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Neuroscience 2009: DISC1 Dominates the Schizophrenia Presence in Chicago

3 November 2009. There was no shortage of DISC1 chatter at the 40th annual meeting of the Society for Neuroscience, held 17-21 October 2009 in Chicago. The highlight was a mini-symposium on Monday, convened by Nicholas Brandon of Pfizer (formerly Wyeth), which covered topics ranging from the role of DISC1 in developing and mature neurons, to the question of whether DISC1 aggregation might contribute to sporadic disease. Co-organizer Akira Sawa of Johns Hopkins, Baltimore, Maryland, unfortunately had to miss the session, but a preview of the mini-symposium was published on October 14 in the Journal of Neuroscience (Brandon et al., 2009). This story covers highlights of the talks as well as posters on DISC1 presented at the meeting.

DISC1 in development
It has been nearly a decade since DISC1 was identified at the breakpoint of a 1:11 chromosomal translocation cosegregating with major mental illness in a large Scottish family (Millar et al., 2000). Since then, a picture has emerged of DISC1 as a multitasking scaffold that associates with a variety of partners to regulate neurogenesis, neuronal migration, and integration during fetal development and in adulthood. Ju Young Kim talked about her recent work from the Ming lab at Johns Hopkins University, Baltimore, Maryland, showing that DISC1 regulates the morphology, migration, and integration of new neurons in the adult hippocampus by negatively regulating the activity of the Akt/mTOR pathway via the actin-binding protein girdin, also known as KIAA1212 (see SRF related news story). This pathway is distinct from another DISC1-regulated cascade that Li-Huei Tsai and colleagues at MIT, Cambridge, Massachusetts, identified recently (see SRF related news story). That work showed that DISC1 regulates GSK3/Wnt/β-catenin signaling, which is critical for the proliferation of neuronal progenitors during development and in adult mice.

Does DISC1 tie into different signaling pathways at different stages of a cell’s life? Kim presented two pieces of data that support this idea. In Tsai’s published experiments, inhibitors of GSK3β reversed the effects of DISC1 loss on neurogenesis. However, Kim reported that GSK3β inhibitors had no effect on DISC1-related changes in morphogenesis and migration in the postnatal hippocampus. Also, expression profiling revealed little girdin in adult neuronal progenitor cells, but Kim noted that expression increased with differentiation.

Karun Singh from the Tsai lab showed new data on another DISC1 interacting protein, suggesting that it might comprise a redundant pathway for β-catenin activation in neuronal progenitor cells. The protein, DIXDC1, is a positive regulator of Wnt signaling that has mostly been studied in cancer (Wang et al., 2009), but is highly expressed in neuronal progenitors and neurons. Singh reported that DIXDC1 interacts with DISC1 in E15 mouse brain. In cell-based assays, either DIXDC1 or DISC1 activated a β-catenin-regulated reporter gene. The two appeared to compensate for each other, as overexpression of one could rescue the activity of the other on gene expression in a knockdown model.

Singh speculated that these parallel pathways could provide a biological explanation for the incomplete penetrance of DISC1 deletions, where overexpression of DIXDC1 might compensate for loss or lower expression of DISC1. In support of that idea, he found that DIXDC1 expression rescued the proliferation defect caused by DISC1 RNAi knockdown in mouse hippocampus. But the two genes were not exactly equivalent. Where DIXDC1 RNAi caused a decrease in progenitor proliferation and a neural migration defect, DISC1 could reverse the progenitor cell defect, but not the migration problem. Singh presented data that DIXDC1 affected cell migration by binding to the DISC1 partner nuclear distribution element 1 (Ndel1) protein.

More evidence for the importance of the GSK3/β-catenin pathway in DISC1 actions comes from the work of Hazel Sive of MIT, who described studies on the zebrafish homolog of DISC1. Knockdown of that gene is embryonic lethal and results in a truncated brain structure with abnormal neuronal patterning. The phenotype, which Sive showed arises from unregulated GSK/β-catenin signaling, can be substantially rescued by expression of human DISC1.

Using the zebrafish, Sive tried to introduce a C-terminal truncation analogous to the human chromosome 1:11 translocation using an antisense oligonucleotide to target the exon8/intron boundary. The treatment resulted in a 50/50 mix of wild-type and truncated mRNAs, but protein levels were not measured. The manipulation caused a severe brain defect, but surprisingly it occurred without activation of GSK3 or loss of β-catenin-driven gene expression.

This brought up the question of what is happening in people with the DISC1 translocation—are GSK3β and β-catenin involved or not? Brandon pointed out that no one has detected truncated protein in blood cells from people with the translocation, and the model of DISC1 pathology invokes haploinsufficiency. However, Brandon said, “We won’t know for sure until we have brain tissue from that family.”

Standing in for the absent Sawa, Atsushi Kamiya, also of Johns Hopkins, spoke about how neurons might balance two opposing roles of DISC1 in neurons: on the one hand, the protein regulates proliferation of neuronal progenitors through the GSK pathway, and on the other, it controls migration. Previous work from Kamiya and Sawa had shown that migration involves DISC1’s interaction with the centrosome and cytoskeletal proteins (see SRF related news story).

According to Kamiya’s new data, the switch between proliferation and migration seems to be the phosphorylation state of DISC1. He showed evidence that only DISC1 that is dephosphorylated at serine 710 (S710) inhibits GSK3β and activates β-catenin-mediated gene expression. In contrast, S710 phosphorylation increases binding of DISC1 with proteins involved in the centrosome pathway.

To look at the switch in cells, they compared the affinity of DISC1 for partner proteins in embryonic neurons. In cells from embryonic day 18, the time of peak migration, the interaction with centrosome-related proteins was stronger than with GSK3β. Conversely, in cells from embryonic day 14, when proliferation is prominent, DISC1 bound more avidly to GSK3. In cells with no DSIC1, the proliferation defect was rescued by an A710 mutant that was incapable of being phosphorylated, but not by the phosphomimic E710, while the opposite was true for the migration defect. The results suggest that phosphorylation of DISC1 may regulate neuronal migration. Kamiya says he is now testing candidate kinases.

Knowledge of how DISC1 participates in development is important, but how will that contribute to understanding mental illness and finding novel therapies? To achieve that goal, Kamiya says, it will be necessary to look past development, and understand DISC1 function in mature neurons. Along those lines, he touched on some new data that were further elaborated in two posters describing a DISC1 knockout recently generated by the Sawa lab in collaboration with Toshifumi Tomoda of the City of Hope/Beckman Research Institute in Duarte, California. The mice, which lack the first three exons of DISC1, do not show neuronal migration defects, but there is some question as to whether they express a C-terminal fragment of DISC1. What the researchers do know is that the mice have axonal and BDNF abnormalities. Cultured cortical neurons from the mice exhibit transport defects, including less movement and secretion of BDNF, which can be reversed by lithium treatment. In vivo, the mice show axonal pathology along the cortico-striatal tract, which was reversed by postnatal expression of DISC1. The mice also show some altered behaviors, such as defects in prepulse inhibition, and the researchers are planning to test whether lithium can rescue the behavioral deficits.

DISC1 in synapses
Nick Brandon echoed Kamiya’s sentiment that while understanding the role of DISC1 in development makes for interesting basic neuroscience, it may not be the shortest path to new treatments. For therapeutic insights, he is focused squarely on the synapse, and so was his talk. DISC1is found in the postsynaptic density in adult human brain (see SRF news story http://www.schizophreniaforum.org/new/detail.asp?id=1270). In cultured cells, the protein localizes around the centrosome at first, and as neurons mature it goes to synapses. Knockdown of DISC1 results in synaptic changes, including enlarged spines and increased surface expression of the GluR1 subunit of the AMPA glutamate receptor in cortical neurons. To understand the role of DISC1 in synapses, and its interaction with the cytoskeleton, Brandon revisited the DISC1 interactome (Camargo et al., 2007), and focused on a synaptic kinase they found there, Traf- and nck-interacting kinase (TNIK). TNIK has been localized to the post-synaptic density in rodent brain, and came up as a gene of interest in two genome wide association studies, one for schizophrenia (Shi et al., 2009) and the other for genes involved in prefrontal cortex function (Potkin et al., 2009). Brandon showed that TNIK is expressed throughout rodent brain, especially in the hippocampus. In the cortex and hippocampus, TNIK colocalized with synaptic DISC1, and both were enriched and associated in the postsynaptic density.

Mapping the interaction showed that DISC1binds TNIK in its kinase domain, while TNIK contacts DISC1 in the region of amino acids 335-348. Brandon then made cell-permeable peptides derived from the DISC1 binding site and showed they could inhibit TNIK activity in cells, and rapidly cause a decrease in key PSD proteins, including PSD95 and GluR1. Depleting TNIK using RNAi in hippocampal neurons resulted in a similar decrease in surface GlurR1, AMPA receptors and activity. Knockdown of DISC1 resulted in higher TNIK levels, and an increase in GluR1. PSD95 was not restored however, but instead was dramatically reduced.

Another partner of DISC1 and TNIK at the synapse is kalirin-7, an activator of the small GTPase Rac-1 that is required for both morphological and functional remodeling of synapses in mature cortical neurons (see SRF related news story). Kalirin-7 has previously been shown to be decreased in schizophrenia brain (see SRF related news story) and kalirin knockout mice show some schizophrenia-related behaviors (see SRF related news story). Brandon showed that kalirin-7 associates with DISC1 close to the TNIK site. A common endpoint for TNIK and kalirin-7 is regulation of the actin cytoskeleton, and in future work Brandon and colleagues will aimed at parsing how DISC1 regulates each of these proteins and through them, cytoskeletal dynamics.

DISC1 in aggregates?
Carsten Korth of the Heinrich Heine University in Düsseldorf, Germany, posed a different question. How might the non-mutant, full-length DISC1 be linked to sporadic illness? So far, people have looked at truncations or outright loss of the protein, but is it possible that a normal DISC1 might be somehow modified and become dysfunctional? Korth takes this idea from the filed of neurodegeneration, where misfolded or aggregated proteins cause dementing or paralyzing diseases including Alzheimer disease (AD), Huntington disease and Parkinson disease. In these disorders, protein aggregates range from large microscopically visible plagues such as in AD, to small aggregates visible only with immunostaining as in some polyglutamate disorders. Could a similar situation pertain to schizophrenia, where patients with negative symptoms and chronic progressive course might be displaying features of neurodegeneration?

To answer this question, Korth and colleagues asked if they could identify insoluble DISC1 in brains of people with schizophrenia. They fractionated homogenates of frozen brain from the Stanley Medical Research Institute Consortium, including 15 subjects with schizophrenia, 15 with bipolar, 15 with major depression and 15 normal. They find a subset of cases show detergent insoluble DISC1 aggregates, and the average level in disease tissue is 4 times higher than normal brain (Leliveld et al., 2008). The aggregates were not restricted to schizophrenia, but were seen in people with bipolar disease and depression as well. There was no correlation between aggregation and any specific clinical features in the patients, Korth said, nor did he see any signs of degeneration in the area where aggregated proteins were seen.

Self-association domains reside in the C-terminal end of DISC1 and a schizophrenia-associated polymorphism, S704C, enhances aggregation of Disc1 C-terminal fragments (Leliveld et al., 2009). In a poster presentation, Svenja Trossbach from the Korth lab showed the same results for full length DISC1.

By looking at proteins overexpressed in cells, or using purified recombinant protein, Korth and colleagues found that aggregated DISC1 loses its ability to associate with Ndel1, suggesting that aggregation might disrupt normal DISC1 function. A poster from Josef Kittler’s lab at University College London lent support to that idea. Presenter Talia Atkin showed that overexpression of DISC1 in cell lines or primary neurons in culture leads to perinuclear aggregation. The aggregates sequester Ndel1 and the cells show disrupted intracellular trafficking of mitochondria and endosomal mitochondrial trafficking.

Korth and colleagues also found a way to enhance DISC1 aggregation by treating cells with dopamine. Dopamine has been previously shown to modify the Parkin protein (LaVoie et al., 2005), and Korth showed that when DISC1-transfected cells were treated with dopamine, the amount of soluble protein was reduced and insoluble multimers increased. Along with the shift of DISC1 to insoluble fractions, adding dopamine caused of loss of Ndel1 interaction. An in vivo experiment using amphetamine-sensitization also increased insoluble dopamine, and the researcher found a correlation between insoluble protein levels and hyperactive behavior measured by open field travel. The model will be useful to look at behavior due to insoluble, full-length wild-type DISC1, Korth said.

Christopher Ross of Johns Hopkins University, Baltimore, Maryland, was at the minisymposium and told SRF that the aggregation data need to be replicated, but he found it the most exciting part of the session.

Posters of note
Two groups used DISC1 mice to look at gene-environment interactions, and in particular the relationship between immune responses and schizophrenia. Maternal infection is a risk factor for schizophrenia, and both groups used injection of poly dI:dC to mimic viral infection and activation of the innate immune system. Daisuke Ibi, from the lab of Kiyofumi Yamada at Nagoya University in Japan, injected neonatal mice expressing a dominate negative DISC1 construct and found synergistic effects on behaviors related to DISC1 expression and on loss of parvalbumin-positive neurons in adult mice (Ibi et al., 2010). In an independent study, Bagrat Abazyan of Johns Hopkins injected pregnant mice of the same dominant-negative DISC1 model with poly dI:dC, and found increases in inflammatory cytokines in the fetal brain 3-6 hours after injection. They also tested the effects of one upregulated cytokine, IL-6, on cortical neurons, and found that while the cytokine normally increased neurite complexity on long neurites in wild type cells, it affected mainly short neurites in cells from the brains of the domaint negative DISC1 mice. The results suggest mutant DISC1 can affect neuronal differentiation by altering the response to IL-6. The mice may provide a model for further analysis of the interactions between immune system activation and genetic susceptibility, the researchers say.—Pat McCaffrey.

Comments on Related News


Related News: Dendritic Spine Research—Putting Meat on the Bones

Comment by:  Amanda Jayne Law, SRF Advisor
Submitted 13 February 2006
Posted 13 February 2006

The formation of dendritic spines during development and their structural plasticity in the adult brain are critical aspects of synaptogenesis and synaptic plasticity. Actin is the major cytoskeletal source of dendritic spines, and polymerization/depolymerization of actin is the primary determinant of spine motility and morphogenesis. Some, but not all, postmortem studies in schizophrenia have identified reduced dendritic spine density in neurons of the hippocampal formation and dorsolateral prefrontal cortex (for review, see Honer et al., 2000); however, little is known about the underlying pathogenic mechanisms affecting synaptic function in the disease.

Many different factors and proteins are known to control dendritic spine development and remodeling (see Ethell and Pasquale, 2005). Comprehensive investigation of the effectors and signaling pathways involved in regulating actin dynamics may provide insight into the molecular mechanisms mediating altered cortical microcircuitry in the disease.

David Lewis and colleagues have previously reported reduced spine density in the basilar dendrites of pyramidal neurons in laminar III of the DLPFC (though this is not clearly a laminar-specific finding). In their current study, Hill et al. extended these investigations to examine gene expression levels for members of the RhoGTPase family of intracellular signaling molecules (e.g., Cdc42, Rac1, RhoA, Duo), and Debrin, an F-actin binding protein, all of which are critical signal transduction molecules involved in spine formation and maintenance. Their aim was to determine whether alterations in the expression of one of more molecules may underlie the reduced spine density seen in the disorder. Hill et al. report that reductions in Cdc42 and Duo mRNA are observed in the DLPFC in schizophrenia and correlate with spine density on deep layer III pyramidal neurons. This paper provides preliminary evidence that "gene expression levels of certain mRNAs encoding proteins known to be key regulators of dendritic spines are reduced in the DLPFC in schizophrenia." However, the paper also reports that these two mRNAs are reduced in lamina where significant reductions in spine density are not observed in schizophrenia. These results may suggest, as the authors discuss, that reduced expression of Cdc42 and Duo might contribute to, but is not sufficient to cause reduced, spine density.

Synaptic dysfunction has received increasing attention as a key feature of schizophrenia’s neuropathology and possibly its genetic etiology (Law et al., 2004). Neuregulin 1 (NRG1), a lead schizophrenia susceptibility gene, is known to be a critical upstream regulator of signal transduction pathways modulating cytoskeletal dynamics, playing pivotal roles in synapse formation and function. We have previously reported that isoform-specific alterations of the NRG1 gene and its primary receptor, ErbB4, are apparent in the brain in schizophrenia and related to genetic risk for the disease (Law et al, 2005a, Law et al, 2005b). Altered NRG1/ErbB4 signaling in schizophrenia may be a pathway to aberrant cortical neurodevelopment and synaptic function via dysregulation of specific intracellular signaling pathways linked to actin. The lack of significant alterations in gene expression levels for proteins such as Rac1 and RhoA in the DLPFC (gray matter, as reported by Hill and colleagues) in schizophrenia might be because the primary defect may not lie with the expression of these molecules but with the upstream modulation of their function and activity. Therefore, investigation of the proteins themselves, their phosphorylation status and activity, will be useful in understanding how genes effect molecular pathways that mediate biological risk for schizophrenia. The study of intracellular signaling cascades may be a route to a closer understanding of the biological mechanisms underpinning the association of genes such as NRG1 and ErbB4 with schizophrenia and their relationship to its neuropathology.

References:

Ethell IM, Pasquale EB. Molecular mechanisms of dendritic spine development and remodeling. Prog Neurobiol. 2005 Feb;75(3):161-205. Epub 2005 Apr 2. Review. Abstract

Honer G, Young C, and Falkai P, 2000. Synaptic Pathology in the Neuropathology of Schizophrenia, Progress and interpretation. Oxford University Press, edited by Paul J Harrison and Gareth W. Roberts, pp105-136.

Law AJ, Weickert CS, Hyde TM, Kleinman JE, Harrison PJ. Reduced spinophilin but not microtubule-associated protein 2 expression in the hippocampal formation in schizophrenia and mood disorders: molecular evidence for a pathology of dendritic spines. Am J Psychiatry. 2004 Oct;161(10):1848-55. Abstract

Law, 2005a. Soc Neurosci Abstract, SFN Annual Meeting, Washington DSC, 2005. Neuregulin1 and schizophrenia: A pathway to altered cortical circuits. Also See SfN 2005 research news: Cortical Deficits in Schizophrenia: Have Genes, Will Hypothesize.

Law 2005b ACNP Abstract, Neuropsychopharmacology, vol. 30, Supplement 1. SNPing away at NRG1 and ErbB4 gene expression in schizophrenia.

View all comments by Amanda Jayne Law

Related News: Architect of Synaptic Plasticity Links Spine Form and Function

Comment by:  Akira Sawa, SRF Advisor
Submitted 29 December 2007
Posted 29 December 2007

Synaptic disturbance in the pathology of schizophrenia is a well-established idea. Lewis’s lab has reported decreased synaptic spine density in brains from patients with schizophrenia (Glantz and Lewis, 2000). Although it is unclear whether this is primary or secondary, expression of kalirin-7-associated molecules is decreased (Hill et al., 2006). Thus, kalirin-7-associated cellular signaling in synaptic spines may have implication for the pathology of schizophrenia. In this sense, I regard the recent publication from Penzes’s lab as very interesting in schizophrenia research.

It is still unclear whether kalirin-7 may interact with genetic susceptibility factors for schizophrenia, such as ErbB4 and DISC1. Until the protein interactions are tested by co-immunoprecipitation at endogenous protein levels, as well as validated by cell staining, we cannot tell whether or not such factors are really associated with the kalirin-7 pathway. This putative protein interaction of kalirin-7 with DISC1 or ErbB4 will be an important issue to address in the future.

In Penzes’s neuronal cultures, he has focused on spine formation in pyramidal neurons, but not in interneurons. Thus, the mechanism proposed in his study will be useful to consider possible pathology in pyramidal neurons in brains of patients with schizophrenia.

View all comments by Akira Sawa

Related News: DISC1: A Matter of Life or Death for Neural Progenitors

Comment by:  Khaled Rahman
Submitted 26 March 2009
Posted 26 March 2009

Mao and colleagues present an impressive body of work implicating GSK3β/β-catenin signaling in the function of Disc1. However, several key experimental controls are missing that detract from the impact of their study, and it is unclear whether this function of Disc1 among its many others is the critical link between the t(1;11) translocation and psychopathology in the Scottish family.

The results of Mao et al. suggest that acute knockdown of Disc1 in embryonic brain causes premature exit from the proliferative cell cycle and premature differentiation into neurons. In fact, they observe fewer GFP+ cells in the VZ/SVZ and greater GFP+ cells within the cortical plate. This is in contrast to the study by Kamiya et al. (2005), in which they find that knocking down Disc1 caused greater retention of cells in the VZ/SVZ and fewer in the cortical plate, suggesting retarded migration. Although the timing of electroporation (E13 vs. E14.5) and examination (E15 vs. P2) differed between the two studies, these results are not easily reconciled.

The authors also suggest that they can rescue the deficits in proliferation by overexpressing human wild-type DISC1, stabilizing β-catenin expression, or inhibiting GSK3β activity, and thus conclude that Disc1 is acting through this pathway. This conclusion, however, rests on an error in logic. If increasing X causes an increase in Y, and decreasing Z causes a decrease in Y, this does not mean that X and Z are operating via the same mechanism. In fact, overexpressing WT-DISC1, stabilizing β-catenin, or inhibiting GSK3β activity all increase proliferation in control cells. Thus, the fact that these manipulations also work in progenitors with Disc1 silenced only tells us that these effects are independent or downstream of Disc1. What are needed are studies that show a differential sensitivity of Disc1-silenced cells to manipulations of β-catenin or GSK3β. In other words, is there a shift in the dose response curves? This is what is to be expected given that Mao et al. show changes in β-catenin levels and changes in the phosphorylation of GSK3β substrates in Disc1 silenced cells.

Furthermore, it is surprising that a restricted silencing of Disc1 in the adult dentate gyrus produces changes in affective behaviors, when total ablation of dentate neurogenesis in the adult produces little effects on depression-related behaviors (Santarelli et al., 2003; Airen et al., 2007). The fact that inhibiting GSK3β increases proliferation in both control and Disc1 knockdown animals to a similar degree suggests that the “rescue” of any behavioral deficits is independent of the drug’s effects on proliferation. Correlating measures of proliferation with behavioral performance would help address this issue.

How this study will lead to new or improved therapeutic interventions is also an open question. Lithium is well known for its mood-stabilizing properties, and this study may point to better, more efficient ways to address these symptoms. However, it is also known that lithium does little for, if not worsens, cognitive symptoms in patients (Pachet and Wisniewski, 2003), and it is this symptom domain that is in dire need of drug development.

It is also important to keep in mind that acute silencing of Disc1 in a restricted set of cells will not necessarily recapitulate the pathogenetic process of a disease-associated mutation. It remains to be seen if similar results are obtained in animal models of the Disc1 mutation (Clapcote et al., 2007; Hikida et al., 2007; Li et al., 2007).

References:

Kamiya A, Kubo K, Tomoda T, Takaki M, Youn R, Ozeki Y, Sawamura N, Park U, Kudo C, Okawa M, Ross CA, Hatten ME, Nakajima K, Sawa A. A schizophrenia-associated mutation of DISC1 perturbs cerebral cortex development. Nat Cell Biol. 2005 Dec 1;7(12):1167-78. Abstract

Santarelli, L. et al. Requirement of hippocampal neurogenesis for the behavioral effects of antidepressants. Science 301, 805–809 (2003). Abstract

Airan, R.D. et al. High-speed imaging reveals neurophysiological links to behavior in an animal model of depression. Science 317, 819-23 (2007). Abstract

Pachet AK, Wisniewski AM. The effects of lithium on cognition: an updated review. Psychopharmacology (Berl). 2003 Nov;170(3):225-34. Review. Abstract

Clapcote SJ, Lipina TV, Millar JK, Mackie S, Christie S, et al. (2007) Behavioral phenotypes of Disc1 missense mutations in mice. Neuron 54: 387–402. Abstract

Hikida T, Jaaro-Peled H, Seshadri S, Oishi K, Hookway C, et al. (2007) Dominant-negative DISC1 transgenic mice display schizophrenia-associated phenotypes detected by measures translatable to humans. Proc Natl Acad Sci U S A 104: 14501–14506. Abstract

Li W, Zhou Y, Jentsch JD, Brown RA, Tian X, et al. (2007) Specific developmental disruption of disrupted-in-schizophrenia-1 function results in schizophrenia-related phenotypes in mice. Proc Natl Acad Sci U S A 104: 18280–18285. Abstract

View all comments by Khaled Rahman

Related News: DISC1: A Matter of Life or Death for Neural Progenitors

Comment by:  Simon Lovestone
Submitted 27 March 2009
Posted 27 March 2009

This is an intriguing paper that builds on a growing body of evidence implicating wnt regulation of GSK3 signaling in psychotic illness (Lovestone et al., 2007).

It is interesting that the authors report that binding of DISC1 to GSK3 results in no change in the inhibitory Ser9 phosphorylation site of GSK3 but a change in Y216 activation site and that this resulted in effects on some but not all GSK3 substrates. This poses a challenge both in terms of understanding the role of GSK3 signaling in schizophrenia and other psychotic disorders and in drug discovery.

The authors cite some of the other evidence for regulation of GSK3 signaling in psychosis, including, for example, the evidence for a role of AKT signaling alteration in schizophrenia and lithium, an inhibitor of GSK3, as a treatment for bipolar disorder. But in both cases, AKT (Cross et al., 1995) and lithium (Jope, 2003), the effect on GSK3 is predominantly via Ser9 phosphorylation and not via Y216. The unstated implication is at least two, possibly three, mechanisms for regulation of GSK3 are all involved in psychotic illness—the auto-phosphorylation at Y216, the exogenous signal transduction regulated Ser9 site inhibition and, if the association of schizophrenia with the wnt inhibitor DKK4 we reported is true (Proitsi et al., 2008), also via the wnt signaling effects on disruption of the macromolecular complex that brings GSK3 together with β-catenin. On the one hand, this might be taken as positive evidence of a role for GSK3 in psychosis—all of its regulatory mechanisms have been implicated; therefore, the case is stronger. On the other hand, GSK3 lies at the intersection point of very many signaling pathways and so is likely to be implicated in many disorders (as it is), and the fact that in cellular and animal models related to psychosis there is no consistent effect on the enzyme is troublesome.

From a drug discovery perspective, those with GSK3 inhibitors in the pipeline will be watching this space carefully. However, it is worth noting that Mao et al. find very selective effects of DISC1 on GSK3 substrates. Despite convincing evidence of an increase in Y216 phosphorylation, which one would expect to increase activity of GSK3 against all substrates, the authors find no evidence of effects on phosphorylation of the GSK3 substrates Ngn2 or C/EBPα. This is somewhat puzzling and merits further attention, especially as in vitro direct binding of a DISC1 fragment to GSK3 inhibited the action of GSK3 on a range of substrates. Might there be more to the direct interaction of DISC1 with GSK3 than a regulation of Y216 autophosphorylation and activation? If, however, GSK3 regulation turns out to be part of the mechanism of schizophrenia or bipolar disorder, then identifying which of the substrates and which of the many activities of GSK3, including on plasticity and hence cognition (Peineau et al., 2007; Hooper et al., 2007), are important in disease will become the critical task.

References:

Lovestone S, Killick R, Di Forti M, Murray R. Schizophrenia as a GSK-3 dysregulation disorder. Trends Neurosci. 2007 Apr 1 ; 30(4):142-9. Abstract

Cross DA, Alessi DR, Cohen P, Andjelkovich M, Hemmings BA. Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature . 1995 Dec 21-28 ; 378(6559):785-9. Abstract

Jope RS. Lithium and GSK-3: one inhibitor, two inhibitory actions, multiple outcomes. Trends Pharmacol Sci . 2003 Sep 1 ; 24(9):441-3. Abstract

Proitsi P, Li T, Hamilton G, Di Forti M, Collier D, Killick R, Chen R, Sham P, Murray R, Powell J, Lovestone S. Positional pathway screen of wnt signaling genes in schizophrenia: association with DKK4. Biol Psychiatry . 2008 Jan 1 ; 63(1):13-6. Abstract

Peineau S, Taghibiglou C, Bradley C, Wong TP, Liu L, Lu J, Lo E, Wu D, Saule E, Bouschet T, Matthews P, Isaac JT, Bortolotto ZA, Wang YT, Collingridge GL. LTP inhibits LTD in the hippocampus via regulation of GSK3beta. Neuron . 2007 Mar 1 ; 53(5):703-17. Abstract

Hooper C, Markevich V, Plattner F, Killick R, Schofield E, Engel T, Hernandez F, Anderton B, Rosenblum K, Bliss T, Cooke SF, Avila J, Lucas JJ, Giese KP, Stephenson J, Lovestone S. Glycogen synthase kinase-3 inhibition is integral to long-term potentiation. Eur J Neurosci . 2007 Jan 1 ; 25(1):81-6. Abstract

View all comments by Simon Lovestone

Related News: DISC1: A Matter of Life or Death for Neural Progenitors

Comment by:  Nick Brandon (Disclosure)
Submitted 27 March 2009
Posted 30 March 2009
  I recommend the Primary Papers

Li-huei Tsai and colleagues have identified another pathway in which the candidate gene DISC1 looks to have a critical regulatory role, namely the wnt signaling pathway, in progenitor cell proliferation. In recent years we have seen that DISC1 has a vital role at the centrosome (Kamiya et al., 2005), in cAMP signaling (Millar et al., 2005), and in multiple steps of adult hippocampal neurogenesis (Duan et al., 2007). They have shown a pivotal role for DISC1 in neural progenitor cell proliferation through regulation of GSK3 signaling using a spectacular combination of cellular and in utero manipulations with shRNAs and GSK3 inhibitor compounds. These findings clearly implicate DISC1 in another “druggable” pathway but at this stage do not really identify new approach/targets, except perhaps to confirm that manipulating adult neurogenesis and the wnt pathway holds much potential hope for therapeutics. Perhaps understanding the mechanism of inhibition of GSK3 by DISC1 in more detail might reveal more novel approaches or encourage more innovative work around this pathway. In addition, I have read the other comment (by Rahman), and though I agree that this work still leaves many questions to be answered, the paper is much more significant and likely reconcilable with previous papers than appreciated. The commentary from Lovestone was very insightful and brings up additional gaps and issues with the present work. Additional experimentation I am sure will tease out more key facets of the DISC1-wnt interaction in the near future.

There are many avenues now to proceed with this work. In particular, from the DISC1-centric view, a GSK3 binding site on DISC1 overlaps with one of the critical core PDE4 binding site. Mao et al. show that residues 211 to 225 are a core part of a GSK3 binding site. Previously, Miles Houslay had shown very elegantly that residues 191-230 form a common binding site (known as common site 1) for both PDE4B and 4D families (Murdoch et al., 2007). It will be important to understand the relationship between GSK3 and PDE4 related signaling in reference to the activity of DISC1 starting at whether a trimolecular complex among DISC1-PDE4-GSK3 can form. Then it will be critical to understand the regulatory interplay among these molecules. For example, it is known that PKA can regulate GSK3 activity (Torii et al., 2008) and the interaction between DISC1 and PDE4, while both GSK3 and PKA can phosphorylate β-catenin (Taurin et al., 2006). The output of these relationships on progenitor proliferation will further deepen insights into the role of DISC1 complexes in neuronal processes. This type of situation is not really surprising for a molecule (DISC1) which has been shown to interact with >100 proteins (Camargo et al., 2007). The context of these interactions in both normal development and disease is likely to be critical to allow understanding of its complete functional repertoire.

Another area where these new findings need to be exploited is in the study of additional animal models. Though the two behavioral endpoint models used in the paper (amphetamine hyperactivity and forced swim test) provide a tantalizing glimpse of the behavioral importance of the complex, it would be critical to look in additional models relevant for schizophrenia and mood disorders. Furthermore, it will be very interesting to look at the effects of GSK3β inhibitors in some of the DISC1 animal models already available and to see if they can reverse all or a subset of reported behaviors. In reviewing a summary of the phenotypes available to date (Shen et al., 2008) there is clearly a number of lines which share the properties with mice injected with DISC1 shRNA into the dentate gyrus and would be of value to look at.

A very exciting paper which I am sure will drive additional research into understanding the role of DISC1 in psychiatry and hopefully encourage drug discovery efforts around this molecular pathway (Wang et al., 2008).

References:

1. Kamiya A, Kubo K, Tomoda T, Takaki M, Youn R, Ozeki Y, Sawamura N, Park U, Kudo C, Okawa M, Ross CA, Hatten ME, Nakajima K, Sawa A. A schizophrenia-associated mutation of DISC1 perturbs cerebral cortex development. Nat Cell Biol . 2005 Dec 1 ; 7(12):1167-78. Abstract

2. Millar JK, Pickard BS, Mackie S, James R, Christie S, Buchanan SR, Malloy MP, Chubb JE, Huston E, Baillie GS, Thomson PA, Hill EV, Brandon NJ, Rain JC, Camargo LM, Whiting PJ, Houslay MD, Blackwood DH, Muir WJ, Porteous DJ. DISC1 and PDE4B are interacting genetic factors in schizophrenia that regulate cAMP signaling. Science . 2005 Nov 18 ; 310(5751):1187-91. Abstract

3. Duan X, Chang JH, Ge S, Faulkner RL, Kim JY, Kitabatake Y, Liu XB, Yang CH, Jordan JD, Ma DK, Liu CY, Ganesan S, Cheng HJ, Ming GL, Lu B, Song H. Disrupted-In-Schizophrenia 1 regulates integration of newly generated neurons in the adult brain. Cell . 2007 Sep 21 ; 130(6):1146-58. Abstract

4. Murdoch H, Mackie S, Collins DM, Hill EV, Bolger GB, Klussmann E, Porteous DJ, Millar JK, Houslay MD. Isoform-selective susceptibility of DISC1/phosphodiesterase-4 complexes to dissociation by elevated intracellular cAMP levels. J Neurosci . 2007 Aug 29 ; 27(35):9513-24. Abstract

5. Torii K, Nishizawa K, Kawasaki A, Yamashita Y, Katada M, Ito M, Nishimoto I, Terashita K, Aiso S, Matsuoka M. Anti-apoptotic action of Wnt5a in dermal fibroblasts is mediated by the PKA signaling pathways. Cell Signal . 2008 Jul 1 ; 20(7):1256-66. Abstract

6. Taurin S, Sandbo N, Qin Y, Browning D, Dulin NO. Phosphorylation of beta-catenin by cyclic AMP-dependent protein kinase. J Biol Chem . 2006 Apr 14 ; 281(15):9971-6. Abstract

7. Camargo LM, Collura V, Rain JC, Mizuguchi K, Hermjakob H, Kerrien S, Bonnert TP, Whiting PJ, Brandon NJ. Disrupted in Schizophrenia 1 Interactome: evidence for the close connectivity of risk genes and a potential synaptic basis for schizophrenia. Mol Psychiatry . 2007 Jan 1 ; 12(1):74-86. Abstract

8. Shen S, Lang B, Nakamoto C, Zhang F, Pu J, Kuan SL, Chatzi C, He S, Mackie I, Brandon NJ, Marquis KL, Day M, Hurko O, McCaig CD, Riedel G, St Clair D. Schizophrenia-related neural and behavioral phenotypes in transgenic mice expressing truncated Disc1. J Neurosci . 2008 Oct 22 ; 28(43):10893-904. Abstract

9. Wang Q, Jaaro-Peled H, Sawa A, Brandon NJ. How has DISC1 enabled drug discovery? Mol Cell Neurosci . 2008 Feb 1 ; 37(2):187-95. Abstract

View all comments by Nick Brandon

Related News: DISC1: A Matter of Life or Death for Neural Progenitors

Comment by:  Akira Sawa, SRF Advisor
Submitted 8 April 2009
Posted 8 April 2009

Mao and colleagues’ present outstanding work sheds light on a novel function of DISC1. Because DISC1 is a multifunctional protein, the addition of new functions is not surprising. Thus, for the past several years, the field has focused on how DISC1 can have distinct functions in different cell contexts (for example, progenitor cells vs. postmitotic neurons, or developing cortex vs. adult dentate gyrus). In addition to Mao and colleagues, I understand that several groups, including ours, have obtained preliminary, unpublished evidence that DISC1 regulates progenitor cell proliferation, at least in part via GSK3β. Thus, I am very supportive of this new observation.

If there might be a missing point in this paper, it is unclear whether suppression of GSK3β occurs in several different biological contexts in brain in vivo. In other words, it is uncertain whether DISC1’s actions on GSK3β are constitutive or context-dependent. How can we reconcile differential roles for DISC1 in progenitor cells in contrast to postmitotic neurons? We have already obtained a preliminary promising answer to this question, which is currently being validated very intensively. These two phenotypes (progenitor cell control and postmitotic migration) may compensate for each other in cortical development; thus, overall cortical pathology looks milder in adults, at least in our preliminary unpublished data using DISC1 knockout mice. We are not sure how this novel function of DISC1 may account for the pathology of Scottish cases. Although I have great respect for the Scottish pioneers of DISC1 study, such as St. Clair, Blackwood, and Muir (I believe that the St. Clair et al., 1990 Lancet paper is one of the best publications in psychiatry), now is the time to pay more and more attention to the question of the molecular pathway(s) involving DISC1 in general schizophrenia (see 2009 SRF roundtable discussion). Unlike the role of APP in Alzheimer’s disease, DISC1 is not a key biological target in general schizophrenia, instead being an entry point to explore much more important targets for schizophrenia. There may be no more need to stick to DISC1 itself in the unique Scottish cases in schizophrenia research. In sum, although there may still be key missing points in this study, I wish to congratulate the authors on their outstanding work.

References:

St Clair D, Blackwood D, Muir W, Carothers A, Walker M, Spowart G, Gosden C, Evans HJ. Association within a family of a balanced autosomal translocation with major mental illness. Lancet . 1990 Jul 7 ; 336(8706):13-6. Abstract

View all comments by Akira Sawa

Related News: DISC1 Players Gird For Adult Neurodevelopment

Comment by:  Kevin J. Mitchell
Submitted 8 October 2009
Posted 8 October 2009

The seminal identification of mutations in DISC1 associated with schizophrenia and other psychiatric disorders raises several obvious questions: what does the DISC1 protein normally do? What are its biochemical and cellular functions, and what processes are affected by its mutation? How do defects in these cellular processes ultimately lead to altered brain function and psychopathology? Which brain systems are affected and how? Similar questions could be asked for the growing number of other genes that have been implicated by the identification of putatively causal mutations, including NRG1, ERBB4, NRXN1, CNTNAP2, and many copy number variants. Finding the points of biochemical or phenotypic convergence for these proteins or mutations may be key to understanding how mutations in so many different genes can lead to a similar clinical phenotype and to suggesting points of common therapeutic intervention.

The papers by Kim et al. and Enomoto et al. add more detail to the complex picture of the biochemical interactions of DISC1 and its diverse cellular functions. The links with Akt and PTEN signaling are especially interesting, given the previous implication of these proteins in schizophrenia and autism. Akt, in particular, may provide a link between Nrg1/ErbB4 signaling and DISC1 intracellular functions.

These studies also reinforce the importance of DISC1 and its interacting partners in neurodevelopment, specifically in cell migration and axonal extension. In particular, they highlight the roles of these proteins in postnatal hippocampal development and adult hippocampal neurogenesis. They also raise the question of which extracellular signals and receptors regulate these processes through these signalling pathways. The Nrg1/ErbB4 pathway has already been implicated, but there are a multitude of other cell migration and axon guidance cues known to regulate hippocampal development, some of which, for example, semaphorins, signal through the PTEN pathway.

Whether or how disruptions in these developmental processes contribute to psychopathology also remains unclear. It seems likely that the effects of mutations in any of these genes will be highly pleiotropic and have effects in many brain systems. The reported pathology in schizophrenia is not restricted to hippocampus but extends to cortex, thalamus, cerebellum, and many other regions. Similarly, while the cognitive deficits receive a justifiably large amount of attention, given that they may have the most clinical impact, motor and sensory deficits are also a stable and consistent part of the syndrome that must be explained. Pleiotropic effects on prenatal and postnatal development, as well as on adult processes, may actually be the one common thread characterizing the genes so far implicated. These new papers represent the first steps in the kinds of detailed biological studies that will be required to make explanatory links from mutations, through biochemical and cellular functions, to effects on neuronal networks and ultimately psychopathology.

View all comments by Kevin J. Mitchell

Related News: DISC1 Players Gird For Adult Neurodevelopment

Comment by:  Peter PenzesMichael Cahill
Submitted 8 October 2009
Posted 8 October 2009

DISC1 disruption by chromosomal translocation cosegregates with several neuropsychiatric disorders, including schizophrenia (Blackwood et al., 2001; Millar et al., 2000). Recent attention has focused on the effects of DISC1 on the structure and function of the dentate gyrus, one of the few brain regions that exhibit neurogenesis throughout life. The downregulation of DISC1 has several deleterious effects on the dentate gyrus, including aberrant neuronal migration (Duan et al., 2007). However, the mechanisms through which DISC1 regulates the structure and function of the dentate gyrus remain unknown. The dentate gyrus and its output to the CA3 area, the mossy fiber, show several abnormalities in schizophrenia and other neuropsychiatric diseases (Kobayashi, 2009). Thus, understanding how a gene associated with neuropsychiatric disease, DISC1, mechanistically impacts the dentate gyrus is an important question with much clinical relevance.

The recent papers by Kim et al. and Enomoto et al. characterize an interaction between DISC1 and girdin (also known as KIAA1212), and reveal how girdin, and the interaction between DISC1 and girdin, impact axon development, dendritic development, and the proper positioning of newborn neurons in the dentate gyrus. Girdin normally stimulates the function of AKT (Anai et al., 2005), and Kim et al. show that DISC1 binds to girdin and inhibits its function. Thus, the loss of DISC1 leaves girdin unopposed, resulting in excessive AKT signaling. Indeed, the developmental defects in neurons lacking DISC1 can be rescued by pharmacologically blocking the activation of an AKT downstream target. However, as shown by Enomoto et al., the loss of girdin produces deleterious effects on neuronal morphology, suggesting that a proper balance of girdin function is crucial.

Collectively, these studies thoroughly characterize the interaction between DISC1 and girdin, and shed much light on the consequences of this interaction on neuronal morphology as well as on the positioning of neurons in the dentate gyrus. The role of girdin in the pathology of neuropsychiatric diseases is unknown, and remains an interesting question for the future. Characterizing the molecules that act up- or downstream of DISC1 remains an important area of investigation and could aid the development of pharmacological interventions in the future. It’s intriguing that DISC1 acting through girdin regulates the activity of AKT as AKT1 was previously identified as a schizophrenia risk gene (Emamian et al., 2004). This suggests a convergence of multiple schizophrenia-associated genes in a shared pathway, and thus it will be important to determine if the DISC1-girdin-AKT1 pathway is particularly vulnerable in neuropsychiatric disorders.

References:

Blackwood DH, Fordyce A, Walker MT, St Clair DM, Porteous DJ, Muir WJ. Schizophrenia and affective disorders--cosegregation with a translocation at chromosome 1q42 that directly disrupts brain-expressed genes: clinical and P300 findings in a family. Am J Hum Genet . 2001 Aug 1 ; 69(2):428-33. Abstract

Millar JK, Christie S, Semple CA, Porteous DJ. Chromosomal location and genomic structure of the human translin-associated factor X gene (TRAX; TSNAX) revealed by intergenic splicing to DISC1, a gene disrupted by a translocation segregating with schizophrenia. Genomics . 2000 Jul 1 ; 67(1):69-77. Abstract

Duan X, Chang JH, Ge S, Faulkner RL, Kim JY, Kitabatake Y, Liu XB, Yang CH, Jordan JD, Ma DK, Liu CY, Ganesan S, Cheng HJ, Ming GL, Lu B, Song H. Disrupted-In-Schizophrenia 1 regulates integration of newly generated neurons in the adult brain. Cell . 2007 Sep 21 ; 130(6):1146-58. Abstract

Kobayashi K. Targeting the hippocampal mossy fiber synapse for the treatment of psychiatric disorders. Mol Neurobiol . 2009 Feb 1 ; 39(1):24-36. Abstract

Anai M, Shojima N, Katagiri H, Ogihara T, Sakoda H, Onishi Y, Ono H, Fujishiro M, Fukushima Y, Horike N, Viana A, Kikuchi M, Noguchi N, Takahashi S, Takata K, Oka Y, Uchijima Y, Kurihara H, Asano T. A novel protein kinase B (PKB)/AKT-binding protein enhances PKB kinase activity and regulates DNA synthesis. J Biol Chem . 2005 May 6 ; 280(18):18525-35. Abstract

Emamian ES, Hall D, Birnbaum MJ, Karayiorgou M, Gogos JA. Convergent evidence for impaired AKT1-GSK3beta signaling in schizophrenia. Nat Genet . 2004 Feb 1 ; 36(2):131-7. Abstract

View all comments by Peter Penzes
View all comments by Michael Cahill

Related News: DISC1 and SNAP23 Emerge In NMDA Receptor Signaling

Comment by:  Jacqueline Rose
Submitted 2 March 2010
Posted 2 March 2010
  I recommend the Primary Papers

The newly published paper by Katherine Roche and Paul Roche reports SNAP-23 expression in neuron dendrites and examines the possible role of this neuronal SNAP-23 protein. To this point, SNAP-23 has traditionally been discussed in reference to vesicle trafficking in epithelial cells (see Rodriguez-Boulan et al., 2005 for review), so it is of interest to determine the function of SNAP-23 in neurons. Suh et al. report that surface NMDA receptor expression and NMDA-mediated currents are inhibited following SNAP-23 knockdown. Further, SNAP-23 knockdown results in a specific decrease in NR2B subunit insertion; previously, the NR2B subunit has been reported to preferentially localize to recycling endosomes compared to NR2A (Lavezzari et al., 2004). Given these findings, it is reasonable to conclude that SNAP-23 may be involved in maintaining NMDA receptor surface expression possibly by binding to NMDA-specific recycling endosomes.

Interestingly, there is recent evidence that PKC-induced NMDA receptor insertion is mediated by another neuronal SNARE protein; postsynaptic SNAP-25 (Lau et al., 2010). It is possible that activity-induced NMDA receptor trafficking is mediated by SNAP-25, while baseline maintenance of NMDA receptor levels relies on SNAP-23. Other evidence to suggest a strictly regulatory role for SNAP-23 in neuronal NMDA insertion is the finding that activity-dependent receptor insertion from early endosomes has previously been reported to be restricted to AMPA-type glutamate receptors (Park et al., 2004). However, it is possible that activity-induced insertion of AMPA receptors occurs via a distinct endosome pool than NMDA receptors; AMPA and NMDA receptor trafficking has been reported to proceed by distinct vesicle trafficking pathways (Jeyifous et al., 2009).

Although SNAP-23 may not be involved in activity-dependent early endosome receptor trafficking, it is possible that SNAP-23 operates in other pathways linked to activity-induced NMDA receptor trafficking. For instance, SNAP-23 may be the SNARE protein by which lipid raft shuttling of NMDA receptors occurs. SNAP-23 has been found to preferentially associate with lipid rafts over SNAP-25 in PC12 cells (Salaün et al., 2005). As well, NMDA receptors have been found to associate with lipid raft associated proteins flotilin-1 and -2 in neurons (Swanwick et al., 2009). Lipid raft trafficking of NMDA receptors to post-synaptic densities has been reported to follow global ischemia (Besshoh et al., 2005), and the possibility remains that under certain circumstances, NMDA trafficking occurs by lipid raft association to SNAP-23.

Taken together, the discovery of post-synaptic SNARE proteins offers several avenues of research to determine their roles and functions in glutamatergic synapse organization. Further, investigating disruption of synaptic receptor organization presents several possibilities for potential etiologies of disorders linked to compromised glutamate signaling like schizophrenia.

References:

Besshoh, S., Bawa, D., Teves, L., Wallace, M.C. and Gurd, J.W. (2005). Increased phosphorylation and redistribution of NMDA receptors between synaptic lipid rafts and post-synaptic densities following transient global ischemia in the rat brain. Journal of Neurochemistry, 93: 186-194. Abstract

Jeyifous, O., Waites, C.L., Specht, C.G., Fujisawa, S., Schubert, M., Lin, E.I., Marshall, J., Aoki, C., de Silva, T., Montgomery, J.M., Garner, C.C. and Green, W.N. (2009). SAP97 and CASK mediate sorting of NMDA receptors through a previously unknown secretory pathway. Nature Neuroscience, 12: 1011-1019. Abstract

Lau, C.G., Takayasu, Y., Rodenas-Ruano, A., Paternain, A.V., Lerma, J., Bennet, M.V.L. and Zukin, R.S. (2010). SNAP-25 is a target of protein kinase C phosphorylation critical to NMDA receptor trafficking. Journal of Neuroscience, 30: 242-254. Abstract

Lavezzari, G., McCallum, J., Dewey, C.M. and Roche, K.W. (2004). Subunit-specific regulation of NMDA receptor endocytosis. Journal of Neuroscience, 24: 6383-6391. Abstract

Park, M., Penick, E.C., Edward, J.G., Kauer, J.A. and Ehlers, M.D. (2004). Recycling endosomes supply AMPA receptors for LTP. Science, 305: 1972-1975. Abstract

Rodriguez-Boulan, E., Kreitzer, G. and Müsch, A. (2005) Organization of vesicular trafficking in epithelia. Nature Reviews: Molecular Cell Biology, 6: 233-247. Abstract

Salaün, C., Gould, G.W. and Chamberlain, L.H. (2005). The SNARE proteins SNAP-25 and SNAP-23 display different affinities for lipid rafts in PC12 cells. Journal of Biological Chemistry, 280: 1236-1240. Abstract

Suh, Y.H., Terashima, A., Petralia, R.S., Wenthold, R.J., Isaac, J.T.R., Roche, K.W. and Roche, P.A. (2010). A neuronal role for SNAP-23 in postsynaptic glutamate receptor trafficking. Nat Neurosci. 2010 Mar;13(3):338-43. Abstract

Swanwick, C.C., Shapiro, M.E., Chang, Y.Z. and Wenthold, R.J. (2009). NMDA receptors interact with flotillin-1 and -2, lipid raft-associated proteins. FEBS Letters, 583: 1226-1230. Abstract

View all comments by Jacqueline Rose

Related News: Stem Cells and TALENs Bring DISC1 Back in the News

Comment by:  Kristen Brennand
Submitted 18 August 2014
Posted 19 August 2014

Johns Hopkins researchers, co-led by Guo-li Ming and Hongjun Song, have done it again! In their second major publication of the summer, this time in Nature, Wen et al. convincingly demonstrate that in human neurons, mutant disrupted in schizophrenia 1 (DISC1) depletes wild-type DISC1 protein, leading to global changes in gene expression and disrupted synaptic vesicle release.

That DISC1 plays a critical role in neuronal development is well established; mutations have been extensively studied in rodents and shown to lead to either accelerated neuronal maturation (Duan et al., 2007) or reduced neurite outgrowth and neuronal connectivity (Kamiya et al., 2005; Pletnikov et al., 2007), depending on the timing and neural subtype expressing mutant protein. More controversial is whether DISC1 plays a causal role in human cases of schizophrenia (Niculescu, 2014; Porteous et al., 2014; Sullivan, 2013), beyond a single t[1;11] [q42.1;q14.3] Scottish pedigree (Millar et al., 2000). In fact, whether the particular frameshift deletion of DISC1 studied in this report, associated with psychosis in one American pedigree, is pathogenic had remained contentious (Green et al., 2006).

Toward resolving these debates, this paper is a tour de force. Beyond just comparing human induced pluripotent stem cell (hiPSC)-derived neurons from two DISC1 patients with differing diagnoses (schizophrenia and major depression), two unaffected related controls, and one unaffected unrelated control, this group produced perfect isogenic controls through TALEN-based gene editing. DISC1 mutant neurons exhibit increased soma size and total dendritic length, together with decreased glutamatergic synapse number and activity. Critically, correction of the DISC1 locus in a patient hiPSC line perfectly rescued these neuronal phenotypes, while introducing the DISC1 frameshift mutation into two control hiPSC lines was sufficient to recapitulate the neuronal defects observed in patients. Given the high penetrance of this DISC1 mutation and the perfect phenotypic amelioration/recapitulation with TALEN-targeted corrections/mutations, the results clearly demonstrate phenotypic effects resulting from the frameshift DISC1 mutation.

Even more intriguingly, the authors demonstrate that mutant DISC1 is capable of depleting wild-type DISC1 levels, likely by increasing wild-type DISC1 ubiquitination and also causing large-scale transcriptional dysregulation in human neurons. Mutant DISC1 seems to function as a hub for transcriptional regulation of many genes (>89) implicated in psychiatric disorders.

The major criticism that this report will encounter will not be the quality of the data presented, which is remarkable in its thoroughness, but rather the persistent condemnation of DISC1 as a rare variant not relevant to the vast majority of schizophrenia cases. The case well made in this paper is that DISC1 is a key hub protein, interacting with and regulating the transcription of any number of genes likely contributing to disease risk across a wide range of psychiatric patients.

References:

Duan X, Chang JH, Ge S, Faulkner RL, Kim JY, Kitabatake Y, Liu XB, Yang CH, Jordan JD, Ma DK, Liu CY, Ganesan S, Cheng HJ, Ming GL, Lu B, Song H. Disrupted-In-Schizophrenia 1 regulates integration of newly generated neurons in the adult brain. Cell . 2007 Sep 21 ; 130(6):1146-58. Abstract

Green EK, Norton N, Peirce T, Grozeva D, Kirov G, Owen MJ, O'Donovan MC, Craddock N. Evidence that a DISC1 frame-shift deletion associated with psychosis in a single family may not be a pathogenic mutation. Mol Psychiatry . 2006 Sep ; 11(9):798-9. Abstract

Kamiya A, Kubo K, Tomoda T, Takaki M, Youn R, Ozeki Y, Sawamura N, Park U, Kudo C, Okawa M, Ross CA, Hatten ME, Nakajima K, Sawa A. A schizophrenia-associated mutation of DISC1 perturbs cerebral cortex development. Nat Cell Biol . 2005 Dec ; 7(12):1167-78. Abstract

Millar JK, Wilson-Annan JC, Anderson S, Christie S, Taylor MS, Semple CA, Devon RS, St Clair DM, Muir WJ, Blackwood DH, Porteous DJ. Disruption of two novel genes by a translocation co-segregating with schizophrenia. Hum Mol Genet . 2000 May 22 ; 9(9):1415-23. Abstract

Niculescu AB. DISCovery in psychiatric genetics. Mol Psychiatry . 2014 Feb ; 19(2):145. Abstract

Pletnikov MV, Xu Y, Ovanesov MV, Kamiya A, Sawa A, Ross CA. PC12 cell model of inducible expression of mutant DISC1: new evidence for a dominant-negative mechanism of abnormal neuronal differentiation. Neurosci Res . 2007 Jul ; 58(3):234-44. Abstract

Porteous DJ, Thomson PA, Millar JK, Evans KL, Hennah W, Soares DC, McCarthy S, McCombie WR, Clapcote SJ, Korth C, Brandon NJ, Sawa A, Kamiya A, Roder JC, Lawrie SM, McIntosh AM, St Clair D, Blackwood DH. DISC1 as a genetic risk factor for schizophrenia and related major mental illness: response to Sullivan. Mol Psychiatry . 2014 Feb ; 19(2):141-3. Abstract

Sullivan PF. Questions about DISC1 as a genetic risk factor for schizophrenia. Mol Psychiatry . 2013 Oct ; 18(10):1050-2. Abstract

View all comments by Kristen Brennand

Related News: Stem Cells and TALENs Bring DISC1 Back in the News

Comment by:  Carsten Korth, SRF Advisor
Submitted 19 August 2014
Posted 19 August 2014

The paper by Wen et al. in Nature is strong in two aspects: First, methodologically the researchers generate optimal isogenic controls of mutant, patient-derived iPS cell lines through TALEN technology, thereby enabling demonstration of (mutant) gene effects in an identical control genetic background and circumventing the use of "normal" controls from independent, apparently healthy individuals where we can never be sure about the future life trajectory of brain disease incidence. Second, by using neurons differentiated from these iPSCs, they demonstrate synaptic pathology and widespread regulation of mental illness-related genes for a DISC1 mutant that had been identified previously in an American pedigree (Sachs et al., 2005).

Based on convincing methodology, and in human neurons, the paper therefore demonstrates a causal relationship of DISC1 to synaptic pathology. The paper also demonstrates that the mutation that could not be adequately linked to disease phenotypes by statistical means (low case numbers; see also Green et al., 2006) is very likely to play a role in developing mental illness phenotypes after all.

Research on the rare gene DISC1 has recently been criticized (Sullivan, 2013; Porteous et al., 2014) as not fitting into the prevalent "common gene-common disease" view boosted by success in GWAS studies (Schizophrenia Working Group of the Psychiatric Genomics Consortium, 2013). Lessons learned from Alzheimer's disease, however, have taught us that the study of rare genes that may not appear in GWAS screens can be extremely rewarding (Bertram et al., 2010). The paper by Wen et al. demonstrates the value of such rare gene research for elucidating cellular disease pathways, supports a role for DISC1 in key components of brain function, and hints at DISC1's role as a master regulator for mental illness-related genes.

It is intriguing that the authors show insolubility of the mutant DISC1 protein that segregates the wild-type soluble form when overexpressed in HEK cells. These results support a view that insoluble DISC1 can contribute to DISC1-dependent cellular pathology (Leliveld et al., 2008; Ottis et al., 2011). In summary, Wen et al. provide an excellent cellular model to further explore human DISC1-specific cellular pathology and DISC1-dependent cellular pathways (Hennah and Porteous, 2009), and help to define DISC1-dependent mental illness, or DISC1opathies (Korth, 2012).

References:

Sachs NA, Sawa A, Holmes SE, Ross CA, DeLisi LE, Margolis RL. A frameshift mutation in Disrupted in Schizophrenia 1 in an American family with schizophrenia and schizoaffective disorder. Mol Psychiatry . 2005 Aug ; 10(8):758-64. Abstract

Sullivan PF. Questions about DISC1 as a genetic risk factor for schizophrenia. Mol Psychiatry . 2013 Oct ; 18(10):1050-2. Abstract

Porteous DJ, Thomson PA, Millar JK, Evans KL, Hennah W, Soares DC, McCarthy S, McCombie WR, Clapcote SJ, Korth C, Brandon NJ, Sawa A, Kamiya A, Roder JC, Lawrie SM, McIntosh AM, St Clair D, Blackwood DH. DISC1 as a genetic risk factor for schizophrenia and related major mental illness: response to Sullivan. Mol Psychiatry . 2014 Feb ; 19(2):141-3. Abstract

Schizophrenia Working Group of the Psychiatric Genomics Consortium. Biological insights from 108 schizophrenia-associated genetic loci. Nature . 2014 Jul 24 ; 511(7510):421-7. Abstract

Bertram L, Lill CM, Tanzi RE. The genetics of Alzheimer disease: back to the future. Neuron . 2010 Oct 21 ; 68(2):270-81. Abstract

Leliveld SR, Bader V, Hendriks P, Prikulis I, Sajnani G, Requena JR, Korth C. Insolubility of disrupted-in-schizophrenia 1 disrupts oligomer-dependent interactions with nuclear distribution element 1 and is associated with sporadic mental disease. J Neurosci . 2008 Apr 9 ; 28(15):3839-45. Abstract

Ottis P, Bader V, Trossbach SV, Kretzschmar H, Michel M, Leliveld SR, Korth C. Convergence of two independent mental disease genes on the protein level: recruitment of dysbindin to cell-invasive disrupted-in-schizophrenia 1 aggresomes. Biol Psychiatry . 2011 Oct 1 ; 70(7):604-10. Abstract

Hennah W, Porteous D. The DISC1 pathway modulates expression of neurodevelopmental, synaptogenic and sensory perception genes. PLoS One . 2009 ; 4(3):e4906. Abstract

Korth C. Aggregated proteins in schizophrenia and other chronic mental diseases: DISC1opathies. Prion . 2012 Apr-Jun ; 6(2):134-41. Abstract

View all comments by Carsten Korth

Related News: Stem Cells and TALENs Bring DISC1 Back in the News

Comment by:  Nick Brandon (Disclosure)
Submitted 18 August 2014
Posted 19 August 2014

The lab of Hongjun Song and Guo-Li Ming have provided a detailed analysis and characterization of a set of iPS cells and derived "cortical" neurons from a small pedigree where a frameshift mutation in the gene DISC1 is purportedly linked to psychiatric illness. There are a number of aspects of this study that are worth highlighting.

Critically, it is important to put the pedigree into some context. The authors state that this American pedigree is very similar to the original Scottish pedigree that led to the cloning of DISC1 (Millar et al., 2000). But this is not the case. The molecular insults in the two pedigrees are very different, while the four base-pair frameshift deletion has been found outside this pedigree. Specifically, the mutation was found in controls and not in patients in a reasonably sized study from the Cardiff group in 2006 (Green et al., 2006). The link between disease causation and the mutation has to be debated.

Where I would like to focus, though, is on the insights into DISC1 biology. I was reasonably impressed by the comparison of mutant and control lines with their isogenic equivalents in some experiments. In particular, the rescue or creation of the deficit in DISC1 protein was a simple but powerful readout of the mutation. This brings a required level of rigor to this type of work.

The other major observation was a clear effect on synapses with the mutant protein. This is not surprising considering previous studies of the protein (Brandon and Sawa, 2011). Observations from this paper, which were minimally characterized, regard the mechanisms by which DISC1 protein levels are decreased with this mutation and how this is specifically related to the decrease in synaptic puncta. The brief data in the supplement showing elevated ubiquitination of wild-type DISC1, perhaps due to changes in protein solubility, are probably critical observations for DISC1 biology and should be an area of further follow up. A role for proteostasis as a driver in mental disorders is growing (Korth, 2012). But at the moment it is not clear that the decreases in protein levels are due to this mechanism or related to changes in neuronal differentiation. Again, for context, it is thought that the expression of DISC1 is quite variable in early development. One of the early studies worth revisiting showed that Disc1 protein levels are regulated in a very dynamic way in the development of the mouse brain (Schurov et al., 2004). So it would be critical to understand whether the authors are measuring a change in development staging or an intrinsic change in the protein.

Finally, the apparent impact of the DISC1 mutation on the transcriptome of the cells was another key observation. I would have liked to have seen RNAseq data from all of the lines plus isogenic controls. Recent publications have suggested that transcriptional variation in iPS derived lines is due to the genetic background (Rouhani et al., 2014), and this would have been an opportunity to analyze this. It would have been of great interest to compare the series of control lines and isogenic control lines, and understand differences between them. This might have also allowed DISC1 mutant effects to be isolated more specifically.

Altogether, though, this is an important paper in the evolution of the use of iPSCs in the study of mental illness and in understanding DISC1 biology in this context.

References:

Brandon NJ, Sawa A. Linking neurodevelopmental and synaptic theories of mental illness through DISC1. Nat Rev Neurosci . 2011 Dec ; 12(12):707-22. Abstract

Green EK, Norton N, Peirce T, Grozeva D, Kirov G, Owen MJ, O'Donovan MC, Craddock N. Evidence that a DISC1 frame-shift deletion associated with psychosis in a single family may not be a pathogenic mutation. Mol Psychiatry . 2006 Sep ; 11(9):798-9. Abstract

Korth C. Aggregated proteins in schizophrenia and other chronic mental diseases: DISC1opathies. Prion . 2012 Apr-Jun ; 6(2):134-41. Abstract

Millar JK, Wilson-Annan JC, Anderson S, Christie S, Taylor MS, Semple CA, Devon RS, St Clair DM, Muir WJ, Blackwood DH, Porteous DJ. Disruption of two novel genes by a translocation co-segregating with schizophrenia. Hum Mol Genet . 2000 May 22 ; 9(9):1415-23. Abstract

Rouhani F, Kumasaka N, de Brito MC, Bradley A, Vallier L, Gaffney D. Genetic background drives transcriptional variation in human induced pluripotent stem cells. PLoS Genet . 2014 Jun ; 10(6):e1004432. Abstract

Schurov IL, Handford EJ, Brandon NJ, Whiting PJ. Expression of disrupted in schizophrenia 1 (DISC1) protein in the adult and developing mouse brain indicates its role in neurodevelopment. Mol Psychiatry . 2004 Dec ; 9(12):1100-10. Abstract

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Related News: Stem Cells and TALENs Bring DISC1 Back in the News

Comment by:  Atsushi Kamiya
Submitted 25 August 2014
Posted 25 August 2014

Ming and Song's group has recently reported an exciting biological characterization of forebrain neurons derived from iPS cells that are produced from the American pedigree where individuals have a frameshift mutation of disrupted in schizophrenia 1 (DISC1), which is associated with mental disorders (Sachs et al., 2005). In this study (Wen et al., 2014), the authors demonstrated deficits in glutamatergic synapses in iPS cell-derived neurons with a frameshift DISC1 mutation. These results are consistent with previous studies showing roles for DISC1 in synaptic function in rodent models (Hayashi-Takagi et al., 2010; Hayashi-Takagi et al., 2014; Wang et al., 2011). The authors also conducted molecular profiling via RNA sequencing in mutant DISC1 neurons. The data indicated altered expression of multiple genes that are involved in not only synaptic functions, but also neuronal development. These data successfully confirm the well-established concept that DISC1 is a multifactorial protein and is involved in diverse developmental cellular processes (Brandon and Sawa, 2011). It is of great interest to test whether iPS-derived neurons from the American pedigree display developmental cellular phenotypes that match those reported in rodent models, such as deficits in cell proliferation and neuronal migration (Kamiya et al., 2005; Mao et al., 2009; Ishizuka et al., 2011).

Notably, the pathological role of the frameshift mutation of DISC1 was confirmed by characterizing different types of isogenic iPS cells with this mutation introduced by TALEN technology. Synaptic deficits caused by the frameshift mutation were rescued by wild-type DISC1 protein expression. Self-dimerization of DISC1 and its aggregated biochemical phenotype were previously reported (Kamiya et al., 2005; Leliveld et al., 2008). Based on the data from in vitro biochemical experiments, the authors propose that depletion of soluble DISC1 by interaction of frameshift mutant DISC1 and wild-type DISC1 might be a pathological mechanism underlying the effect of the frameshift mutation. One should be cautious in interpreting the data, as Craddock and colleagues reported that a frameshift mutation in DISC1 found in the American pedigree has been found in control subjects (Green et al., 2006). Thus, further genetic characterization of this mutation is awaited in order to make a conclusive statement of the potential pathological role of this frameshift mutation. As the authors described, a major question in biological psychiatry is how iPS cell technology could be utilized to obtain novel insights which could not be achieved by using animal models. Further investigation of DISC1 using iPS technology in more translational ways is of particular interest for the next major breakthrough in our field.

References:

Wen Z, Nguyen HN, Guo Z, Lalli MA, Wang X, Su Y, Kim NS, Yoon KJ, Shin J, Zhang C, Makri G, Nauen D, Yu H, Guzman E, Chiang CH, Yoritomo N, Kaibuchi K, Zou J, Christian KM, Cheng L, Ross CA, Margolis RL, Chen G, Kosik KS, Song H, Ming GL. Synaptic dysregulation in a human iPS cell model of mental disorders. Nature. 2014 Aug 17. Abstract

Sachs NA, Sawa A, Holmes SE, Ross CA, DeLisi LE, Margolis RL. A frameshift mutation in Disrupted in Schizophrenia 1 in an American family with schizophrenia and schizoaffective disorder. Mol Psychiatry. 2005 Aug;10(8):758-64. Abstract

Hayashi-Takagi A, Takaki M, Graziane N, Seshadri S, Murdoch H, Dunlop AJ, Makino Y, Seshadri AJ, Ishizuka K, Srivastava DP, Xie Z, Baraban JM, Houslay MD, Tomoda T, Brandon NJ, Kamiya A, Yan Z, Penzes P, Sawa A. Disrupted-in-Schizophrenia 1 (DISC1) regulates spines of the glutamate synapse via Rac1. Nat Neurosci. 2010 Mar; 13(3):327-32. Abstract

Hayashi-Takagi A, Araki Y, Nakamura M, Vollrath B, Duron SG, Yan Z, Kasai H, Huganir RL, Campbell DA, Sawa A. PAKs inhibitors ameliorate schizophrenia-associated dendritic spine deterioration in vitro and in vivo during late adolescence. Proc Natl Acad Sci U S A. 2014 Apr 29; 111(17):6461-6. Abstract

Wang Q, Charych EI, Pulito VL, Lee JB, Graziane NM, Crozier RA, Revilla-Sanchez R, Kelly MP, Dunlop AJ, Murdoch H, Taylor N, Xie Y, Pausch M, Hayashi-Takagi A, Ishizuka K, Seshadri S, Bates B, Kariya K, Sawa A, Weinberg RJ, Moss SJ, Houslay MD, Yan Z, Brandon NJ. The psychiatric disease risk factors DISC1 and TNIK interact to regulate synapse composition and function. Mol Psychiatry. 2011 Oct; 16(10):1006-23. Abstract

Brandon NJ, Sawa A. Linking neurodevelopmental and synaptic theories of mental illness through DISC1. Nat Rev Neurosci. 2011 Dec; 12(12):707-22. Abstract

Kamiya A, Kubo K, Tomoda T, Takaki M, Youn R, Ozeki Y, Sawamura N, Park U, Kudo C, Okawa M, Ross CA, Hatten ME, Nakajima K, Sawa A. A schizophrenia-associated mutation of DISC1 perturbs cerebral cortex development. Nat Cell Biol. 2005 Dec; 7(12):1167-78. Abstract

Mao Y, Ge X, Frank CL, Madison JM, Koehler AN, Doud MK, Tassa C, Berry EM, Soda T, Singh KK, Biechele T, Petryshen TL, Moon RT, Haggarty SJ, Tsai LH. Disrupted in schizophrenia 1 regulates neuronal progenitor proliferation via modulation of GSK3beta/beta-catenin signaling. Cell. 2009 Mar 20; 136(6):1017-31. Abstract

Ishizuka K, Kamiya A, Oh EC, Kanki H, Seshadri S, Robinson JF, Murdoch H, Dunlop AJ, Kubo K, Furukori K, Huang B, Zeledon M, Hayashi-Takagi A, Okano H, Nakajima K, Houslay MD, Katsanis N, Sawa A. DISC1-dependent switch from progenitor proliferation to migration in the developing cortex. Nature. 2011 May 5; 473(7345):92-6. Abstract

Kamiya A, Kubo K, Tomoda T, Takaki M, Youn R, Ozeki Y, Sawamura N, Park U, Kudo C, Okawa M, Ross CA, Hatten ME, Nakajima K, Sawa A. A schizophrenia-associated mutation of DISC1 perturbs cerebral cortex development. Nat Cell Biol. 2005 Dec; 7(12):1167-78. Abstract

Leliveld SR, Bader V, Hendriks P, Prikulis I, Sajnani G, Requena JR, Korth C. Insolubility of disrupted-in-schizophrenia 1 disrupts oligomer-dependent interactions with nuclear distribution element 1 and is associated with sporadic mental disease. J Neurosci. 2008 Apr 9; 28(15):3839-45. Abstract

Green EK, Norton N, Peirce T, Grozeva D, Kirov G, Owen MJ, O'Donovan MC, Craddock N. Evidence that a DISC1 frame-shift deletion associated with psychosis in a single family may not be a pathogenic mutation. Mol Psychiatry. 2006 Sep; 11(9):798-9. Abstract

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