NOS1AP protein levels are altered in BA46 and cerebellum of patients with schizophrenia

Dear Editors, Brzustowicz and colleagues (2004) identified significant linkage disequilibrium between schizophrenia and markers within the gene encoding nitric oxide synthase 1 (neuronal; NOS1) adaptor protein (NOS1AP; also termed carboxyl-terminal PDZ ligand of nNOS or CAPON). Quantitative real-time PCR (qRT-PCR) analysis of mRNA from human postmortem dorsolateral prefrontal cortex further revealed that expression of the short isoform of the NOS1AP gene (NOS1AP-S) is significantly increased in patients with schizophrenia (Xu, et al., 2005). More recently, the group also identified a functional risk allele within NOS1AP and showed that this change increased NOS1AP mRNA expression in a cell culture system (Wratten, et al., 2009). Despite these recent reports establishing linkage between NOS1AP and schizophrenia, little is known about NOS1AP protein expression in the brains of affected patients. Previous reports described two distinct NOS1AP isoforms: full-length NOS1AP-L (10 exons, ~75kD) and NOS1AP-S, a C-terminal specific transcript that encodes only the PDZ domain (Jaffrey, et al., 1998; Xu, et al., 2005). We have now identified a novel isoform, NOS1AP-S’ (Figure 1A), in mouse and human tissue using qRT-PCR (data not shown). To evaluate the expression levels of these three NOS1AP isoforms in human brain tissue, postmortem samples from Brodmann’s Area (BA) 46, BA11, Medial Temporal Lobe (MTL), Occipital Lobe (OL), and cerebellum of unaffected patients and those with schizophrenia were obtained from the Human Brain and Spinal Fluid Resource Center (Los Angeles, CA) and subjected to Western blotting with normalization to GAPDH or actin, as previously described (Xu, et al., 2005). Investigators were blinded to all subject information until after statistical analysis. The logarithms of the normalized values for subjects with schizophrenia and unaffected control patients within the same brain region were compared using the standard t-test. Correction for testing of multiple expression levels was made using permutations of case/control labels. Secondary examination of linear models with other covariates was based on the AICc model selection criterion (Burnham, 2002). The L (p = 0.0067; reported p-values are nominal), S´ (p = 0.0082) and S (p = 0.0041) isoforms were increased in BA46 of patients with schizophrenia (Figure 1B) at a nominal significance level, although only the increase in the S isoform was significant (p