed in neurons within various brain areas including the olfactory bulb, cortex, hippocampus, hypothalamus, cerebellum and brain stem. During development, DISC1 protein is detected at all stages, from E10 to 6 months old, with two significant peaks of protein expression of a DISC1 isoform at E13.5 and P35. Interestingly, these time points correspond to critical stages during mouse development, the active neurogenesis period in the developing brain and the period of puberty. Together, these results suggest that DISC1 may play a critical role in brain development, consistent with the neurodevelopmental hypothesis of the etiology of schizophrenia.

Disrupted in Schizophrenia 1 and Nudel form a neurodevelopmentally regulated protein complex: implications for schizophrenia and other major neurological disorders.

Disrupted In Schizophrenia 1 (DISC1) was identified as a potential susceptibility gene for schizophrenia due to its disruption by a balanced t(1;11) (q42;q14) translocation, which has been shown to cosegregate with major psychiatric disease in a large Scottish family. We have demonstrated that DISC1 exists in a neurodevelopmentally regulated protein complex with Nudel. The complex is abundant at E17 and in early postnatal life but is greatly reduced in the adult. Nudel has previously been shown to bind Lis1, a gene underlying lissencephaly in humans. Critically, we show that the predicted peptide product resulting from the Scottish translocation removes the interaction domain for Nudel. DISC1 interacts with Nudel through a leucine zipper domain and binds to a novel DISC1-interaction domain on Nudel, which is independent from the Lis1 binding site. We show that Nudel is able to act as a bridge between DISC1 and Lis1 to allow formation of a trimolecular complex. Nudel has been implicated to play a role in neuronal migration, together with the developmental variation in the abundance of the DISC1-Nudel complex, may implicate a defective DISC1-Nudel complex as a neurodevelopmental cause of schizophrenia.

Edg2 receptor distribution in adult rat brain.

The Edg (endothelial differentiation gene) receptors are recently discovered G-protein coupled receptors which are activated by endogenous lysophospholipids. The cellular activities mediated by Edg receptors are reminiscent of those normally associated with Trk receptor activation and include modulation of cell growth, differentiation, proliferation and migration as well as apoptotic and cytoskeletal effects. In this study we have investigated immunohistochemically the distribution of one family member, the Edg2 receptor, within the adult rat brain and shown the protein expression to be most prominent in white matter tract regions. This suggests a possible role for the Edg2 receptor in nerve cell myelination.

Edg2 receptor functionality: gialpha1 coexpression and fusion protein studies.

Recombinant receptor cell lines are widely used in G-protein-coupled receptor selectivity studies. To unequivocally interpret the results of such studies, it is essential that the host cell line does not endogenously express the receptor of interest and in addition is unresponsive to the receptor's natural ligand. Here we describe an approach to overcome such difficulties associated with orphan receptors or, as in the present case, receptors whose endogenous ligand ubiquitously affects mammalian cells. The functional heterologous assay system described is for the hEdg2 receptor, which uses lysophosphatidic acid as its endogenous ligand. Once activated, this receptor mediates its effects via multiple secondary messenger pathways, including a Gi-coupled pathway. We have transiently expressed a pertussis toxin-insensitive hEdg2 receptor-ratGialpha1 fusion protein into human embryonic kidney cells and have monitored the ability of compounds to stimulate [(35)S]GTPgammaS binding in membranes prepared from these cells after pretreatment with toxin. Because the assay conditions used favor Gi-mediated responses and because endogenous Gialpha subunits are rendered inactive, the response measured is, by definition, fusion protein-mediated. Consequently, we have developed an assay that monitors definitively Edg2 receptor-mediated responses in a mammalian cell line. A limited structure activity relationship study suggests that the lysophospholipid carbon chain has a role in receptor activation and in addition indicates that certain modifications to the phosphate group are tolerated.