High frequency of large intragenic deletions in the Fanconi anemia group Agene

Fanconi anemia (FA) is an autosomal recessive disorder exhibiting chromosom al fragility, bone-marrow failure, congenital abnormalities, and cancer. At least eight complementation groups have been described, with group A accou nting for 60%-65% of FA patients. Mutation screening of the group A gene (F ANCA) is complicated by its highly interrupted genomic structure and hetero geneous mutation spectrum. Recent reports of several large deletions of FAN CA, coupled with modest mutation-detection rates, led us to investigate whe ther many deletions might occur in the heterozygous state and thus fail to be detected by current screening protocols. We used a two-step screening st rategy, in which small mutations were detected by fluorescent chemical clea vage of the FANCA transcript, and heterozygosity for gross deletions was de tected by quantitative fluorescent multiplex PCR. We screened 26 cell lines from FA complementation group A for FANCA mutations and detected 33 differ ent mutations, 23 of which were novel. Mutations were observed in all 26 ce ll lines and included 43 of a possible 52 mutant alleles (83%). Of the muta nt alleles, 40% were large intragenic deletions that removed up to 31 exons from the gene, indicating that this may be the most prevalent form of muta tion in FANCA. Several common deletion breakpoints were observed, and there was a highly significant correlation between the number of breakpoints det ected in a given intron and the number of Alu repeats that it contained, wh ich suggests that Alu-mediated recombination may explain the high prevalenc e of deletions in FANCA. The dual screening strategy that we describe may b e useful for mutation screening in other genetic disorders in which mutatio n-detection rates are unexpectedly low.