Unravelling the molecular phenotype of adrenocortical dysplasia (acd) mice

The acd mouse is a model for congenital adrenal hypoplasia, displaying small adrenal glands and a lack of cortical zonation. The gene mutated in acd mice (Acd) is an orthologue of a human gene encoding for a member of the telomere cap complex (TCC), which regulates telomerase access to the telomere and prevents uncontrolled processing by the DNA-repair machinery. The purpose of this study is to further describe the molecular phenotype of the acd mouse. Failure in telomere protection results in either apoptosis or senescence. In isolated mouse embryonic fibroblasts (MEFs) from acd mice we observe no differences in cumulative population doublings or induction of senescence (β-Gal pH 6.0). To further show an association of Acd with the TCC we generated anti-Acd antibodies. In a modified chromatin immunoprecipitation assay crosslinked protein-DNA complexes were immunoprecipitated, dot-blotted and hybridized with telomeric repeat sequences. The signal ratio of anti-Acd/preimmune serum precipitation was higher in wild type compared to acd mice giving evidence for an in vivo association of Acd with the TCC. Telomeric dysfunction usually results in chromosomal aberrations and subsequent genomic instability. In metaphase spreads from acd MEFs we found a higher number of these chromosomal abnormalities in DAPI stains. Currently we are characterizing this phenotype by SKY analysis. In view of the fact that most animal models deficient in proteins of the TCC are lethal we investigated the possibility of acd being a hypomorphic allele. In quantitative RT-PCR analysis we show a minimal amount of wild type Acd mRNA in acd MEFs but not in any tissue from adult mutant animals. In summary we show an association of Acd with telomeric repeats and an accumulation of chromosomal aberrations in acd MEFs. To further investigate the role of Acd and telomeric integrity in adrenocortical development we are crossing acd mice and p53 deficient mice to determine if p53 mediated growth arrest due to telomeric failure contributes to the observed adrenocortical phenotype in acd mice.