Identification of a Functional Risk Variant for Pemphigus Vulgaris in the <i>ST18</i> Gene

<div><p>Pemphigus vulgaris (PV) is a life-threatening autoimmune mucocutaneous blistering disease caused by disruption of intercellular adhesion due to auto-antibodies directed against epithelial components. Treatment is limited to immunosuppressive agents, which are associated with serious adverse effects. The propensity to develop the disease is in part genetically determined. We therefore reasoned that the delineation of PV genetic basis may point to novel therapeutic strategies. Using a genome-wide association approach, we recently found that genetic variants in the vicinity of the S<i>T18</i> gene confer a significant risk for the disease. Here, using targeted deep sequencing, we identified a PV-associated variant residing within the <i>ST18</i> promoter region (p<0.0002; odds ratio = 2.03). This variant was found to drive increased gene transcription in a p53/p63-dependent manner, which may explain the fact that ST18 is up-regulated in the skin of PV patients. We then discovered that when overexpressed, ST18 stimulates PV serum-induced secretion of key inflammatory molecules and contributes to PV serum-induced disruption of keratinocyte cell-cell adhesion, two processes previously implicated in the pathogenesis of PV. Thus, the present findings indicate that ST18 may play a direct role in PV and consequently represents a potential target for the treatment of this disease.</p></div

ST18 promotes PV serum- and PV IgG-induced release of pro-inflammatory cytokines.

<p>NHEKs were transfected with an ST18 expression vector (ST18, dark grey) or control empty vector (EV, light grey). (a,b,c) Supernatants were collected 2, 4, 6 and 24 hours post exposure to PV serum or control serum and (a) TNFα, (b) IL-1α and (c) IL-6 secretion was measured as described in Materials and Methods; (d,e,f) Supernatants were collected 6 hours (IL-1α and IL-6) or 24 hours (TNFα) post exposure to PV and control serum or PV and control IgG and (d) TNFα, (e) IL-1α and (f) IL-6 secretion was measured as described in Materials and Methods. Results represent the mean of three independent experiments and are expressed as the relative cytokine secretion in percentage, compared to control (empty vector) ± SE (*p<0.05, **p<0.01, ***p<0.001 and ****p<0.0001 by 2-tailed t test).</p

ST18 promotes PV serum and PV IgG-induced cell-cell disadhesion.

<p>We used a dispase-based dissociation assay to evaluate the effect of ST18 overexpression on cell adhesion. NHEKs transfected with a ST18 expression vector (ST18) or with a control vector (EV) were grown to confluency in the presence of PV serum and control serum or PV IgG and control IgG. (a) Epidermal sheets were released from the tissue plates and subjected to mechanical stress as described in Materials and Methods; (b) the resulting fragments were counted. Results represent the mean of three independent experiments and are expressed as number of fragments ± SE (*p<0.05, **p<0.01, ***p<0.001 by 2-tailed t test).</p

Targeted deep sequencing of the ST18 gene locus.

<p>Deep sequencing of the ST18 gene locus in 16 PV patients led to identification of 789 genetic variants (black dots), depicted along the ST18 gene locus (X axis, chr8:53,023,392–53,373,519, GRCh37/hg19 assembly; Y axis, negative log-transformed P-values of association score). A case-control association analysis against the 1000 genome project data revealed a large PV-associated haplotype block (in red) residing within an ST18 intron and harboring rs17315309 (arrows).</p