Dysregulation of DLX5 by Position Effect in a Family with a Novel Phenotype of Craniofacial Defects and Hearing Loss
Precisely regulated temporal and spatial patterns of gene expression are essential for proper human development. Cis-acting regulatory elements, some located at large distances from their corresponding genes, play a critical role in transcriptional control of key developmental genes and disruption of these regulatory elements can lead to disease. We report a three generation family with five affected members in whom an inv(7)(q21.3q35) segregates with craniofacial defects and mixed hearing loss. FISH mapping of both inv(7) breakpoints and further delineation of the breakpoint regions by Southern blotting was employed to facilitate cloning and sequencing of both breakpoints. The 7q21.3 breakpoint lies about 80 kb centromeric to DLX5. While the breakpoint does not disrupt DLX5, the abnormal phenotype observed in the family is similar to defects seen in DLX5 knockout mice, suggesting that the family’s phenotype could be due to altered DLX5 regulation. DLX5 is paternally imprinted in human lymphoblastoid cell lines and other tissues. In four of five family members the inv(7) is known to occur on the maternal allele, therefore, dysregulation of DLX5 on this allele could lead to complete absence of DLX5 expression in some tissues. Results of reverse transcriptase-PCR performed on RNA isolated from lymphoblastoid cell lines derived from three family members indicate that DLX5 is not expressed, supporting altered DLX5 expression as the cause of the family’s disorder. To find the regulatory element responsible for the aberrant expression pattern, we used bioinformatic approaches to analyze 5 kb of sequence deleted at and additional sequences centromeric to the 7q21.3 breakpoint for the presence of conserved non-genic regions and transcription factor binding sites. Several sequences were identified in these regions that may act as DLX5 enhancers. We are currently generating transgenic mice to test these sequences for in vivo enhancer activity by determining if they can drive expression of a reporter gene in relevant tissues of mouse embryos. Through these experiments, we aim to discover the etiology of this disorder and to gain valuable insight into the complex regulation of DLX5, a critical developmental gene.