Genes at human chromosome 5q31.1 regulate delayed-type hypersensitivity responses associated with Leishmania chagasi infection

Abstract

Visceral leishmaniasis (VL) caused by Leishmania chagasi is endemic to northeast Brazil. A positive delayed-type hypersensitivity skin test response (DTH+) is a marker for acquired resistance to disease, clusters in families and may be genetically controlled. Twenty-three single nucleotide polymorphisms (SNPs) were genotyped in the cytokine 5q23.3-q31.1 region IRF1-IL5-IL13-IL4-IL9-LECT2-TGFBI in 102 families (323 DTH+; 190 DTH-; 123 VL individuals) from a VL endemic region in northeast Brazil. Data from 20 SNPs were analyzed for association with DTH+/- status and VL using family-based, stepwise conditional logistic regression analysis. Independent associations were observed between the DTH+ phenotype and markers in separate linkage disequilibrium blocks in LECT2 (OR 2.25; P=0.005; 95% CI=1.28-3.97) and TGFBI (OR 1.94; P=0.003; 95% CI=1.24-3.03). VL child/parent trios gave no evidence of association, but the DTH- phenotype was associated with SNP rs2070874 at IL4 (OR 3.14; P=0.006; 95% CI=1.38-7.14), and SNP rs30740 between LECT2 and TGFBI (OR 3.00; P=0.042; 95% CI=1.04-8.65). These results indicate several genes in the immune response gene cluster at 5q23.3-q31.1 influence outcomes of L. chagasi infection in this region of Brazil.

Figure 1

Drawing of the chromosome 5q23.3-q31.1 region showing relative positions of genes and SNPs (not to scale). Boxes indicate the locations of genes including both exons and introns. Letters referring to SNPs (dotted vertical arrows; open dots are SNPs associated with the DTH− phenotype; grey-filled dots are SNPs associated with the DTH+ phenotype) are defined in Table 2. Distances between the start of the 5' UTR of IRF1 and end of the 3'UTR of IL4, the 5'UTR of SEC24 and 3'UTR of DCOL, and the 5'UTR of IL9 and 3'UTR of TGFBI are indicated with dotted horizontal arrows. Solid horizontal arrows indicate the direction of the coding region for selected genes. Brackets refer to markers in LD clusters (H to O; R to V) as defined by D' (see figure 2). Markers not shown on the figure were not analyzed statistically, either because there was only one allele or they were out of Hardy Weinberg Equilibrium. (See methods section for details.)

Figure 2

LD plot generated using the default settings in Haploview available from the HapMap Project site (http://www.hapmap.org/). LD between pairs of markers across chromosome 17q11.1-q22 was determined within Haploview using (A) D' or (B) r² statistics. LD values between markers are indicated at the intercept of the two markers on the matrix. Empty boxes indicate that the LD value is 1. Intensity of colour on the red/pink or black/grey scale indicates the degree of confidence in the LD value. Two main haplotype blocks outlined within the black triangles lie within this region of 5q23.3-q31.1: rs31530 (H) -rs30740 (O) encompassing LECT2; and rs4976360 (R) - rs7725447 (V) encompassing TGFBI. All SNPs used had minor allele frequencies >0.1 (see table 2).

Figure 3

Shows the SYNPLOT output for the multiple alignment of (top to bottom) human, dog, cow and mouse sequences carried out across the region LECT2 to TGFBI on chromosome 5q31.1 using LAGAN. On the 4-species alignments red boxes define exons at the two genes, blue boxes indicate repetitive sequence, green boxes show GENSCAN auto-annotated putative exons, vertical black bars show the positions of SNPs (J to U, see table 2) used in this study. The plot shows degree of conservation of sequence across all 4 species, on a vertical scale 0-100%. CNS are defined as regions of conserved sequence that are approaching or similar in magnitude to that observed at the known exons for the two genes. Examples of potential regions of CNS of interest are indicated by vertical red arrows, including the single CNS showing high conservation between these mammalian species and chicken (data not shown).