The accumulated findings reported here suggest that variation in SORCS1 sequence, expression, and function may influence the development of AD. Although the identity of the specific AD-associated sequence variations in SORCS1 remains to be determined, our results imply that (1) there are different AD-associated allelic variants in the SORCS1 gene in different populations; (2) these variants are likely to be in intronic regulatory sequences that effect cell type–specific or tissue-specific expression of SorCS1; and (3) that genetic variation in SORCS1 might affect AD risk by altering the physiological role of SorCS1 in the processing of APP holoprotein.
Similar to observations on the SORCS1
no single SNP or haplotype was associated with AD in all datasets. In addition, the direction of the effect of some of the disease-associated alleles differed between some of the datasets, which we attribute to population differences in LD with the true genetic effector. Another potential explanation for these series-related differences could be confounding genetic or environmental factors that are influential in 1 dataset but not the others. Several issues diminish the possibility that the association between SORCS1
and AD is spurious. First, the association was initially identified in the NIA-LOAD family 6k dataset (rs7082289; SNP 1) using conservative family-based association tests, which are less sensitive to confounding due to population stratification. Second, several alleles and haplotypes were associated with altered risk for AD in at least 2 unrelated data sets. Third, the associations of several SNPs remained significant in meta-analyses that included the Toronto dataset. Differences in the LD patterns between the datasets are a possible explanation for the inverse direction of the association with AD in the Toronto dataset as compared to the other Caucasian datasets. It is possible that the minor alleles of significant SNPs are in linkage with risk alleles in some datasets or populations but in linkage with protective variants in others. Of note, the direction of the effect of SNP 11 in the Toronto dataset was similar to the direction of the effect in the Caribbean Hispanic and MIRAGE African American datasets. Alternatively, the genotyped variants are not the disease-causing variants but rather identify protective or harmful disease-modifying variations in SORCS1
. Fourth, the finding of association with different SNPs in different ethnic groups is a not an unusual observation in complex diseases.38
The occurrence of pathogenic mutations across multiple domains of disease genes (allelic heterogeneity) and the absence of these variants in some datasets or ethnic groups (locus heterogeneity) are frequently observed in both monogenic and complex traits.39
Because the 16 genotyped SNPs do not cover the whole genetic variation in the SORCS1
gene (see Supporting Information ), it is possible that additional polymorphisms in nontagged regions of the gene are associated with AD risk.
Also, it needs to be acknowledged that the sample sizes of the individual datasets were modest. Thus, it remains possible that larger individual datasets would have detected additional genotype-phenotype associations with smaller effect sizes or allele frequencies. Our meta-analyses of all Caucasian datasets, which included in total 2,309 cases and 3,482 controls, confirmed the findings for SNPs 4–9 derived from the individual study samples, and in addition pointed to SNP 13 as significantly associated with AD.
Our finding of a role of SORCS1 in AD is also supported by the results of our RT-PCR and brain microar-ray analyses. We found significantly lower SorCS1 expression levels in amygdala from AD brains compared to the controls, and decreases in SorCS1 expression levels at several probe sites were associated with closely located SNPs. In contrast, when tissue from regions that are less affected by the AD process (ie, cerebellum and occipital lobe) was used, SorCS1 expression did not differ between AD cases and controls. However, while plausible and consistent with our results from the genetic association studies and cell biological experiments, alternative explanations for these differences in expression levels must be discussed. It remains possible that SORCS1 expression levels in the amygdala were influenced by differential cell loss that is more pronounced in AD brains. Alternatively, differential expression between AD and control brains exists throughout the brain but could be more pronounced in the amygdale, and smaller differences in the control brain regions (ie, cerebellum and occipital cortex) may not have been detected due to small sample size.
Our data also revealed that high levels of SorCS1 result in modest (~30%) reductions in both Aβ
40 and Aβ
42, which would be protective. Conversely, the RNAi knockdown of SorCS1 on APP processing had the inverse observation of an increase in Aβ
levels (2–3-fold). In addition, the APP-GV assay, which detects γ
demonstrated that the reduction of SorCS1 leads to a greater than 3-fold increase of γ
-secretase activity on APP processing. Reminiscent of AD, the APP protein level and maturation was found to be unaffected, suggesting altered trafficking and/or increased activity of γ
-secretase, leading to the generation of Aβ
as likely mechanisms.
The mechanisms by which overexpression of SorCS1 transcripts modulates Aβ production is not immediately clear, but likely involves binding of SorCS1 to the APP holoprotein or to its processing enzymes, possibly separating APP away from BACE1 and γ-secretase cleavage.
In summary, while the genetic and biochemical data both infer a relationship between SorCS1 and the AD process, it is presently unclear how this is mediated. It is tempting to speculate that intragenic, noncoding polymorphisms in SORCS1 might account for the modest, yet consistent association with risk for AD, and might act by modulating SorCS1 expression. In particular, we hypothesize that high basal levels of SorCS1 expression in some individuals might have a protective effect, whereas low levels of expression may lead to elevated APP processing into Aβ as found in AD. Additional studies will be needed to determine whether carriers of alleles associated with differential risk for AD are indeed protected and that protection arises because of high levels of expression of SorCS1.