Evidence for cis-eQTL in stimulated <i>versus</i> non-stimulated cells.

<p>For each gene, we plotted the SNP with the lowest <i>P</i> value obtained under an additive model in one condition (stimulated or non-stimulated) against the <i>P</i> value obtained under the alternative condition. Red and grey dashed lines correspond to the 0.01 and to the 0.5 FDR to classify response eQTL (reQTL). Green dots are general cis-eQTL (found in both conditions). Blue dots are reQTL specific to cells stimulated with <i>M</i>. <i>leprae</i> sonicate while pink dots are reQTL specific to untreated cells. For this figure, reQTL are variants that exhibit a significant <i>P</i> value for genotype-phenotype association in one condition at an FDR of 0.01 and not in the other condition at an FDR of 0.5 (without taking the entire 200-kb tested regions per gene into account). The orange cloud corresponds to all the variants detected as being cis-eQTL at an FDR of 0.01.</p

Family based sample and study design.

<p>Two sets of families were employed: those with T1R-affected offspring and those with leprosy but T1R-free offspring. The T1R-affected subset comprised 229 offspring belonging to 221 families while the T1R-free subset comprised 229 offspring in 209 families. Offspring were matched by clinical leprosy subtype in the two family sets. In a first analysis stage, the transmission disequilibrium test (TDT) was used to estimate significance of association of <i>LRRK2</i> variants with disease in each subset. In a second stage, a formal heterogeneity test was performed to identify <i>LRRK2</i> variants preferentially associated with T1R.</p

Haplotype analysis of independent signal of association with T1R.

<p>Haplotype analysis of independent signal of association with T1R.</p

Evidence for cis-eQTL in stimulated <i>versus</i> non-stimulated cells.

<p>For each gene, we plotted the SNP with the lowest <i>P</i> value obtained under an additive model in one condition (stimulated or non-stimulated) against the <i>P</i> value obtained under the alternative condition. Red and grey dashed lines correspond to the 0.01 and to the 0.5 FDR to classify response eQTL (reQTL). Green dots are general cis-eQTL (found in both conditions). Blue dots are reQTL specific to cells stimulated with <i>M</i>. <i>leprae</i> sonicate while pink dots are reQTL specific to untreated cells. For this figure, reQTL are variants that exhibit a significant <i>P</i> value for genotype-phenotype association in one condition at an FDR of 0.01 and not in the other condition at an FDR of 0.5 (without taking the entire 200-kb tested regions per gene into account). The orange cloud corresponds to all the variants detected as being cis-eQTL at an FDR of 0.01.</p

LRRK2 variants preferentially associated with T1R.

<p>LRRK2 variants preferentially associated with T1R.</p

Host versus pathogen control of <i>LRRK2</i> expression levels.

<p><i>LRRK2</i> expression levels for 53 unrelated subjects are indicated on the y-axis and stratified according to rs2404580 genotypes on the x-axis. The left panel represents baseline expression while the right panel indicates gene expression levels following stimulation with <i>M</i>. <i>leprae</i> antigen.</p

Multivariate analysis of <i>LRRK2</i> variants with T1R

<p>Multivariate analysis of <i>LRRK2</i> variants with T1R</p

Proposed mechanism for LRRK2 in T1R.

<p>The LRRK2 M2397T amino acid substitution affects protein turnover. The methionine variant of LRRK2 displays a half-life of approximately 8 hours while the half-life of the threonine variant is 18 hours [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0004412#pntd.0004412.ref034" target="_blank">34</a>]. LRRK2 arrests the NFAT transcription factor in the cytoplasm through a complex mechanism mediated by Ca²⁺ [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0004412#pntd.0004412.ref036" target="_blank">36</a>]. This prevents NFAT to migrate to the nucleus and trigger the expression of pro-inflammatory cytokines [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0004412#pntd.0004412.ref035" target="_blank">35</a>]. The M2397 allele is in tight linkage disequilibrium with alleles of SNPs that promote an increase in <i>LRRK2</i> expression creating a compensatory mechanism to counterbalance the shorter LRRK2-M2397 half-life. This compensatory mechanism is abrogated in the presence of <i>M</i>. <i>leprae</i> antigen. Hence, the effect of the M2397T amino acid substitution is most pronounced in the presence of <i>M</i>. <i>leprae</i> antigen.</p

Enrichment of shared risk-alleles associated with T1R leprosy and IBD phenotypes.

<p>GWAS loci for (A) Inflammatory Bowel Disease [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006637#pgen.1006637.ref024" target="_blank">24</a>], (B) Schizophrenia [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006637#pgen.1006637.ref025" target="_blank">25</a>], (C) Human height [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006637#pgen.1006637.ref026" target="_blank">26</a>] and (D) Human blood metabolites [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006637#pgen.1006637.ref027" target="_blank">27</a>] are represented by a single SNP with the strongest evidence for association. The count of SNPs is given below each central blue pie (panels A—D) for each disease. The light blue pie slices indicate the numbers of SNPs that were not available for comparisons with the leprosy families. The proportions of shared SNPs with T1R-free leprosy (left light green pie) and T1R leprosy (right light orange pie) are indicated as pies for each of the GWAS sets (panel A—D). The pies in panel E represents the totality of SNPs tested in the present study for T1R-free and T1R-affected families. The yellow slices indicate the proportion of SNPs with nominal evidence for association with T1R-free leprosy or T1R/leprosy. The darker shade of yellow represent the proportion of SNPs that are significantly heterogeneous between T1R free and affected leprosy patients. The hypergeometric test used as baseline the proportion of nominally associated GWAS SNPs to estimate the significance of enrichment of the T1R/leprosy and T1R-free leprosy GWAS SNPs among the four selected disease phenotypes.</p

Study design to identify genetic variants associated with T1R in three independent samples.

<p>In all samples T1R-affected subjects were matched with T1R-free subjects according to their clinical sub-type of leprosy. For the discovery phase, two sets of families with leprosy-affected offspring (T1R-free or T1R-affected) were selected from our records of Vietnamese leprosy families. A transmission disequilibrium test (TDT) was used to evaluate the non-random transmission of alleles from parents to offspring in the T1R-affected and T1R-free families independently. Next, a formal heterogeneity test was used to contrast the associations in the two family sets and to identify specific associations with the T1R-affected subset. Odds ratios were estimated by conditional logistic regression using the un-transmitted allele as a pseudosib control and compared to the actual offspring case in a matched case-control design. Subsequently, we recruited independent leprosy cases from Vietnam and Brazil for replication and validation of associations in the discovery phase. In these population-based case-control designs T1R-affected were compared with T1R-free subjects using a logistic procedure with adjustment for age at leprosy diagnosis and gender.</p