Functional characterization of the human Huntington's disease gene promoter

The genetic basis of neurodegeneration in Huntington's disease (HD) has bee n identified as a (CAG)(> 37) repeat expansion in a gene of unknown functio n. Interestingly, patients with the same expanded (CAG), repeat length may have markedly different ages at onset. Based on experiences in animal model s the level of expression might be one of the modifying factors. To gain in sight into the regulation of the human HD gene we functionally analyzed 226 6 bp of the HD gene promoter region. This region lacks a TATA and a CAAT bo x, is GCrich, and it has several consensus sequences for SP1, AP-2 and AP-4 binding sites. The stretch between nucleotides -49 and -198 relative to th e first ATG is highly conserved between human and rodents and it harbors se veral potential binding sites for transcription factors. We analyzed deleti on mutants fused with the chloramphenicol acetyltransferase (CAT) reporter gene in transfected, huntingtin expressing neuronal (NS20Y) and non-neurona l (CHO) cell lines. Partial deletion of the evolutionarily conserved part o f the promoter significantly reduces the activity in both neuronal and non- neuronal cells indicating that the core promoter activity is located betwee n nucleotides -221 and 4, relative to the +1 translation start site. Bindin g affinities of DNA-protein interactions were defined by electrophoretic mo bility shift assays and the protected nucleotide positions were determined by DNase I footprinting. (C) 2001 Elsevier Science BY All rights reserved.