A comprehensive analysis of 22q11 gene expression in the developing and adult brain

Abstract

Deletions at 22q11.2 are linked to DiGeorge or velocardiofacial syndrome (VCFS), whose hallmarks include heart, limb, and craniofacial anomalies, as well as learning disabilities and increased incidence of schizophrenia. To assess the potential contribution of 22q11 genes to cognitive and psychiatric phenotypes, we determined the CNS expression of 32 mouse orthologs of 22q11 genes, primarily in the 1.5-Mb minimal critical region consistently deleted in VCFS. None are uniquely expressed in the developing or adult mouse brain. Instead, 27 are localized in the embryonic forebrain as well as aortic arches, branchial arches, and limb buds. Each continues to be expressed at apparently constant levels in the fetal, postnatal, and adult brain, except for Tbx1, ProDH2, and T10, which increase in adolescence and decline in maturity. At least six 22q11 proteins are seen primarily in subsets of neurons, including some in forebrain regions thought to be altered in schizophrenia. Thus, 22q11 deletion may disrupt expression of multiple genes during development and maturation of neurons and circuits compromised by cognitive and psychiatric disorders associated with VCFS.

Fig. 1.

(a) The top shows a scale map of the proximal 1.5-Mb portion of chromosome 22q11 consistently deleted in DiGeorge syndrome (containing 27 of 32 transcripts examined) showing transcription units (orange bars) and low-copy repeats (LCRs, white boxes). The bottom shows that a majority of mouse orthologs are in a segment of mouse chromosome 16. (b) PCR array of 22q11.2 orthologs at sites of nonaxial M/E interaction: the frontonasal mass/forebrain (FNM/FB), branchial arches (BA), aortic arches and heart (AAH), and forelimb buds (FLB). Expression is presented in chromosomal order, centromeric (top) to telomeric (bottom). Brackets and arrows represent genes that share bidirectional promoters. The LCR bracket indicates genes found in human LCRs (not on chromosome 16 in mouse). β-actin and GAPDH are positive controls. (c–h). Whole embryonic day (E) 10/E10.5 mouse embryo immunolocalization of five 22q11 proteins shows correspondence between message and protein localization for this subset of orthologs. (i–n) Whole E10/E10.5 mouse embryo in situ hybridization of seven 22q11 orthologs shows focal expression at nonaxial inductive sites.

Fig. 2.

(a) PCR array showing expression of 22q11 orthologs in whole E10 and E14 mouse embryos and in the brain (anterior from the medulla) from E12 through adulthood. A no-RT lane is included as a negative control for sample contamination, and β-actin and GAPDH were used as positive controls. (b–d) Immunolocalization (green) of three 22q11 proteins in the developing neocortex at E16 shows expression in distinct domains of the developing cortex (cytoarchitecture shown with blue nuclear counterstain). vz, ventricular zone; iz, intermediate zone; cp, cortical plate. (e–h) Immunolocalization (green) of four 22q11 proteins in the adult neocortex suggests subtle variations in laminar frequency of labeled cells. Layer identity, demonstrated by blue nuclear counterstain, is shown at left.

Fig. 3.

(a) PCR array of microdissected adult forebrain regions and other CNS domains shows expression of the 22q11 orthologs. (b) A PCR array of several organs shows widespread expression of most 22q11 orthologs outside the CNS.

Fig. 4.

Immunolocalization (green) of six 22q11 proteins in the adult mouse brain shows expression in subsets of neurons in the striatum, hippocampus, and cerebellum; a blue nuclear counterstain indicates the position of cell bodies. Double labeling with glial (second row, red) or neuronal (third row, red) markers suggests that the cells expressing these 22q11 proteins are neurons.

Fig. 5.

PCR expression array of selected human 22q11 genes. Human cDNA pools were obtained from BD Biosciences Clontech. cDNAs from whole brain and organ tissues were evaluated, and samples of forebrain and other CNS regions are included at the far right.