The accumulated findings reported herein suggest that variation in the LRRTM3
sequence, expression, and function may influence the development of AD. Although the identity of the specific AD-associated sequence variations in LRRTM3
remains to be determined, our results from the association studies imply (1) that there are different AD-associated allelic variants in the LRRTM3
gene in different populations and (2) that some of these variants are likely to be in intronic regulatory sequences that affect cell type–specific or tissue-specific expression of LRRTM3.
The results from our γ-secretase assays suggest that genetic variation in LRRTM3
might affect AD risk by altering the physiologic role of LRRTM3 in the processing of APP holoprotein. These 2 findings are complementary but independent. We could not examine, and therefore could not conclude, that the specific polymorphisms associated with AD in our data sets affect γ-secretase processing of APP or LRRTM3 levels in any direction. We could only demonstrate independent effects of genetic variants in LRRTM3 on AD risk and of LRRTM3 knockdown on γ-secretase processing. However, both independent findings are in line with a role of LRRTM3 in AD as has been suggested before—namely, that genetic variations in LRRTM3
are associated with AD6–10
and that LRRTM3 may promote APP processing through an effect on APP cleavage.5,10
Several issues diminish the possibility that the association between LRRTM3
and AD is spurious. First, several alleles and their corresponding haplotypes were associated with altered AD risk in 2 unrelated data sets from different ethnic groups. Second, the strength of the association (OR and P
value) of rs10997477 with AD increased in a meta-analysis of both data sets. Although in the Caribbean Hispanic data set a haplotype (rs10822970|rs2619652|rs2764813) that was significantly associated with AD was located in the same LD block as SNP rs10997477, which was significant in the NIALOAD data set, our results are consistent with the notion that there are different disease-associated variants in different ethnic groups. The occurrence of pathogenic mutations across multiple domains of disease genes (allelic heterogeneity) and the absence of these variants in some data sets or ethnic groups (locus heterogeneity) are frequently observed in both mono-genic and complex traits.34,35
An alternative explanation for the fact that different SNPs are associated with AD in the 2 ethnic groups may be differences in LD patterns. It is likely that the genotyped variants are not the disease-causing variants but rather are in LD with causative protective or harmful disease-modifying variations in LRRTM3
As previously described, rs1925608, rs7082306, and rs1925609 belong to a distinct LD block in intron 2 containing several regulatory regions or transcription-factor binding sites. Thus, it seems likely that these 3 SNPs point to the same disease-associated variant. The fact that this block is not in LD with regions outside LRRTM3
supports the notion that the genetic association of LRRTM3
with AD is independent of a potential association of CTNNA3
with AD. Our finding of a role of LRRTM3
in AD is also supported by the results of our cell experiments demonstrating an effect of LRRTM3 on γ-secretase processing of APP. These observations are consistent with the findings by Majercak et al5
that small-interfering RNAs targeting LRRTM3 inhibit the secretion of Aβ40 and Aβ42.
The fact that the effect sizes of associated SNPs were small (OR, 1.1–1.2) is expected for a common disease and in line with the recently detected novel AD susceptibility loci identified by large genome-wide association studies.36–40
It has to be acknowledged that the sample sizes of both individual data sets were modest and we had 80% power to detect effect sizes of OR of 1.18 or larger. Thus, it remains possible that larger individual data sets would have detected additional genotype-phenotype associations with smaller effect sizes or allele frequencies. It is also possible that additional polymorphisms in nontagged regions of the gene are associated with AD risk. A second limitation is that we were not able to directly examine the effect of specific disease-causing mutations on γ-secretase processing of APP or LRRTM3 levels. That would require resequencing of LRRTM3 because the polymorphisms identified in the present study likely are not the disease-causing variants but rather are in LD with causative protective or harmful disease-modifying variations.
In summary, our findings from genetic epidemio-logic and functional analyses provide modest support for a role of LRRTM3 in AD. Sequencing studies are needed to identify the specific disease-causing mutations and to examine whether they are associated with differences in APP processing through effects on function or level of LRRTM3. If our findings are confirmed, this would hold the promise of an LRRTM3 as a therapeutic target for AD and other related amyloid disorders.