Individuals with 22q11.2 microdeletions show behavioral and cognitive deficits and are at high risk of developing schizophrenia. We analyzed an engineered mouse strain carrying a chromosomal deficiency spanning a segment syntenic to the human 22q11.2 locus. We uncovered a previously unknown alteration in the biogenesis of microRNAs (miRNAs) and identified a subset of brain miRNAs affected by the microdeletion. We provide evidence that the abnormal miRNA biogenesis emerges because of haploinsufficiency of the Dgcr8 gene, which encodes an RNA-binding moiety of the 'microprocessor' complex and contributes to the behavioral and neuronal deficits associated with the 22q11.2 microdeletion.

Using a relatively dense genetic map of 72 single-nucleotide polymorphisms (SNPs) distributed across the entire 1.5-Mb locus on chromosome 22q11 associated with susceptibilit to schizophrenia 1,2 , we previously identified two subregions that were consistently associated with the disease 3,4 . In the distal subregion, we detected an association signal with five neighboring SNPs distributed over a haplotypic block of 80 kb encompassing six known genes 4 . One of these five SNPs, rs175174, had the strongest association of all 72 SNPs that we tested. Here we show that rs175174 regulates the level of the fully functional transcript by modulating the retention of intron 4 of the gene ZDHHC8, which encodes a putative transmembrane palmitoyltransferase. Zdhhc8-knockout mice had a sexually dimorphic deficit in prepulse inhibition, a gene dosage-dependent decrease in exploratory activity in a new environment and a decreased sensitivity to the locomotor stimulatory effects of the psychomimetic drug dizocilpine (MK801). SNP rs175174 shows differences in transmission distortion between sexes in individuals with schizophrenia. Our results indicate that there is an unexpected connection between impaired palmitate modification of neuronal proteins and the psychiatric phenotypes associated with microdeletions of chromosome 22q11.SNP rs175174 is located in intron 4 of ZDHHC8 (previously annotated as KIAA1292; Fig. 1a,b), and the potential effect of rs175174 genotype on splicing warranted further investigation. We prepared total or poly(A) + RNA from fresh (nontransformed) human lymphocytes and subjected it to reverse transcription, PCR amplification using primers located in exons 1 and 8 and sequencing. This analysis identified a reproducible, complex pattern of alternative splicing around intron 4 (similar analysis of exons 8-11 showed no evidence for alternative splicing; data not shown). We identified two main products of comparable abundance in both the total (Fig. 1c) and the poly(A) + RNA fractions (data not shown), derived either from the fully spliced form or from an unspliced form retaining intron 4 (Fig. 1c). Sequence analysis of the introns and their flanking regions identified a suboptimal 5′ splice site (Fig. 1b) that could reduce the efficiency of splicing, leading to the retention of intron 4 (refs. 5,6). Examples of intron retention have been found in transcripts from several species 7 . SNP rs175174 is located in the middle of intron 4, embedded in a highly conserved heptanucleotide motif (Fig. 1d). The retained intron introduces a stop codon in the open reading frame that could lead to premature termination of translation (Fig. 1d).We constructed minigenes from the 'risk' and 'nonrisk' alleles containing exon 4, intron 4 and exon 5 along with flanking genomic sequences and transfected them into human embryonic kidney (HEK293) cells. Initial semiquantitative PCR amplification analysis showed that the pattern of splicing observed in lymphocytes was recapitulated in total RNA prepared from HEK293 cells transiently...

Microdeletions of 22q11.2 represent one of the highest known genetic risk factors for schizophrenia. It is likely that more than one gene contributes to the marked risk associated with this locus. Two of the candidate risk genes encode the enzymes proline dehydrogenase (PRODH) and catechol-O-methyltransferase (COMT), which modulate the levels of a putative neuromodulator (L-proline) and the neurotransmitter dopamine, respectively. Mice that model the state of PRODH deficiency observed in humans with schizophrenia show increased neurotransmitter release at glutamatergic synapses as well as deficits in associative learning and response to psychomimetic drugs. Transcriptional profiling and pharmacological manipulations identified a transcriptional and behavioral interaction between the Prodh and Comt genes that is likely to represent a homeostatic response to enhanced dopaminergic signaling in the frontal cortex. This interaction modulates a number of schizophrenia-related phenotypes, providing a framework for understanding the high disease risk associated with this locus, the expression of the phenotype, or both.