Fine mapping of a gene responsible for regulating dietary cholesterol absorption; founder effects underlie cases of phytosterolaemia in multiple communities

Abstract

Sitosterolaemia (also known as phytosterolaemia, MIM 210250) is a rare recessive autosomal inherited disorder, characterised by the presence of tendon and tuberous xanthomas, accelerated atherosclerosis and premature coronary artery disease. The defective gene is hypothesised to play an important role in regulating dietary sterol absorption and biliary secretion, thus defining a molecular mechanism whereby this physiological process is carried out. The disease locus was localised previously to chromosome 2p21, in a 15 cM interval between microsatellite markers D2S1788 and D2S1352 (based upon 10 families, maximum lodscore 4.49). In this study, we have extended these studies to include 30 families assembled from around the world. A maximum multipoint lodscore of 11.49 was obtained for marker D2S2998. Homozygosity and haplotype sharing was identified in probands from non-consanguineous marriages from a number of families, strongly supporting the existence of a founder effect among various populations. Additionally, based upon both genealogies, as well as genotyping, two Amish/Mennonite families, that were previously thought not to be related, appear to indicate a founder effect in this population as well. Using both homozygosity mapping, as well as informative recombination events, the sitosterolaemia gene is located at a region defined by markers D2S2294 and Afm210xe9, a distance of less than 2 cM.

Figure 1

Sitosterolaemia pedigrees. The pedigrees analysed in this study are shown above. All parents are shown as obligate carriers. The one known consanguineous marriage, pedigree 3400, is as indicated. Although family members for pedigree 3800 are known, they were not available for analyses.

Figure 2

Multipoint analyses of markers genotyped in the sitosterolaemia pedigrees. Multipoint analysis was performed using SIMWALK, using markers indicated in Table 1, as described in Methods. Although pedigree 4000 showed a recombination event for markers Afm210xe9 and D2S4016 (see Results and Figure 3), since these markers have a recombination distance of 0.0, this event is not detected by SIMWALK (see text).

Figure 3

Recombination events that narrow the localisation of the sitosterolaemia gene. The lower limit of the sitosterolemia gene is defined by a recombination event detected in family 200, and the upper limit is defined by a recombination in pedigree 700. The parents in pedigree 700 are not available for genotype analyses. Additionally, two contiguous markers, Afm210xe9 and D2S4016, were recombinant in pedigree 4000, placing the sitosterolaemia locus either above or below these markers.

Figure 4

A summary of informative recombination events and homozygosity detected in all of the pedigrees. A indicates all the informative recombinant events detected, B shows probands that showed contiguous homozygosity for more than three markers, and where parental alleles were fully informative, either by direct analyses, or based upon predicted genotypes. Although the recombination events localise the sitosterolaemia gene to one of two regions, when combined with the homozygosity patterns, the gene can be localised to lie between D2S2294 and Afm210xe9 (boxed area, see text). Note that only pedigree 3400 is known to be consanguineous.

Figure 5

Haplotype sharing in some of the pedigrees. Comparison of haplotypes within the narrowed area of interest showed that the probands from the Amish and Mennonite families (left hand side), showed significant haplotype sharing as did two Japanese probands (middle panel). Additionally, a Norwegian and a Finnish proband (right hand side) also showed both homozygosity and haplotype sharing.

Figure 6

Genealogical analyses of the Amish and Mennonite families in this study. A genealogical analysis of the parents from pedigrees 2700 and 2200 (as indicated) was performed as described in Methods. Five possible founders, that link all four parents, were identified. An approximate time index is as shown in the left margin for temporal orientation. As can be seen, individual 74 is the least related to all of the remaining three obligate carriers. Of the possible founders, the Yoder line has the most frequent interconnections, and the Blank lines the least. The small dots indicate a generation. The ‘Amish’ mutation is at least 200 years old.