Abstract
We previously reported a schizophrenia-associated polymorphic CT di-nucleotide repeat (DNR) at the 5′-untranslated repeat (UTR) of DPYSL2, which responds to mammalian target of Rapamycin (mTOR) signaling with allelic differences in reporter assays. Now using microarray analysis, we show that the DNR alleles interact differentially with specific proteins, including the mTOR-related protein HuD/ELAVL4. We confirm the differential binding to HuD and other known mTOR effectors by electrophoretic mobility shift assays. We edit HEK293 cells by CRISPR/Cas9 to carry the schizophrenia risk variant (13DNR) and observe a significant reduction of the corresponding CRMP2 isoform. These edited cells confirm the response to mTOR inhibitors and show a twofold shortening of the cellular projections. Transcriptome analysis of these modified cells by RNA-seq shows changes in 12.7% of expressed transcripts at a false discovery rate of 0.05. These transcripts are enriched in immunity-related genes, overlap significantly with those modified by the schizophrenia-associated gene, ZNF804A, and have a reverse expression signature from that seen with antipsychotic drugs. Our results support the functional importance of the DPYSL2 DNR and a role for mTOR signaling in schizophrenia.
Highlights
DPYSL2 encodes a cytosolic protein best known as collapsin response mediator protein 2 (CRMP2), a member of a five-gene family named for their involvement in axonal growth cone collapse.Mammals express three DPYSL2 transcripts, A, B and C, each differing in an alternative first exon
The results suggest that the 13DNR allele has a weaker response to mammalian target of Rapamycin (mTOR) signaling, and is more sensitive to mTOR inhibition
We have demonstrated that the di-nucleotide repeat (DNR) variant in the 5ʹ-untranslated repeat (UTR) of the DPYSL2B transcript has strong effects on the expression of the corresponding protein isoform, and provided additional evidence that this is likely due to changes in its response to mTOR signaling
Summary
DPYSL2 encodes a cytosolic protein best known as collapsin response mediator protein 2 (CRMP2), a member of a five-gene family named for their involvement in axonal growth cone collapse.Mammals express three DPYSL2 transcripts, A, B and C, each differing in an alternative first exon. DPYSL2 encodes a cytosolic protein best known as collapsin response mediator protein 2 (CRMP2), a member of a five-gene family named for their involvement in axonal growth cone collapse. Most published literature, including this report, has focused on the B transcript, which we refer to as DPYSL2B with the corresponding protein isoform CRMP2B. Altering the expression of Crmp[2] in cultured rat hippocampal neurons significantly affects axonal growth.[15,16] Further, multiple proteomic studies report aberrant expression of CRMP2 in the brains of schizophrenics.[17,18,19,20] Most recently, a study showed downregulation of the Crmp[2] protein in the prefrontal cortex and hippocampus of prenatally stressed rats.[21]
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