Projects / Programmes
Cell signalling of Toll-like receptors
| Code |
Science |
Field |
Subfield |
| 1.05.00 |
Natural sciences and mathematics |
Biochemistry and molecular biology |
|
| Code |
Science |
Field |
| P004 |
Natural sciences and mathematics |
Biochemistry, Metabolism |
| Code |
Science |
Field |
| 3.04 |
Medical and Health Sciences |
Medical biotechnology |
innate immune response, cell signaling, Toll-like receptors, signaling adapter, TIR domain, MyD88, inflammation, infection, B-cell lymphom, protein structure, signaling lipids, phosphatidic acid
Researchers (16)
Organisations (3)
Abstract
Toll-like receptors (TLRs) represent a family of innate immune receptors that recognize molecular patters characteristic either for the invading microorganisms or for tissue damage. Activation of TLRs leads to inflammation, which occurs in infection, autoimmune diseases, cancer and in many other diseases. Activation of the TLR signaling is triggered by agonist-induced receptor dimerization and is mediated in all but one TLRs by the adapter molecule MyD88. MyD88 comprises a structured death domain (DD) at the N-terminus and C-terminal Toll/interleukin-1 receptor (TIR) as well as an unstructured intermediary (INT) segment between DD and TIR and an N-terminal segment. DD and TIR domain mediate homotypic interaction with DD of IRAK signaling kinases and TIR domains of TLRs, respectively. However the role of unstructured subdomains of MyD88 is unclear. We propose to investigate the biochemical background and functional consequences of the previously unknown properties of MyD88 and its subdomains in four tasks:
1. Based on our initial results we propose that the INT segment of MyD88 binds to the DD of IRAK4 downstream signaling kinase and that the peptide segment from this domain can inhibit cell activation mediated by MyD88. Confirmation of the cellular target of INT will help us elucidate the molecular mechanism of TLR signaling. We will determine the specificity of inhibition of different TLRs and IL-1R and investigate the therapeutic potential of cell permeabilizing INT peptide for the inhibition of inflammation in the endotoxaemic animal model of sepsis.
2. We propose that the N-terminal unstructured segment of MyD88 mediates binding of MyD88 to membrane microdomains enriched with phosphatidic acid (PA), which is a signaling lipid produced by several biosynthetic pathways. Our initial results show that MyD88 and its N-terminal peptide specifically bind PA. We plan to investigate the effect of decreasing or increasing the level of PA on cell signaling though TLRs by up- or down-regulating activity of enzymes involved in PA conversion. We propose that PA participates in a positive feedback loop to sensitize TLR signaling. Confirmation of the role of this signaling lipid for activation of the innate immune system will have a significant impact on the understanding of signaling crosstalk in immune response.
3. Recently point mutations within the TIR domain of MyD88 have been described in B-cell lymphoma, which require constitutive MyD88 activity for the survival of cancerous cells. We propose that those oncogenic mutations, (e.g. L265P) disrupt the fold of a TIR domain of MyD88, leading to its constitutive activity. This suggests that a previously unknown function of the TIR domain is to prevent the constitutive activity of MyD88. We will investigate the effect of those mutations on MyD88 and isolated TIR domain using biochemical, biophysical and cell activation experiments. We will design and test the phenotype of new mutations within the core of TIR domain. We expect that results will be able to explain the high frequency of those mutations in cancer. We will additionally investigate the potentials for therapeutic treatment of cancer with constitutively active MyD88 using INT peptide (prepared in task 1).
4. Several pathogenic bacteria produce immunosuppressing proteins that contain TIR domain homology (TcpB)1 with an N-terminal extension containing a coiled-coil motif. We propose that the physiological function of this N-terminal extension is to augment the inhibition of TLRs. We will investigate the specificity of inhibition of an artificially dimerized TIR domain of MyD88. This will allow us to explain the evolutionary advantage for bacteria to adopt the dimerization motif and provide additional structural information on the molecular mechanism of MyD88 interaction with TLRs. Strong inhibitors of MyD88 signaling are also potentially useful for the inhibition of excessive inflammat
Significance for science
Within the project we made significant advances towards understanding the molecular mechanism of signaling through the adapter protein MyD88, which is a central mediator of TLR signaling and IL1R. We have identified a new, so far unknown role of specific sub-domains of the protein MyD88, such as intermediate INT domain and the N-terminal end of the protein prior to DD. We found that the dimerization of the TIR domain is a key step in the activation of which enables the dimerization to trigger activation or coupled by coiled-coil induced dimerization to cause greater inhibition, which is interesting for the development of new therapeutic strategies. We explained the molecular mechanism of activation of the mutants in the TIR domain of MyD88, which are associated with B-cell lymphoma, and physiological effects, resulting from these results.
Significance for the country
The results of the project are important for Slovenia, since the results are relevant to the health, since the innate immunity is important for infectious diseases as well as for other inflammatory diseases and cancer. Research in this area allows the contact with the world class research in the field of immunology, which is particularly important for health. The important role of the group members for training qualified researchers who are able to solve complex scientific and developmental problems. Success in this field of science and intense international involvement the promotion of Slovenia as the scientifically developed country and for the promotion of science as a key factor in the successful development of Slovenia.
Most important scientific results
Annual report
2011,
2012,
2013,
final report,
complete report on dLib.si
Most important socioeconomically and culturally relevant results
Annual report
2011,
2012,
2013,
final report,
complete report on dLib.si
