Scientists link schizophrenia risk gene to early cortical neuron changes

Researchers at King's College London have identified the biological nature and timing of changes in human cortical neurons caused by altering activity of a schizophrenia-associated gene in developing human neurons. This discovery links a genetic risk factor to cellular changes in neurons; an essential step for understanding the neurobiology of this mental illness and developing future treatments. 

Schizophrenia is estimated to be one of the most heritable psychiatric conditions, with a strong developmental aspect. Large scale human genomic studies have identified many genetic variants which are thought to increase the likelihood of schizophrenia. 

However, the link between these genetic risk variants and the underlying neurobiology of schizophrenia is less well understood. Addressing this knowledge gap provides vital information that could ultimately help develop therapies for the disorder.

This new research, published in Science Advances, from neuroscientists at the Institute of Psychiatry, Psychology & Neuroscience (IoPPN), starts to bridge the knowledge gap between genetics and their neural consequences that lead to symptoms of schizophrenia. 

While previous large-scale genetic studies have identified genetic risk factors for schizophrenia, they don't tell you when in development that gene is active or which cell type it's expressed in. To get at this information we needed to use precision functional genomics."

Professor Deepak Srivastava, Professor of Molecular Neuroscience at IoPPN King's College London and joint senior author on the paper

Relatively little is known about the mechanism of the first schizophrenia-related gene to be identified from genomic data, ZNF804A. The study identifies a specific type of neuron where ZNF804A is most active in an important developmental window. The findings also establish a novel link between two previously identified cellular processes associated with the gene: synaptic regulation and protein production regulation. 

Dr Laura Sichlinger, Research Fellow at University of Pennsylvania and first author on the study said: "Schizophrenia is a highly complex disorder. It has both a genetic and environmental component. There are 287 loci so far identified by genomic studies in humans. To be able to understand what the genes normally do in neurons is a step forward in understanding the biology of the disorder." 

Brain development is a carefully coordinated process triggered by sequentially activated genes that choreograph the precise maturation of different types of neurons and support cells in the brain. To understand developmental disorders, it is essential to identfy the timing of gene activation. 

The study confirmed that ZNF804A is most active early in development, consistent with previous studies that showed it to be highly expressed in the brain during the second trimester of neurodevelopment. 

The new research uncovered that ZNF804A was most active in glutamatergic neurons in this developmental period. Crucially, this helped the researchers focus their investigation on this type of neuron, at this particular developmental stage. 

To understand how ZNF804A contributes to the underlying neurobiology and ultimately symptoms of schizophrenia, researchers prevented the gene from functioning as it would normally in these glutamatergic neurons. To do this they employed a gene-editing approach called CRISPR-Cas9. This method works by cutting out part of the DNA in a specific gene, meaning it will be less able to be translated in its corresponding protein. Essentially, it will be able to do less of its normal function in the cell. 

By looking at the changes that happened after interfering with ZNF804A, researchers could infer what the gene might be doing in development and what types of cellular processes might be altered in neurons with schizophrenia-related mutations. 

Scientists then used a microscope to look at the junctions, called synapses, between neurons with supressed ZNF804A gene activity. These junctions are run by a series of proteins sitting on the neuronal membrane. Some sit on the neuron sending the signal; some on the neuron receiving the signal. Changes in the numbers of these synaptic proteins can impact how the neurons send and receive signals. 

The microscopy images revealed that there were more proteins at the synapses between the glutamatergic neurons, suggesting they might be more electrically excitable than normal. 

This was confirmed by chemically stimulating the neurons causing them to be more electrically active. The neurons which had less ZNF804A gene responded more than normal ones. 

Some of the proteins that sit at the synapse can be created through a process called 'protein translation' in which a biological blueprint (called mRNA) of the protein is read in, and the corresponding protein is produced. Normally if more proteins are being made in a neuron, scientists will see evidence of more translation. 

Neurons are cells with distinctive shapes, much like trees with many branching projections. The junctions between neurons can form at many parts of the neuron but often lie on the smallest branches called dendrites. To get proteins to these synapses, neurons must transport ribosomes (the machinery that builds new proteins) to the ends of the dendrite branch. This provides an ideal way to regulate how much protein is made at specific neuronal junctions: by controlling where the ribosomes are, and how many are available to make new proteins. 

The schizophrenia risk gene ZNF804A has previously been associated with cells' protein translation machinery. However, it was unknown how this related to links to synapses and signalling between neurons. 

The new study found that the neurons with impaired ZNF804A had more synapses and they had more protein production locally in their dendrites, providing a crucial link between these two cellular functions of ZNF804A. This paves the way towards a comprehensive mechanistic understanding of the role this gene plays in neuronal development. 

Professor Anthony Vernon, Professor of Neuropsychopharmacology at IoPPN, King's College London and joint senior author on the paper said: "We want to stress that these specific genetic manipulations of developing neurons do not mimic the full complement of genetic risk linked to schizophrenia. Rather, they are a tool that allow us to understand what specific risk genes, in this case, ZNF804A control in a cell and developmental timepoint specific manner. This in turn illuminates the biological processes and pathways that may be affected by specific schizophrenia-linked genetic mutations, such as those in ZNF804A. The next step is to use these tools at scale to ask whether and how the diverse array of risk genes linked to schizophrenia may converge on similar pathways and produce similar phenotypes."

This research was funded by the UK Medical Research Council (MRC Centre for Neurodevelopmental Disorders, MRC Doctoral Training Partnership), Royal Society UK, Brain and Behavior Foundation and the National Centre for the Replacement, Refinement and Reduction of Animals in Research.