Familial combined hyperlipidemia (FCHL), characterized by elevated levels of serum total cholesterol, triglycerides or both 1,2 , is observed in about 20% of individuals with premature coronary heart disease 1 . We previously identified a locus linked to FCHL on 1q21-q23 in Finnish families with the disease 3 . This region has also been linked to FCHL in families from other populations 4-6 as well as to type 2 diabetes mellitus 7-12 . These clinical entities have several overlapping phenotypic features, raising the possibility that the same gene may underlie the obtained linkage results. Here, we show that the human gene encoding thioredoxin interacting protein (TXNIP) on 1q, which underlies combined hyperlipidemia in mice 13 , is not associated with FCHL. We show that FCHL is linked and associated with the gene encoding upstream transcription factor 1 (USF1) in 60 extended families with FCHL, including 721 genotyped individuals (P = 0.00002), especially in males with high triglycerides (P = 0.0000009). Expression profiles in fat biopsy samples from individuals with FCHL seemed to differ depending on their carrier status for the associated USF1 haplotype. USF1 encodes a transcription factor known to regulate several genes of glucose and lipid metabolism 14-17 .To identify the gene on 1q21 associated with FCHL, we initially sequenced four functionally relevant regional candidates: TXNIP, USF1, retinoid X receptor gamma (RXRG) and apolipoprotein A-II (APOA2). In parallel, we carried out a functionally unbiased genetic analysis of 60 single-nucleotide polymorphisms (SNPs) in 26 genes in 42 families with FCHL, including the 31 families in the original linkage study 3 . We then genotyped the ten SNPs most likely to be relevant in the extended sample of 60 families of FCHL (Supplementary Table 1 online). Fifty SNPs were located in a 5.8-Mb region flanking the peak markers D1S104 and D1S1677 (Fig. 1). All the families that we studied included a proband with severe coronary heart disease and an abnormal lipid phenotype and an average of 5-6 members affected with FCHL.We sequenced the entire TXNIP gene and the 2,000-bp upstream DNA region in 60 FCHL probands. Of the 20 SNPs identified, none resulted in amino acid changes, and all were rare, with a maximal 7% allele frequency. We also did not observe the nonsense mutation causing hyperlipidemia in mice 13 . We genotyped the four most common SNPs in the 60 families with FCHL but found no evidence of association Table 2 for distances, SNP numbers and LD clusters of these SNPs). (c) The SNPs associated with triglyceride levels in men and (d) the SNPs associated with FCHL and triglycerides in all family members.

We performed a genomewide scan for genes that predispose to low serum HDL cholesterol (HDL-C) in 25 well-defined Finnish families that were ascertained for familial low HDL-C and premature coronary heart disease. The potential loci for low HDL-C that were identified initially were tested in an independent sample group of 29 Finnish families that were ascertained for familial combined hyperlipidemia (FCHL), expressing low HDL-C as one component trait. The data from the previous genome scan were also reanalyzed for this trait. We found evidence for linkage between the low-HDL-C trait and three loci, in a pooled data analysis of families with low HDL-C and FCHL. The strongest statistical evidence was obtained at a locus on chromosome 8q23, with a two-point LOD score of 4.7 under a recessive mode of inheritance and a multipoint LOD score of 3.3. Evidence for linkage also emerged for loci on chromosomes 16q24.1-24.2 and 20q13.11, the latter representing a recently characterized region for type 2 diabetes. Besides these three loci, loci on chromosomes 2p and 3p showed linkage in the families with low HDL-C and a locus on 2ptel in the families with FCHL.

Coronary heart disease (CHD) is a complex disorder constituting a major health problem in Western societies. To assess the genetic background of CHD, we performed a genomewide linkage scan in two study samples from the genetically isolated population of Finland. An initial study sample consisted of family material from the northeastern part of Finland, settled by a small number of founders approximately 300 years ago. A second study sample originated from the southwestern region of Finland, settled approximately 2,000 years ago. Families were ascertained through probands exhibiting premature CHD, defined as >50% stenosis of at least two coronary arteries at a young age, as verified by coronary angiography. Both study samples and the pooled data set provided evidence for linkage in two chromosomal regions. A region on chromosome 2q21.1-22 yielded two-point LOD scores of 3.2, 1.9, and 3.7, in the affected sib-pair (ASP) analyses of the northeastern, southwestern, and pooled study samples. The corresponding multipoint maximum-likelihood scores (MLSs) for these three study samples were 2.4, 1.3, and 3.0. In addition, a region on chromosome Xq23-26 resulted in two-point LOD scores of 1.9, 3.5, and 2.9 and in multipoint MLSs of 3.4, 3.1, and 2.5, respectively. In conclusion, this study identifies two loci likely to contribute to premature CHD: one on chromosome 2q21.1-22 and another on chromosome Xq23-26.