AIRE-PHD fingers are structural hubs to maintain the integrity of chromatin-associated interactome

Abstract

Mutations in autoimmune regulator (AIRE) gene cause autoimmune polyendocrinopathy candidiasis ectodermal dystrophy. AIRE is expressed in thymic medullary epithelial cells, where it promotes the expression of peripheral-tissue antigens to mediate deletional tolerance, thereby preventing self-reactivity. AIRE contains two plant homeodomains (PHDs) which are sites of pathological mutations. AIRE-PHD fingers are important for AIRE transcriptional activity and presumably play a crucial role in the formation of multimeric protein complexes at chromatin level which ultimately control immunological tolerance. As a step forward the understanding of AIRE-PHD fingers in normal and pathological conditions, we investigated their structure and used a proteomic SILAC approach to assess the impact of patient mutations targeting AIRE-PHD fingers. Importantly, both AIRE-PHD fingers are structurally independent and mutually non-interacting domains. In contrast to D297A and V301M on AIRE-PHD1, the C446G mutation on AIRE-PHD2 destroys the structural fold, thus causing aberrant AIRE localization and reduction of AIRE target genes activation. Moreover, mutations targeting AIRE-PHD1 affect the formation of a multimeric protein complex at chromatin level. Overall our results reveal the importance of AIRE-PHD domains in the interaction with chromatin-associated nuclear partners and gene regulation confirming the role of PHD fingers as versatile protein interaction hubs for multiple binding events.

Aire’s plant homeodomain(PHD)-2 is critical for induction of immunological tolerance

“…Such observations suggest that Aire might participate in multiple multiprotein complexes, differentially dependent on its individual domains. This notion is consistent with recent 3D structural data on a tandem Aire PHD1-PHD2 fragment, which revealed the two domains to be independent and noninteracting (13). This study also showed that point mutations in PHD1 that have a major impact on PTA gene expression had relatively minor effects (less than two-fold) on the interaction of Aire with protein partners involved in chromatin binding/structure or transcription (e.g., RuvBL2, SMC1A, DNA-PKcs, and Msh6).…”

“…It seems not to operate like a traditional transcription factor, binding to a promoter and/or enhancer element and inducing transcript initiation, but instead to somehow release RNA polymerase (RNA-Pol)II pausing and promote transcript elongation (9,10). Aire targets and regulates genes as a component of one or more large multiprotein complexes (11)(12)(13). Its numerous associates include sets of proteins involved in nuclear transport, chromatin binding/structure, transcription, and pre-mRNA processing (12).…”

“…PHD1 has also been reported to be a vital link in Aire's interaction with DNA-PKcs (20) and to harbor E3 ubiquitin-ligase activity essential for transcriptional induction (21), although the NMR solution structure and subsequent functional assessments did not support this latter role (22). The function of Aire's more carboxyl-terminal PHD2 remains an enigma, with arguments both for and against a critical role in transcriptional transactivation (11,13,16,21,(23)(24)(25).…”

Induction de la tolérance centrale dans le thymus par le facteur de transcription Aire

“…La reconnaissance de cette marque épigénétique est nécessaire à la transcription des TRA [25][26][27] (Figure 2B). Le domaine PHD2, quant à lui, n'interagit pas directement avec le nucléosome mais contribue à la structure protéique tridimensionnelle de Aire qui est essentielle à son activité transcriptionnelle [28]. Le domaine PHD1 est aussi impliqué dans l'interaction de Aire avec la kinase nucléaire, DNA-PK (DNA dependent protein kinase).…”

The Mechanism of Tissue-Restricted Antigen Gene Expression by AIRE

“…These findings raise the conundrum of how AIRE can regulate such a large repertoire of divergent genes. Recent work from others and us has addressed this question and revealed the mechanisms of action of this enigmatic protein (26)(27)(28)(29)(30)(31).…”