Connection established: Missing linker between intracellular pathogen detection and inflammation.
To detect harmful pathogens the immune system is equipped with a variety of receptors, among them the so called “Toll-like receptors”. These receptors received their name as they are homologs to the toll proteins in fruit flies whose discoverer Christiane Nüsslein-Volhard was so excited by them that she yelled “Das ist ja toll!” (German for “that’s amazing!”). But what are these amazing receptors? Humans have 10 of these receptors which sit on cell membranes outside and inside of a variety of cells such as immune cells or cells with barrier functions. Activation of those receptors stimulates different downstream pathways, such as NFkB, MAPK and interferon, to mature the cell, or prepare it to act functionally.
All of these 10 receptors have different specificities for molecules that are common among pathogens. The most prominent of these Toll-like receptors (TLRs) is TLR4 which sits on the cell surface. It recognizes lipopolysaccharides (LPS) which are components of the outer membrane of gram-negative bacteria.
TLR3, 7, 8 and 9 are receptors that sit inside the cells in so called endosomal compartments. The function of these endosomal proteins is to detect molecular patterns of pathogens that are taken up and processed by the host cells. These include double-stranded (ds) and single-stranded (ss) RNA, and CpG-rich ssDNA which are present in viruses and bacteria. It is important to note that human cells also contain RNA and DNA species which can trigger TLR signaling, however, due to the specific location of intracellular TLRs inside the endosomal compartment this is normally not the case. Under certain conditions such as cell death, host RNA and DNA can appear outside of cells, enter the endosomal compartment through endocytosis and trigger TLR signaling. Many other mechanisms are in check to prevent the recognition, but in some genetically prone individuals this might result in TLR activation and subsequent autoimmunity.
Many genes have been associated with a higher risk to develop autoimmune diseases, however, for most of them the underlying mechanisms (the exact molecular cause(s)) are not fully understood. One such gene is SLC15A4, which has been reported to play an important role in interferon production after TLR7 stimulation and subsequent autoantibody production. This appeared to be caused through the amino acid transporter function of this protein.
In a recent paper, a group of researchers led by Giulio Superti-Furga located in Vienna, Austria further investigated the role of SLC15A4 in TLR signaling. The author used a tandem-affinity protein purification system to identify potential interactors of SLC15A4. This system uses ectopic, or artificial, expression of a protein of interest that is fused to two different protein tags allowing co-purification with specific interactors which can subsequently be detected by mass spectrometry. With this approach they repeatedly identified the interactor CXorf21, which has also been associated with autoimmunity and autoinflammation. This interaction which the authors confirmed in varying cell models seem to play an important role downstream of TLR7 and TLR8. If CXorf21 or SLC15A4-deficient cells were stimulated with the ssRNA mimetic R848 they showed a severe reduction of cytokine and chemokine expression compared to control cells. Similar results were seen if TLR9 was stimulated suggesting that the presence of these two molecules is a general requirement for proper endosomal TLR function. Strikingly, the production of these molecules was unaltered if other, plasma-membrane-localized TLRs were stimulated. In further experiments the authors then showed that specifically the activation of the interferon pathway was impaired as expression of genes associated with this pathway were altered in knockout cells. Furthermore, the phosphorylation of IRF5, a member of this pathway was altered whereas other pathways such as NFkB and MAPK where unaltered. To understand the molecular requirements of the interaction of these two proteins for TLR signaling, the authors did a series of experiments in which they reconstituted the absence of either protein with varying mutants of the original proteins which let them to identify regions in SLC15A4 and CXorf21 that are important for their interaction and signaling. They identified a conserved motif in the C-terminus of CXorf21 which is homolog to a region in IRF5. This region is important for CXorf21 to act as an adaptor protein for proper activation of IRF5 downstream of endosomal TLRs. Consequently, the authors renamed the protein CXorf21 as ‘TLR adaptor interacting with SLC15A4 on the lysosome’ (TASL).
The work of the Superti-Furga lab identifies a critical linker between TLR and IRF5 through SLC15A4 and TASL. TASL seems to show similarities with other adaptor proteins such as TRIF and MAVS in a way that it shares a motif for dimerization. This is of particular interest as this domain can be potentially used as a target for drug intervention. Thus, we might be able in the future to treat autoimmune diseases associated with drugs that specifically dampen the overreacting immune response towards self-nucleic acids.
By Nina Serwas
This article:
Further reading:
Gay, N. J., Symmons, M. F., Gangloff, M. & Bryant, C. E. Assembly and localization of Toll-like receptor signalling complexes. Nat. Rev. Immunol. 14, 546–558 (2014).
Kobayashi, T. et al. The histidine transporter SLC15A4 coordinates mTOR-dependent inflammatory responses and pathogenic antibody production. Immunity 41, 375–388 (2014).
Pelka, K., Shibata, T., Miyake, K. & Latz, E. Nucleic acid-sensing TLRs and autoimmunity: Novel insights from structural and cell biology. Immunol. Rev. 269, 60–75 (2016).
Ve, T., Gay, N. J., Mansell, A., Kobe, B. & Kellie, S. Adaptors in Toll-Like Receptor Signaling and Their Potential as Therapeutic Targets. Curr Drug Targets 13, 1360–1374 (2012).
Schematic of SLC15A4- and TASL-dependent IRF5 activation by TLR7, TLR8 and TLR9 from the primary paper, Heinz et al.