Molecular Characterization of Novel Mechanisms for Interleukin 17 and its Essential Signaling Mediator Act1 in Airway Epithelial Cells.

摘要

The role of the proinflammatory cytokine interleukin-17 (IL-17) in the airway has been under investigation for the last 12 years. Many studies have been published demonstrating its pleiotropic role in the production of chemokines, cytokines, mucins, and antimicrobial proteins. Many in vivo models have demonstrated the association of IL-17 in airway inflammation in response to allergens as well as environmental insults, as well as chronic inflammatory lung diseases such as cystic fibrosis, chronic obstructive pulmonary disease and asthma.;However, relatively little is known regarding the downstream signaling mechanisms by which IL-17 mediates its many effects. What is known is that the signaling is complex; IL-17 has been shown to involve multiple signaling pathways, including: the canonical NF-kappaB pathway, multiple mitogen-activated protein (MAP) kinase pathways, as well as C/EBP, PI-3 kinase and JAK. Both cell type and target gene appear to determine the signaling pathway involved downstream of IL-17. Little is known regarding the protein-protein interactions that mediate these signaling events. What is known is that the intracellular protein Act1 (also known as CIKS) is an important downstream mediator of IL-17 induced signaling. Cells derived from Act1-deficient mice are largely unresponsive to IL-17A stimulation. Act1 has been shown to contain both TRAF6 and IL-17 receptor binding sites, and therefore acts as an intermediate to connect the activated IL-17 receptor complex to pathways downstream of TRAF6. However, Act1 has additional functions as well. It can also bind to the IL-25, CD40 and BAFF-R receptors, and has recently been shown to act as a U-box type E3 ubiquitin ligase. Recently, a single nucleotide polymorphism (SNP) that encodes a loss-of-function mutation in the gene for Act1, TRAF3IP2, was associated with psoriasis, an autoimmune inflammatory skin disease, and psoriatic arthritis. As IL-17 is associated with the pathogenesis of psoriasis, this is counter-intuitive to the known functions of Act1, indicating that Act1 may have other functions as well.;This dissertation details a novel nuclear function for Act1 as a transcriptional enhancer. Subsequent RNA-seq comparison of cells ectopically expressing Act1 and IL-17A stimulated cells identified a number of cornified envelope constituents whose expression is up-regulated by both Act1 and IL-17. The cornified envelope is a structure formed in the outermost layer of stratified squamous epithelia. Finally, we detail the use of a yeast two hybrid assay to identify novel Act1 interacting proteins, and further characterize the interaction of Act1 with one of these proteins, COMMD1 (copper m etabolism (Murr1) domain containing 1), which might represent a new target of Act1's ubiquitin ligase activity.