Abstract
In humans, SHH haploinsufficiency results in holoprosencephaly (HPE), a defect in anterior midline formation. Despite the importance of maintaining SHH transcript levels above a critical threshold, we know little about the upstream regulators of SHH expression in the forebrain. Here we describe a rare nucleotide variant located 460 kb upstream of SHH in an individual with HPE that resulted in the loss of Shh brain enhancer-2 (SBE2) activity in the hypothalamus of transgenic mouse embryos. Using a DNA affinity-capture assay, we screened the SBE2 sequence for DNA-binding proteins and identified members of the Six3 and Six6 homeodomain family as candidate regulators of Shh transcription. Six3 showed reduced binding affinity for the mutant compared to the wild-type SBE2 sequence. Moreover, Six3 with HPE-causing alterations failed to bind and activate SBE2. These data suggest a direct link between Six3 and Shh regulation during normal forebrain development and in the pathogenesis of HPE.
| Original language | English |
|---|---|
| Pages (from-to) | 1348-1353 |
| Number of pages | 6 |
| Journal | Nature Genetics |
| Volume | 40 |
| Issue number | 11 |
| DOIs | |
| Publication status | Published - Nov 2008 |
Bibliographical note
Funding Information:We thank the families for their participation in these studies. We also thank J. Richa and his staff at the University of Pennsylvania Transgenic and Mouse Chimeric Facility for their assistance in transgenic mouse production. We are grateful to V. Cheung, D. Kessler and T. Kadesch for their helpful comments on the manuscript. We are also grateful to K. Ewens and W. Ankener (R. Spielman laboratory) for the control human genotyping data and P. Bovolenta (Instituto Cajal, CSIC, Madrid, Spain) for kindly providing the human Six3 and Six6 expression constructs. This work was supported by NIH grants R01 NS39421 from NINDS (D.J.E.), R01 NS052386 (G.O.), March of Dimes grant #1-FY05-112 (D.J.E.), a Pew Scholar Award in the Biomedical Sciences (D.J.E.), Cancer Center Support CA-21765 (G.O.), the American Lebanese Syrian Associated Charities (ALSAC) (G.O.) and the Division of Intramural Research, National Human Genome Research Institute, National Institutes of Health (M.M.).