Evidence for Common Ancestral Origin of a Recurring BRCA1 Genomic Rearrangement Identified in High-Risk Hispanic Families

Diagram of the 3-primer BRCA1 del (ex 9-12) assay. A, genomic structure of breakpoint region with numbered exons. Solid blocks, BRCA1 exons 8 to 13; close crosshatch bar, intron 8; distinct crosshatch patterns, introns involved in rearrangement; open bars, introns 9 to 11. Bracket below the diagram, positions of the respective breakpoints. Primer positions for the specific rearrangement assay (P₁, P₂, P₃) are indicated along with respective orientation. Although not detectable in assay, predicted product for P₁ and P₃ (15.4 kb) wild-type allele is indicated by parentheses above diagram. B, depiction of the wild-type (P₁ and P₂) and rearrangement mutation-specific (P₁ and P₃) PCR products. C, photograph of ethidium bromide–stained agarose gel with cases (first five lanes) and controls (wt, water). Size standard indicated on right. Clean bands of 1,145 and 742 bp represent the wild-type and mutant alleles in positive cases (lanes 1-5), while the wild-type control (lane 6) shows only the wild-type allele.

BRCA1 deletion exon 9 to 12 breakpoint localized within AluSp sequence in intron 8 and AluSx sequence in intron 12. The sequence of the 2.7-kb product spanning the breakpoint was determined and mapped against BRCA1 genomic sequence [National Center forBiotechnology Information (NCBI) accession no. L78833] by RepeatMasker v3.1.X software (A.F.A. Smit, R. Hubley and P. Green RepeatMasker at http://repeatmasker.org). The breakpoint is depicted as a consensus block of identity; thus, the precise point of recombination and which respective Alu element contributed to that cannot be determined. The intronic breakpoint region is depicted, with unique intron 8 sequence leading to the specific AluSp (single underline) element, adjacent to the consensus block (boxed), and the 3′ AluSx element (double underline) in intron 12 sequence.

BRCA1 protein functional domains and predicted frameshift and premature truncation. A, adapted from Welcsh et al., 2000 (42): Translated BRCA1 exons and distribution of known/putative functional domains for BRCA1 protein. B, depiction of truncated coding sequence resulting from deletion of exons 9 to 12. C, predicted frameshift and premature stop codon that would result from direct splicing of the remaining exons (8 joined to 13), presuming no use of cryptic or alternate splice sites.

BRCA1 del (ex 9-12) haplotype. The haplotype was determined from the mother (2025-14) and daughter (2025-1) pair (in bold type). No determination was made for marker D17S1327 as the genotype is equivalent in both mother and daughter. Dash, deleted marker D17S1323 in the mutant allele. For the other four cases, a genotype was established. All the individuals carrying the mutation carried the alleles associated with the haplotype, suggesting that this is a founder mutation.