Abstract
BackgroundPolymorphisms at the G72/G30 locus on chromosome 13q have been associated with schizophrenia or bipolar disorder in more than ten independent studies. Even though the genetic findings are very robust, the physiological role of the predicted G72 protein has thus far not been resolved. Initial reports suggested G72 as an activator of D-amino acid oxidase (DAO), supporting the glutamate dysfunction hypothesis of schizophrenia. However, these findings have subsequently not been reproduced and reports of endogenous human G72 mRNA and protein expression are extremely limited. In order to better understand the function of this putative schizophrenia susceptibility gene, we attempted to demonstrate G72 mRNA and protein expression in relevant human brain regions.MethodsThe expression of G72 mRNA was studied by northern blotting and semi-quantitative SYBR-Green and Taqman RT-PCR. Protein expression in human tissue lysates was investigated by western blotting using two custom-made specific anti-G72 peptide antibodies. An in-depth in silico analysis of the G72/G30 locus was performed in order to try and identify motifs or regulatory elements that provide insight to G72 mRNA expression and transcript stability.ResultsDespite using highly sensitive techniques, we failed to identify significant levels of G72 mRNA in a variety of human tissues (e.g. adult brain, amygdala, caudate nucleus, fetal brain, spinal cord and testis) human cell lines or schizophrenia/control post mortem BA10 samples. Furthermore, using western blotting in combination with sensitive detection methods, we were also unable to detect G72 protein in a number of human brain regions (including cerebellum and amygdala), spinal cord or testis. A detailed in silico analysis provides several lines of evidence that support the apparent low or absent expression of G72.ConclusionOur results suggest that native G72 protein is not normally present in the tissues that we analysed in this study. We also conclude that the lack of demonstrable G72 expression in relevant brain regions does not support a role for G72 in modulation of DAO activity and the pathology of schizophrenia via a DAO-mediated mechanism. In silico analysis suggests that G72 is not robustly expressed and that the transcript is potentially labile. Further studies are required to understand the significance of the G72/30 locus to schizophrenia.
Highlights
Polymorphisms at the G72/G30 locus on chromosome 13q have been associated with schizophrenia or bipolar disorder in more than ten independent studies
We sought to investigate expression of G72 mRNA across a variety of human CNS regions, using Clontech multiple tissue Northern (MTN) (Multiple Tissue Northern) human brain blots
While further studies are required to understand the significance of the G72/30 locus to schizophrenia, we propose that if native G72 protein exists at all, it is expressed at such low levels that any physiological role is called in to question
Summary
Polymorphisms at the G72/G30 locus on chromosome 13q have been associated with schizophrenia or bipolar disorder in more than ten independent studies. Initial reports suggested G72 as an activator of D-amino acid oxidase (DAO), supporting the glutamate dysfunction hypothesis of schizophrenia. These findings have subsequently not been reproduced and reports of endogenous human G72 mRNA and protein expression are extremely limited. G72 and G30 are overlapping genes transcribed from opposite strands on chromosome 13q33 They were initially identified within a 65 kb region containing markers associated with schizophrenia in two independent disease cohorts [1]. Genetic association between the G72/G30 locus and both schizophrenia and bipolar disorder has subsequently been reported in several studies and is supported by a recent meta-analysis [2]. As NMDA receptor hypofunction has been implicated in the pathophysiology of schizophrenia, the report of a functional interaction between G72 and DAO suggests a pathway whereby G72 could modulate DAO activity, D-serine levels and NMDA receptor activity and, contribute to the disease pathology
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