Introduction
Psoriasis vulgaris [MIM 177900] is a chronic inflammatory dermatosis with a prevalence of 0.5-5%, depending on race and countriy. Psoriasis is a multi-factorial disease involving the interaction of several genes as well as environmental factors like streptococcal infections, physical trauma, drug usage, and stress. Genome-wide scans have revealed linkage to several re¬gions across the genome. Linkage has been shown on chro¬mosomes 6p21, 1q21, 1p and 16q. Suggestive linkage inclu¬des regions at chromosomes 3p, 8q, 15p and 20p. Other stu¬dies have also shown chromosomes 17q and 4q to be of interest but these regions have not been replicated in any genome-wide scans (reviewed in 1). All the genomewide studies have revealed highly significant linkage to a region on the major histocompatability complex (MHC) at 6p21.3, designated “psoriasis susceptibility region 1” (PSORS1). Within that region, HLA-Cw6 was the first allele to show a strong association with familial psoriasis; and HLA-Cw6 is still the only marker allele present in all investigated ethnic groups .Our own group refined PSORS1 to a region of approximately 170 kb telemoric of HLA-C (2). A case control study in a Japanese population using microsa¬tellite markers distributed over a 1060 kb region surrounding the HLA-C locus located PSORS1 to an interval of 111 kb telomeric of the HLA-C gene (3). Interestingly these two studies, al¬though based on different ethnic groups, showed an overl¬apping DNA interval of approximately 110 kb sugges¬ting that PSORS1 is likely to be located here. Seven genes have been identified within this candidate DNA interval including POU5F1 or octamer transcription factor 3 gene (OTF3), the cell growth regulated gene TCF19, HCR gene encoding #-helix coiled coil rod homologue protein, SPR1 encoding small proline rich protein, S gene or corneodesmosin (CDSN), SEEK1 and STG. TCF19 is far less poly¬morphic than the other genes in the region and showed no association with psoriasis, suggesting that the true gene(s) for psoriasis could be telomeric to TCF19. However, several other studies also investigating the relation of psoriasis with loci centromeric of HLA-Cw, i.e. MICA, TNF#, and HLA-DRB1 and -DQB1.These studies demon¬stra¬ted that the prevalence of the risk alleles of these loci seems to be propably lower than that of PSORS1 or Cw6, but that the presence of alleles of these loci results in higher risk for psoriasis HLA-Cw. In order to further clarify the psoriasis risk region in the MHC, we examined a 1,923 kb DNA interval located from 910 kb telomeric to 1013 kb centomeric of HLA-C for psoriasis muta¬tion(s).
Patients and Methods
Patients and controls: 530 patients from 56 families with multiple occurance of chronic plaque type psoriasis (age at onset < 40) and 696 healthy controls recruited in the Kiel area were included into the study. For confirmatory purposes, an independent cohort (480 patients, 1468 unaffected) derived from Ann Arbor, MI, was genotyped for a subset of the SNPs (n=40). SNP- and HLA- Genotyping: A total of 99 SNPs was selected encompassing 1,923 kb on 6p21.3 from DDR1 to BTNL2 (table 1). SNP genotyping was done by TaqManR technology. SNPs were selected either from the literature or from the Kiel NGFN SNP program. HLA genotypes (HLA-B, Cw, DRB1, and –DQB1) were estimated using the DYNAL Allset PlusR genotyping kits or by PCR-based HLA genotyping-tests as published (4). Statistical analysis: For case-control and transmission disequilibrium analysis, odds ratios and p-values were calculated for single markers and for marker haplotypes. All calculations were done using the program HAPLOVIEW (5). Haplotype construction: For construction of SNP haplotypes, either HAPLOVIEW or a self-written program (http://www.hlainformatik.de/) were used. Shortly, this program uses the segregation infor¬ma¬tion of chromosome 6 as provided by HLA data and is able to define SNP haplotypes as far as siblings and parents are not heterozygous at a given position. Stratification for PSORS1: Patients and probands were designated to be positive for PSORS1 if they were positive for HLA-Cw6.
Results:
Association analysis: Several regions in the investigated area yielded highly significant association with psoriasis (CDSN, HCR, POU5F1, HCG2-II, KIA0055hom) all p<0.00001; fig 1). All of these loci are already known to demonstrate strong association with psoriasis. However, in both cohorts the calculation of odds ratios and family based transmission disequilibrium ratios yielded the highest values for loci located centro¬meric of HLA-Cw, close to or within the central MHC (fig. 1). The highest odds ratios were observed in both cohorts for P5-1, MICB, and the TNF# promoter polymorphism at position –238.
Haplotype construction: Using 530 individuals from 56 families of the primary cohort, we were able to deduce the SNP sequence of a total of 307 chromosomes. Among those, the phase of 83% of the SNPs could unambigously be identified. Analysis of SNP blocks both by HAPLOVIEW and se¬quence comparison revealed an extremely well conserved block of 30 kb in the region telomeric of HLA-C which comprises 20 SNPs and the genes HCR, TCF19, and POU5F1. This block (block I in fig. 2) is present in 101 of the 307 chromosomes, and 37 of the 46 affected founders are positive for this haplotype. Of the 56 chromosomes positive for Cw6 (18%), 52 (93%) are positive for Block I, and the SNPs which yielded the strongest association (figure 1) are all present in this block.
A further block (block III, Fig. 2) contains the three SNPs which yielded the highest odds ratios. Again, these three SNPS are in almost perfect linkage disequilibrium (LD) and form a well conserved haplotype which in turn is in very close linkage with block I (31 out of 33 chromosomes; 94%). However, block III is less frequent than block I (33 vs 101 chromo¬so¬mes).
Haplotype association: Individuals positive for PSORS1 and Block III were selected and the strenght of asso¬ciation was calculated in dependence of the presence or absence of the respective haplotypes. In both cohorts, the frequency of psoriasis was significantly increased when block III was present in indi¬viduals positive for block I when compared with individuals positive only for block I but negative for block III. Block III occurred only very rarely in the absence of PSORS1 (or HLA-Cw6, respectively). Table 2 shows the results of this analysis.
Discussion:
Psoriasis is a common, immunologically-mediated, hyper¬roliferative skin disease that is influenced by multiple genes, including a major gene in the major histocompatibility complex. Recently discussed candidate genes within the MHC are CDSN, HCR, and HLA-Cw6. However, because the penetrance of the disease allele at this locus is only about 10%, and based on recurrence risk(6) and linkage(1) analysis, it is apparent that additional loci also influence susceptibility to psoriasis. Such additional sus¬ceptibility regions migth be PSORS2-PSORS6 which have been shown to be in significant linkage with the disorder (1). We have analysed association and odds ratios of a total of 99 SNP marker located within the MHC I and MHC III. Not surprisingly, markers located within or close to the PSORS1 region as described in the literature yielded the highest significance. The respective marker alleles are very frequent among patients (> 80%, data not shown) but do not carry high relative risks. In contrast, we observed three markers centromeric of PSORS1, which yield much higher odds ratios (fig. 1) but are less frequent in patients. In order to clarify this discrepancy, we deduced the SNP frequencies of the MHC chromosomes present in the investigated families. These investigations demonstrated that the PSORS1 haplotype is in very close linkage with a second haplotype located centro¬meric of HLA-Cw. The presence of this haplotype signi¬ficantly increases the risk to develop psoriasis, but only if the PSORS1 haplo¬type is also present. Without PSORS1, this haplotype does not confer any risk for psoriasis, indicating that there is an interaction of at least two different genes. Genes lo¬ca¬ted in the centromeric region significantly asso¬ciated with psoriasis are P5.1, TNF# and MICB. Due to their immuno¬logical functions, at least TNF# and MICB are well suited pso¬riasis susceptibility candidates. Taken together, these data indicate that in fact genes additional to the one present in PSORS1 are necessary to develop psoriasis. One of such a gene might also be present in the human MHC, located centromeric of PSORS1 close to the central MHC. Since the psoriasis gene located in PSORS1 is probably in¬volved in the epidermal differentiation dysre¬gu¬la¬tion cha¬rac¬teristic for psorisis, this second gene might be reponsible for the immunological dysregulation of psoriasis.
Project Status
The SNP based gene scan of the MHC is now finished. Due to the observed extreme LD within the investigated areas, genotyping probably won’t provide futher information. Functional analyses now have to show the role of the interacting genes located in that region.
Outlook
To further clarify the role of a second psoriasis gene in the MHC, (i) functional studies will be performed to investigate the role of the identified candidates in the pathogenetics of psoriasis. (ii) The chip based genome wide scan will be used to identify further additional genes in the human genome which interact with PSORS1.
Lit.: 1. Elder JT. Psoriasis clinical registries, genetics, and genomics. Ann Rheum Dis. 2005 Mar;64 Suppl 2:ii106-7. 2. Nair RP et al. Localization of psoriasis-susceptibility locus PSORS1 to a 60-kb interval telomeric to HLA-C. Am J Hum Genet. 2000 Jun;66(6):1833-44. 3. Oka A et al. Association analysis using refined micro¬satellite markers localizes a susceptibility locus for psoriasis vulgaris within a 111 kb segment telomeric to the HLA-C gene. Hum Mol Genet. 1999 Nov;8(12):2165-70. 4. Welsh K et al. Molecular typing for the MHC with PCR-SSP . Rev Immunogenet. 1999;1(2):157-76. 5. Barrett JC et al. Haploview: analysis and visualization of LD and haplotype maps. Bioinformatics. 2005 Jan 15;21(2):263-5. 6: Elder JT et al. The genetics of psoriasis. Arch Dermatol 1994;130(2):216-24.
