Holoprosencephaly (HPE) is a brain malformation characterized by the failure of the forebrain to divide into hemispheres during development. While it is estimated that HPE affects 1 in 5,000-10,000 live births, the real incidence may by as high as 1 in 200-250 fetuses, owing to the high rates of spontaneous abortion of those severely affected. Although not the sole culprit, mutations in the Six3 gene can cause HPE.
Dr. Oliver and colleagues are helping to understand why.
"This work adds another player and another step towards deciphering the molecules and mechanisms involved in vertebrate head development in normal and pathological conditions, " explains Dr. Oliver.
The researchers found that in mice, the protein encoded by the Six3 gene (SIX3) secures the identity of anterior (forebrain) neural cells by blocking out a posteriorizing signal that sweeps through the putative forebrain during day 8.0-8.5 of embryogenesis, and determines the identity of more posterior brain structures. This posteriorizing signal is transmitted through the Wnt family of secreted, cell-to-cell signaling proteins.
To examine the role of SIX3 in vivo, Dr. Oliver and colleagues generated transgenic mice deficient in Six3. These Six3-mutant mice died at birth, lacking the rostral forebrain and most anterior head structures, like the eyes and nose. Dr. Oliver and colleagues determined that the SIX3 protein binds to regulatory elements within the Wnt1 gene, suppressing its expression in the putative forebrain region, and thereby enabling these cells to adopt an anterior cell fate. In Six3-mutant mice these cells are not given the proper choice of anterior fate, and thus the forebrain fails to form normally.
Interestingly, the researchers also demonstrated that this genetic pathway of forebrain regionalization has been conserved throughout evolution from fish to mammals. In mutant zebrafish in whom Wnt1 is aberrantly over-expressed, forebrain development fails, and these fish are termed "headless" mutants. In collaboration with Dr. Lila Solnica-Kretzel and researchers at Vanderbilt University, Dr. Oliver and colleagues showed that the forced expression of mouse Six3 messenger RNA in headless zebrafish mutants effectively down-regulated Wnt1 expression and rescued the headless phenotype.
"It is clear that small differences in the expression of Wnt proteins along the future brain region are critical for the determining the final identity of the different brain territories. We have now determined that in the most rostral region, Six3 expression is required in order to guarantee that no Wnt1 protein will reach that territory," states Dr. Oliver.
Journal
Genes & Development