Gene expression profiling in facioscapulohumeral muscular dystrophy

Abstract

Facioscapulohumeral muscular dystrophy (FSHD) is the third most common form of inherited muscle disease following Duchenne and myotonic dystrophy. FSHD is initially characterized by an asymmetric, progressive weakness of the facial and pectoral girdle muscles. The disorder is inherited in an autosomal dominant fashion with nearly complete penetrance. FSHD results from a deletion of integral copies of a 3.3-kb tandem repeat unit on the long arm of chromosome 4 (D4Z4). A "short" EcoR1 fragment (< 35 kb) containing D4Z4 segregates with the disease, whereas the size of this polymorphic locus in the normal population ranges between 35 and 300 kb. The immediate proximity of the 3.3-kb tandem repeat to the telomere and its sequence similarity to constituitive heterochromatin suggest that this repeat, deleted in FSHD, lies in telomeric heterochromatin. This conclusion is further supported by the fact that, despite intense efforts over the past seven years, there have been no protein coding transcripts identified fr om this repeat sequence. Integral deletions of the heterochromatic D4Z4 repeat instead appear to disrupt the normal expression of adjacent genes, a phenomenon akin to position effect variegation in Drosophila and telomere silencing in yeast. The genes affected by the D4Z4 polymorphism have not been identified, as the 4qter region is rich in repetitive and pseudogene sequences. We are currently using the Affymetrix GeneChip system to examine large-scale differential gene expression in FSHD muscle tissue and myoblast cell lines. In addition, we have used the GeneChip system to examine the regulatory mechanism disrupted by deletions of the heterochromatic repeat D4Z4. Differential gene expression in yeast strains carrying telomeric half-YACs encompassing the FSHD region have been analysed in an effort to identify yeast homologues of genes involved in heterochromatin formation at the D4Z4 locus. A number of genes involved in DNA interstrand crosslink repair were found to be upregulated, perh aps unveiling the initial mutational mechanism in this disease. Thus, we will likely yield immense insight into both the regulatory and mutational mechanisms of the human disease FSHD as well as elucidate the primary cause of this disorder through microarray analysis.Exposure of cells to anticancer agents can result in altered gene expression. In general, only a single gene or a small number of genes are studied. To improve the efficiency and value of such experiments, we are interested in using DNA array technology to identify, on a genome-wide scale, drug-responsive genes which might serve both as surrogate markers for drug action and as future targets for drug development. As a means to establish this approach in our Centre we have carried out two exploratory projects. In the first project, we examined gene expression in p53 wild-type and mutant ovarian cell lines established from two patients with ovarian cancer who achieved a complete remission following treatment. Neither line can grow in vitro and were passaged as subcu...