25 May 2012. A single gene within a chromosomal region associated with autism, schizophrenia, and other brain disorders controls head size in zebrafish, according to a study published in Nature on 17 May. Led by Nicholas Katsanis at Duke University, Durham, North Carolina, the study fingers KCTD13, a relatively unknown gene lying within the 16p11.2 region implicated in head size in humans: duplications and deletions of this region are associated with microcephaly and macrocephaly, respectively. Whether autism or schizophrenia are also linked to KCTD13 is unclear, but the findings offer up a new genetic controller of brain development, which is thought to go awry in both disorders.
The study uses zebrafish to efficiently identify the dosage-sensitive genes among the 29 genes contained in the 16p11.2 locus. This region has been associated with a variety of phenotypes by copy number variants (CNVs): deletions of the 16p11.2 locus are associated with autism, obesity, and macrocephaly, whereas duplications are associated with both autism and schizophrenia, anorexia, and microcephaly. The “mirror phenotype” of head size gave the researchers a chance to explore the effects of each gene in the region on the anatomy of the experimentally amenable zebrafish, with only KCTD13 falling out as a controller of head size (see SRF conference report). Along with a recent flurry of imaging genetics studies of brain size in humans (see SRF related news story), the findings on head size might prove to be a useful intermediate phenotype through which more complex disorders may be understood (Meyer-Lindenberg and Weinberger, 2006).
First author Christelle Golzio and colleagues began by injecting human transcripts of the different 16p11.2 genes into zebrafish embryos. Zebrafish have versions of all but five of these genes, making most of these injections overexpression experiments. Four days after injections, overexpression of only one—KCTD13—affected head size, giving a significantly smaller head than controls. The researchers then knocked down expression of the zebrafish version of KCTD13, and found a corresponding increase in head size compared to controls. This kind of sliding scale was not seen for body length, which argues that KCTD13 was not simply delaying or accelerating body development.
KCTD13 encodes a protein that interacts with proliferating cell nuclear antigen (PCNA), a protein that helps with DNA replication. This suggests that KCTD13 may be involved with neurogenesis, and consistent with this, the researchers found that KCTD13 was strongly expressed in the developing brain. When the researchers stained zebrafish embryos with markers for cell birth and death, they found that the microcephaly induced by KCTD13 overexpression was accompanied by decreased neural proliferation and increased cell death relative to controls, whereas the macrocephaly induced by KCTD13 downregulation came with only increases in proliferation. These shifts in proliferation and death were apparent even before the head size differences developed, which argues that an altered balance in cell birth and/or death drove the microcephaly and macrocephaly.
Moving into mammals
In mice, KCTD13 is expressed during development in cortex, striatum, hippocampus, and the olfactory tubercle. To test whether downregulation of KCTD13 in mice could recapitulate the increased head size found in zebrafish, the researchers designed short hairpin RNAs (shRNAs) against the mouse version of KCTD13 and injected them into the ventricular space of developing brains in mouse embryos. Two days later, they found a twofold increase in newborn cells compared to controls, consistent with a similar relationship between KCTD13 expression and head size; an overexpression experiment was not reported.
As for humans, the researchers noted a report last year of a smaller than usual deletion within the 16p11.2 locus (118 kb versus the usual 600 kb) that included only five genes, including KCTD13; three members of the family carrying this deletion also had autism (Crepel et al., 2011). In the new study, Katsanis’ team screened this restricted region in 518 people with autism and 8328 controls. This turned up a KCTD13-specific deletion in an exon in one case of autism. The person also harbored another CNV nearby, however, obscuring a potential link between KCTD13 and autism.
The KCTD13 findings don’t preclude a role for other genes in head size—indeed, the researchers reported that combinations of KCTD13 and other 16p11.2 gene transcripts (either MAPK3 or MVP) enhanced microcephaly in zebrafish. Still, the study illustrates how this approach can offer a fast track to function for the many, many genes implicated by CNVs in psychiatric disorders. People carrying these CNVs often come with physical anomalies, which can seem secondary to their psychiatric diagnoses. But these anatomical features might be key clues to understanding the fine grain of a CNV, and potentially, more complex disorders like schizophrenia.—Michele Solis.
Golzio C, Willer J, Talkowski ME, Oh EC, Taniguchi Y, Jacquemont S, Reymond A, Sun M, Sawa A, Gusella JF, Kamiya A, Beckmann JS, Katsanis N. KCTD13 is a major driver of mirrored neuroanatomical phenotypes of the 16p11.2 copy number variant. Nature. 2012 May 16;485(7398):363-7. Abstract