Tight Clustering of Extracellular BP180 Epitopes Recognized by Bullous Pemphigoid Autoantibodies

Abstract

Bullous pemphigoid is a blistering skin disease associated with autoantibodies against the BP180 antigen, a transmembrane component of the hemidesmosome. Anti-BP180 antibodies have been demonstrated to be pathogenic in a passive transfer mouse model. One extracellular site on human BP180 (MCW-1) was previously shown to be recognized by 50-60% of bullous pemphigoid sera. To facilitate the identification of additional autoantibody-reactive epitopes, recombinant forms of the BP180 ectodomain were generated using both bacterial and mammalian expression systems. One recombinant protein, sec180e, that was expressed in COS-1 cells and that contained the entire BP180 ectodomain, provided us with a tool to detect conformational epitopes. Bullous pemphigoid sera immunoadsorbed against the major noncollagenous NC16A domain no longer reacted with sec180e, indicating that autoantibody reactivity to the BP180 ectodomain is restricted to the NC16A region. Immunoblot analysis of bullous pemphigoid sera immunoadsorbed with a series of recombinant NC16A peptides revealed the presence of three novel autoantigenic sites that, along with the MCW-1 epitope, are clustered within the N-terminal 45 amino acid stretch of NC16A. All 15 bullous pemphigoid sera tested reacted with a recombinant protein containing this BP180 segment. No disease-associated epitopes were detectable within the remaining 28 amino acids of NC16A. Thus, bullous pemphigoid patient autoantibodies react with a set of epitopes on the BP180 ectodomain that are highly clustered. This autoantibody-reactive region on human BP180 shows overlap with the corresponding murine BP180 site that is targeted by antibodies that are pathogenic in the mouse model of bullous pemphigoid. These findings suggest new directions for the development of diagnostic and therapeutic tools for this disease.

What’s new in bullous pemphigoid

“…18,19 In addition, several major antigenic sites have been identified within the N-terminal 45 amino acid stretch of NC16A, while the remaining 28 amino acids of NC16A were shown to have no BP-associated epitopes. 17 Several studies have confirmed that the serum levels of anti-COL17 NC16A autoantibodies correlate with the disease severity of BP. 20,21 In addition to these clinical observations, the pathogenicity of anti-COL17 NC16A IgG antibodies from BP patients was shown by in vitro study.…”

“…5). 16,17 Epitope mapping using various fragments of COL17 has shown that sera from most BP patients recognize NC16A. 16 ELISA analysis using recombinant COL17 NC16A demonstrated that 94-96% of BP sera reacts to COL17 NC16A peptides.…”

Pemphigoid gestationis: new insights into the pathogenesis lead to novel diagnostic tools

“…4). Four of these epitopes (NC16A1 through NC16A3) are also recognised by sera from patients with bullous pemphigoid and linear IgA disease 44,45 . In contrast, PG sera do not react with epitope NC16A4, which is a major target of autoantibodies in lichen planus pemphigoides 46,47 .…”

Context-Dependent Regulation of Collagen XVII Ectodomain Shedding in Skin

“…Previous studies have demonstrated that COL17 is the major autoantigen (autoAg) not only for BP but also for other pemphigoid disorders, including linear IgA bullous dermatosis (LAD), mucous membrane pemphigoid, pemphigoid gestationis and lichen planus pemphigoides 3 . The major epitopes for BP autoAbs tightly cluster within the juxtamembranous non-collagenous NC16A domain of COL17, and the majority of IgG autoAbs react with this region 6,7 . Passive transfer experiments of rabbit IgG 8 and autoAbs from BP patients 9 into wild-type (WT) or transgenic (Tg) neonatal mice have demonstrated pathogenic roles of antibodies (Abs) to the NC16A domain in blister formation.…”