Transcriptomic analysis of postmortem brain identifies dysregulated splicing events in novel candidate genes for schizophrenia

Abstract

The diverse spatial and temporal expression of alternatively spliced transcript isoforms shapes neurodevelopment and plays a major role in neuronal adaptability. Although alternative splicing is extremely common in the brain, its role in mental illnesses such as schizophrenia has received little attention. To examine this relationship, postmortem brain tissue was obtained from 20 individuals with schizophrenia (SZ) and 20 neuropsychiatrically normal comparison subjects. Gray matter samples were extracted from two brain regions implicated in the disorder: Brodmann Area 10 and caudate. Affymetrix Human Gene 1.0 ST arrays were used on four subjects per group to attain an initial profile of differential expression of transcribed elements within and across brain regions in SZ. Numerous genes of interest with altered mRNA transcripts were identified by microarray through the differential expression of particular exons and 3' untranslated regions (UTRs) between diagnostic groups. Select microarray results--including dysregulation of ENAH exon 11a and CPNE3 3'UTR--were verified by qRTPCR and replicated in the remaining independent sample of 16 SZ patients and 16 normal comparison subjects. These results, if further replicated, clearly illustrate the importance of Identifying transcriptomic variants in expression studies, and implicate novel candidate genes in the disorder.

Figure 1. Exonic Expression and Alternative 3′UTR Usage of CPNE3in: A) BA10; and B) CAUD

Microarray results of differential probe expression of Copine3 (CPNE3) in Brodman Area 10 (A) and Caudate (B) in postmortem brain tissue samples from normal control subjects (NC) (n=4) and individuals with schizophrenia (SZ) (n=4). The interaction of diagnosis and exon ID was highly significant in both brain regions; in fact, it was the only gene for which a Bonferroni-corrected threshold for significance of this term was met. *There was a statistically significant difference in probe-level expression between diagnostic groups at the most distal end of the 3′UTR in both BA10 (p=9.7e-4) and CAUD (p=4.0e-5), indicating that a truncated transcript may be expressed in SZ patients.

Figure 2. Exonic Expression and Alternative Splicing of ENAH in: A) BA10; and B) CAUD

Microarray results of differential exon expression of Enabled Homolog (ENAH) in Brodman Area 10 (A) and Caudate (B) in postmortem brain tissue samples from normal control subjects (NC) (n=4) and individuals with schizophrenia (SZ) (n=4). The interaction of diagnosis and exon ID was significant only in BA10, not CAUD, indicating the possibility of regionally specific differential splice-variant expression between the groups. *There was a statistically significant difference in expression between diagnostic groups at exon 11a in BA10 (p=0.047) but not CAUD (p=0.489), indicating increased expression of the short (11a) isoform in SZ patients in BA10 only.