The Speckled Protein (SP) Family: Immunity's Chromatin Readers

Abstract

Chromatin 'readers' are central interpreters of the epigenome that facilitate cell-specific transcriptional programs and are therapeutic targets in cancer and inflammation. The Speckled Protein (SP) family of chromatin 'readers' in humans consists of SP100, SP110, SP140, and SP140L. SPs possess functional domains (SAND, PHD, bromodomain) that dock to DNA or post-translationally modified histones and a caspase activation and recruitment domain (CARD) to promote multimerization. Mutations within immune expressed SPs associate with numerous immunological diseases including Crohn's disease, multiple sclerosis, chronic lymphocytic leukemia, veno-occlusive disease with immunodeficiency, as well as Mycobacterium tuberculosis infection, underscoring their importance in immune regulation. In this review, we posit that SPs are central chromatin regulators of gene silencing that establish immune cell identity and function.

Keywords: Aire; B cells; CLL; Crohn’s disease; Mycobacterium tuberculosis; PML nuclear bodies; SP100; SP110; SP140; SP140L; VODI; chromatin; epigenetics; macrophages; multiple sclerosis.

Figure 1.. Functional Domains and Disease-Associated Mutations in Human Speckled Proteins (SPs) and Autoimmune Regulator (Aire)

SPs and Aire express highly similar caspase activation and recruitment domains (CARDs), Sp100, Aire, NucP41/75, DEAF-1 shared domains (SANDs), and plant homeodomains (PHDs). The SP family PHD is similar to Aire’s PHD1. Aire contains two unique PHDs and lacks a bromodomain (BRD). Highlighted are representative mutations associated with immunological diseases. Depicted are full-length SP isoforms: SP100 isoform C (SP100-C), SP110 isoform C (SP110-C), SP140 isoform A (SP140-A), and SP140-like (SP140L) isoform A (SP140L-A). Protein domains are depicted with corresponding amino acid residue positions underneath. The synonymous mutation depicted in SP140 is a cytidine-to-thymidine nucleotide mutation. Δ1, one-nucleotide deletion; +7, seven-nucleotide insertion. This figure was created using BioRender (https://biorender.com/).

Figure 2.. Genomic Organization of Speckled Protein (SP) Genes and Evolution of SP140.

(A) Schematic of SP genes on human chromosome 2 and Sp genes on mouse chromosome 1. (B) Here, we use SP140 as a model protein of the SP family to show the high degree of evolution in these proteins and the importance of studying this family in both mouse and human. Mouse Sp140, human SP140, rhesus monkey SP140, and zebrafish SP140 amino acid sequences were aligned using ClustalW, which determined that there was 34.7% pairwise identity between mouse and human at the amino acid level. Sequence similarity was determined using the Blosum62 score matrix and a threshold of 1. A schematic of human SP140 with functional domains is shown above the alignment. The intrinsically disordered region (IDR) is indicated on the human schematic but is mostly absent in the mouse. Abbreviations: BRD, bromodomain; CARD, caspase activation and recruitment domain; PHD, plant homeodomain; SAND, Sp100, Aire, NucP41/75, DEAF-1 shared domain.

Figure 3.. Outline of All Known Human Isoforms of Speckled Proteins (SPs).

Functional domains are color coded and indicated in the key: CARD, caspase activation and recruitment domain; SAND, Sp100, Aire, NucP41/75, DEAF-1 shared domain; PHD, plant homeodomain; BRD, bromodomain; HMG, high-mobility group protein domain. SP110 isoform A (SP110-A) contains a truncated BRD. In the SP140 alignment, most isoforms differ by exon usage in the intrinsically disordered region (IDR). Numbering above these IDR exons corresponds to exon numbering in full-length human SP140 isoform A (SP140-A). SP140L, SP140-like.

Figure 4.. Expression of Speckled Proteins (SPs) in Murine Immune Cell Subsets.

Sp100, Sp110, and Sp140 expression in indicated mouse leukocytes based on RNA-seq data (Immgen.org). Abbreviations; DC, dendritic cell; GN, granulocyte; HSC, hematopoietic stem cell; ILCs, innate lymphoid cells; Mφ, macrophage; Mo, monocyte.

Figure 5.. Mechanism of Action to date for Speckled Protein 140 (SP140) for Immune Cell Identity and Function.

Lineage-defining transcription factors (TFs) for mature macrophages (such as PU.1) drive SP140 transcription. SP140 in turn docks to repressed chromatin marked by H3K27me3 and maintains the silencing of lineage-inappropriate genes, such as HOX, OLIG, and FOX, for the preservation of cell identity and function.

Figure 6.. Alignments of Functional Domains of Human Speckled Protein (SP) Isoforms and Autoimmune Regulator (Aire).

Sequences were aligned in Geneious using ClustalW. Red shading of residue and bar graphs above sequences corresponds to percentage identity at each residue. Stars indicate conserved residues that are mutated in Aire in patients with autoimmune polyendocrine syndrome type 1 (APS-1). In (B), a black box indicates the KDWK-like DNA-binding motif [11]. In (D), the arrow points to the position where SPs lack a conserved asparagine residue previously reported to anchor bromodomains to histone tail acetyl groups via hydrogen bonds [13]. Abbreviations: BRD, bromodomain; CARD, caspase activation and recruitment domain; PHD, plant homeodomain; SAND, Sp100, Aire, NucP41/75, DEAF-1 shared domain; SP140L, SP140-like protein; SP100-C, SP100 isoform C.