MECHANISMS OF PIN1 REGULATION OF IRAK-M STABILITY IN TLR/IL-1R SIGNALING AND STRUCTURE DETERMINATION OF IRAK-M DEATH DOMAIN
Asthma is a chronic inflammatory sickness of the airways caused by environmental and genetic factors. The mechanisms of activation of the innate immunity signaling pathways that cause asthma are not fully understood yet. This study investigated the novel targets for the development of new asthma therapeutics. Innate immunity provides the first line of defense against bacterial and viral pathogens. Toll-like receptors (TLRs) act as sentinels to detect specific pathogen-associated molecular patterns (PAMPs). TLR-initiated signaling cascades trigger inflammatory, allergic and non-allergic responses via NF-kappaB signaling. Loss of proper innate immunity regulation leads to inflammatory diseases such as asthma. IRAK-M is a known negative regulator of TLR signaling and is known to assemble into the activated TLR signaling complex to attenuate downstream signaling. Prolyl peptide bonds, such as in the phosphorylated Ser/Thr-Pro (pS/T-P) motifs recognized by Pin1, can exist in two distinct isomer conformations, cis and trans, that exchange on a slow time scale (exchange time constant of several minutes). Pin1 can accelerate the isomerization rate of pS/T-P motifs by orders of magnitude. The NMR experiments reported here show that Pin1 directly interacts with and isomerizes the phosphoS110-P111 peptide bond in a phosphopeptide corresponding to the IRAK-M sequence 103-124 (pIRAK-M), and also acts on the corresponding peptide harboring the phosphomimetic mutation S110E. Assembly of IRAK-M into the TLR signaling complex is mediated by its N-terminal death domain. Oligomerization is a hallmark of Death Domains (DD). The IRAKM-DD has eluded structure determination due to aggregation. Here, we report the 1H, 13C and 15N backbone and side chain resonance assignments for a double-mutant IRAK-M Death Domain (R56D, Y61E) that is a highly soluble monomer well suited for NMR studies. Furthermore, we solved IRAK-M Death Domain structure and simulated docking prediction.
Death Domain; IRAK-M; NMR Structure; Biophysics; NMR; Biology; Asthma
Nicholson, Linda K.
Oswald, Robert Edward; Mao, Yuxin
Ph. D., Biophysics
Doctor of Philosophy
dissertation or thesis