Linkage and association studies identify a novel locus for Alzheimer disease at 7q36 in a Dutch population-based sample

Abstract

We obtained conclusive linkage of Alzheimer disease (AD) with a candidate region of 19.7 cM at 7q36 in an extended multiplex family, family 1270, ascertained in a population-based study of early-onset AD in the northern Netherlands. Single-nucleotide polymorphism and haplotype association analyses of a Dutch patient-control sample further supported the linkage at 7q36. In addition, we identified a shared haplotype at 7q36 between family 1270 and three of six multiplex AD-affected families from the same geographical region, which is indicative of a founder effect and defines a priority region of 9.3 cM. Mutation analysis of coding exons of 29 candidate genes identified one linked synonymous mutation, g.38030G-->C in exon 10, that affected codon 626 of the PAX transactivation domain interacting protein gene (PAXIP1). It remains to be determined whether PAXIP1 has a functional role in the expression of AD in family 1270 or whether another mutation at this locus explains the observed linkage and sharing. Together, our linkage data from the informative family 1270 and the association data in the population-based early-onset AD patient-control sample strongly support the identification of a novel AD locus at 7q36 and re-emphasize the genetic heterogeneity of AD.

Figure 1

Pedigree of family 1270. A blackened symbol represents a patient, and an unblackened symbol represents an unaffected individual or an at-risk individual with unknown phenotype. The Roman numbers to the left of the pedigree denote generations; Arabic numbers above the symbols denote individuals. All numbering is in accordance with that of van Duijn et al. (1994b). The Arabic numbers below the symbols denote age at onset for patients and either age at last examination or age at death for unaffected individuals and at-risk individuals with unknown phenotype. An asterisk (*) indicates an individual who was included in the linkage analysis. The arrow identifies the proband with early-onset AD (age 47 years).

Figure 2

Brain CT scan of patient III-21 at age 82 years, showing marked cortical and moderate subcortical atrophy, most pronounced in the temporal and frontal regions, with secondary dilatation of the lateral and third ventricles. Mainly frontally localized periventricular leukoencephalomalacia is present as well.

Figure 3

Genomewide linkage data of the 10-cM density scan of family 1270, with use of the ABI Linkage Mapping Set MD10, version 2, across all chromosomes. Linkage analysis at chromosomes 10, 12, and 21 was performed at a 5-cM density, with use of markers from the ABI Linkage Mapping Set HD-5, version 2. Two-point LOD scores for all markers were calculated using MLINK of the computer package Linkage, version 5.1, and are shown relative to their position on the genetic map (Dib et al. 1996).

Figure 3

Genomewide linkage data of the 10-cM density scan of family 1270, with use of the ABI Linkage Mapping Set MD10, version 2, across all chromosomes. Linkage analysis at chromosomes 10, 12, and 21 was performed at a 5-cM density, with use of markers from the ABI Linkage Mapping Set HD-5, version 2. Two-point LOD scores for all markers were calculated using MLINK of the computer package Linkage, version 5.1, and are shown relative to their position on the genetic map (Dib et al. 1996).

Figure 3

Genomewide linkage data of the 10-cM density scan of family 1270, with use of the ABI Linkage Mapping Set MD10, version 2, across all chromosomes. Linkage analysis at chromosomes 10, 12, and 21 was performed at a 5-cM density, with use of markers from the ABI Linkage Mapping Set HD-5, version 2. Two-point LOD scores for all markers were calculated using MLINK of the computer package Linkage, version 5.1, and are shown relative to their position on the genetic map (Dib et al. 1996).

Figure 4

Disguised linkage pedigree of family 1270 (fig. 1). Haplotypes are based on 17 informative markers at 7q36. Haplotypes for individuals from the first and second generations were inferred from genotype data of siblings and offspring. The risk haplotype was arbitrarily set for individual I-2. For confidentiality reasons, haplotypes are shown only for patients; the number of at-risk individuals included in the genotyping are indicated by numbers within diamonds.

Figure 5

Linkage pedigrees of Dutch families with early-onset AD who share haplotypes at 7q36. Blackened symbols represent patients with AD; unblackened symbols represent unaffected individuals or at-risk individuals with unknown phenotype. Roman numbers to the left of the pedigree denote generations; Arabic numbers above the symbols denote individuals. All numbers are in accordance with those used in the complete pedigrees published by van Duijn et al. (1994b). For patients, the age (in years) at onset is shown below the symbol. An asterisk (*) indicates that DNA was available for genetic analysis. The arrow identifies the proband. To protect privacy of at-risk individuals, the shared haplotypes are shown only for patients. Haplotypes for individual III-20 in family 1242 and individual III-4 in family 1034 were reconstructed from genotype data obtained from their siblings and offspring. For family 1125, only genotype data are given, since segregation data were not informative for deduction of haplotypes.

Figure 6

Expression analysis of PAXIP1. A, Northern blot of multiple tissues (FirstChoice Human Blot 1 [Ambion]) hybridized with pooled PAXIP1 cDNA fragments C1 (exons 1–9) and C2 (exons 8–22)—obtained by PCR amplification with use of primers C1F (5′-CGCCGCGGAGCCTCCCGGGCCGCCG-3′), C1R (5′-CAGGTGGCCAGCAGTTGCTTATCAGACATC-3′), C2F (5′-GAAGGCTTCTTATTGGGATG-3′), and C2R (5′-TGAAGGAAGCGCAGCAGCTCC-3′)—in a mixture of cDNA (brain, liver, testis, colon, and lymphoblast). B, β-Actin. Transcript lengths (kb) are specified at the left and were calculated on the basis of the Millennium Marker bands marked on the blot. C, RT-PCR of RNA extracted from seven different human brain regions and lymphoblasts of PAXIP1 C-terminal fragment C2 (exons 8–22). Predicted PCR fragment length of C2 is 1,487 bp.