Abstract
Aggregate-like biomolecular assemblies are emerging as new conformational states with functionality. Aire, a transcription factor essential for central T cell tolerance, forms large aggregate-like assemblies visualized as nuclear foci. Here we demonstrate that Aire utilizes its caspase activation recruitment domain (CARD) to form filamentous homo-multimers in vitro, and this assembly mediates foci formation and transcriptional activity. However, CARD-mediated multimerization also makes Aire susceptible to interaction with promyelocytic leukemia protein (PML) bodies, sites of many nuclear processes including protein quality control of nuclear aggregates. Several loss-of-function Aire mutants, including those causing autoimmune polyendocrine syndrome type-1, form foci with increased PML body association. Directing Aire to PML bodies impairs the transcriptional activity of Aire, while dispersing PML bodies with a viral antagonist restores this activity. Our study thus reveals a new regulatory role of PML bodies in Aire function, and highlights the interplay between nuclear aggregate-like assemblies and PML-mediated protein quality control.
Highlights
Aggregate-like biomolecular assemblies are emerging as new conformational states with functionality
We found that purified mouse Aire (mAire) caspase activation recruitment domain (CARD) forms filaments (Fig. 1b), and that the filaments are of non-amyloid type, as evidenced by Congo Red and thioflavin T staining assays (Fig. 1c, d)
We found that mAire aa 1–174, which harbors CARD and the nuclear localization signal (NLS) has very low expression levels, so we fused this to monomeric GFP (Fig. 5a)
Summary
Aggregate-like biomolecular assemblies are emerging as new conformational states with functionality. A transcription factor essential for central T cell tolerance, forms large aggregate-like assemblies visualized as nuclear foci. Our study reveals a new regulatory role of PML bodies in Aire function, and highlights the interplay between nuclear aggregate-like assemblies and PML-mediated protein quality control. Recent chromatin immunoprecipitation experiments suggest that Aire recognizes general epigenetic features and alters chromatin structure including superenhancer and Polycomb-repressed regions, which may lead to indirect regulation of multiple target genes[13,14,15,16] This idea is further supported by the stochastic nature of target gene expression at a single cell level[15,17,18], and the variance of Aire-dependent target genes depending on the cell type[19]. It has been shown that Aire foci formation is correlated with Aire transcriptional activity[4] and that these foci are distinct from other membrane-less nuclear granules characterized to date, such as promyelocytic leukemia protein (PML) bodies[26]
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