We conducted a genomewide ordered subset linkage analysis for AD using admixture proportion as a covariate in AAs. Our results showed that admixture proportion can be a useful covariate to identify the subset of families which are more homogeneous and thereby increase power for linkage detection in AAs. We identified four ordered subsets with statistically significant increases in lod scores on chromosomes 4, 12, 15 and 22.
Our most interesting finding was on chromosome 4, where we observed the strongest linkage signal (lod = 4.2) at 180 cM in a subset of 44 families with an African ancestry proportion ranging from 0.858 to 0.996. Linkage evidence for alcohol-related phenotypes on chromosome 4 has been reported in many previous studies. However, the locus identified here does not appear to overlap with any chromosome 4 linkage signals reported previously, with the exception of the study by Wilhelmsen (Wilhelmsen et al. 2003
), which yielded modest evidence for linkage to the trait of “level of response to alcohol” (peak lod = 1.4, 170–190 cM). The 1-lod support interval (177–185 cM) for the chromosome 4 peak region contains 50 genes and harbors a particularly promising AD risk candidate gene, GLRA3
. This gene encodes the alpha-3 subunit of the neuronal glycine receptor, a primary mediator of neuronal inhibition in the brain regions known to be sensitive to ethanol (Perkins et al. 2010
The second strongest linkage signal (lod = 3.2) obtained by the current OSA is located at 46 cM on chromosome 22 in a subset of 33 families with a high proportion of African ancestry ranging from 0.885 to 0.996. Wilhelmsen et al. (Wilhelmsen et al. 2003
) found a suggestive linkage (lod = 2.9) peak to the trait of “level of response to alcohol” in the region of 20–30 cM on chromosome 22; however, this linkage region does not overlap the 1-lod support interval (45–49 cM) for our finding. However, considering the small number of families contributing to our observed linkage signal, and the low mapping location precision possible with ASPs, it is possible that these findings are in fact reflective of the same risk gene. Three candidate genes located in the 1-lod support interval of the linkage peak on chromosome 22 are particularly interesting: PACSIN2
, which encodes protein kinase C and casein kinase substrate in neurons; NPTXR
, which encodes the neuronal pentraxin receptor; and SYNGR1
, which encodes an integral membrane protein associated with presynaptic vesicles in neurons.
We also identified two regions with suggestive evidence of linkage on chromosomes 12 and 15 in two ordered subsets with lower proportions of African ancestry (i.e., ranging from 0.443 to 0.813 and from 0.443 to 0.801 for chromosomes 12 and 15, respectively). We did not identify compelling candidate genes from the 1-lod support interval for these two regions.
We also examined the characteristics of the 124 families and the families in the subsets to evaluate whether the increased linkage signals in the subset families could be due to the basic demographic difference between the overall families and the subsets. Supplementary Table 1
illustrates sex and age distribution among the total set of families and the subset families. We did not find any significant difference in age or sex distribution between the subsets and the overall families, suggesting the increased linkage signals in the subsets may not be accounted for by these basic family characteristics.
Samples included in the current study were ascertained mostly for DSM-IV cocaine or opioid dependence. Therefore, we had to consider the possibility that the linkage signals detected for AD in AAs could be attributable to other substance dependence (SD), such as cocaine dependence (CD) and opioid dependence (OD). We examined this alternate explanation for the findings by directly testing linkage for CD or OD in the same subset of families identified for AD on chromosomes 4, 12, 15 and 22.We found a significant linkage on chromosome 4 at 180 cM for CD (LOD = 3.49). Another suggestive linkage signal for CD was also identified on chromosome 22 at 46 cM (LOD = 2.93).The linkage results from other SD might indicate that the linkage signal is not specific for AD and could reflect a shared liability between different substances
Although genome-wide association studies (GWAS) have successfully identified common risk variants associated with complex disorders, the heritability of most complex disorders remains largely unexplained. It is now widely believed that the “missing heritability” could be attributable to rare variants of high penetrance or structural variation that is not well tagged by the current GWAS approach (Manolio et al. 2009
). Compared with GWAS design, linkage analysis has the advantage of enriching signals for the same gene/functional unit that may harbor multiple rare variants of large effect or inherited CNVs segregating in families. With the growing evidence for the role of rare variants and CNVs in psychiatric disorders (Stefansson et al. 2008
; Walsh et al. 2008
; Williams et al. 2010
), the regions discovered by linkage may complement the GWAS approach in pinpointing rare variants of high penetrance that cannot be captured using the current GWAS design. Thus, further investigation through targeted deep sequencing studies of the linkage regions identified by the current OSA may identify novel genes containing multiple rare or uncommon AD risk variants with large effects.
In summary, our results demonstrate that the genetic ancestry proportion can be used as a covariate to reduce genetic heterogeneity and enhance the detection of linkage for AD in AA samples. The current OSA approach based on admixture proportion could be applied to any linkage study for complex traits conducted in AA or other admixed populations.