Session 3H: LUBAC-Dependent Epithelial Signaling Regulates TRAIL Expression During Influenza A Virus Infection
Session Number
Session 3H: 2nd Presentation
Advisor(s)
Dr. Laura Dada, Northwestern University
Location
Room A117
Start Date
26-4-2018 12:40 PM
End Date
26-4-2018 1:25 PM
Abstract
The linear ubiquitin chain assembly complex (LUBAC), consists of three proteins; HOIP, the catalytic unit whose E3-ligase activity is necessary for the formation of “head-to-tail” linear ubiquitin chains, and SHARPIN and HOIL, which provide stability to the complex (Tokunaga et al. 2009). Downstream of inflammatory stimuli, LUBAC is required for the robust activation of NF-κB and subsequent upregulation of NF-kB-regulated inflammatory cytokines (Tokunaga and Iwai, 2012). During Influenza A Virus (IAV) infection, alveolar epithelial cells (AEC) are targeted for viral replication and orchestrate the inflammatory response by producing cytokines, which recruit inflammatory cells (Short et al. 2014). While infiltrating immune cells are necessary for viral clearance, they contribute to the excessive production of cytokines, which promotes AEC death, damages the alveolar epithelial barrier, and impairs proper lung function (Short et al. 2013). One of the cytokines produced, TNF-related apoptosis-inducing ligand (TRAIL), is expressed by macrophages in response to IAV-induced epithelial signaling (Herold et al. 2008). TRAIL interacts with its receptor, Death Receptor 5 (DR5), on AEC to induce death and enhance morbidity and mortality during influenza infection (Herold et al. 2008). To investigate whether TRAIL expression is driving influenza-induced mortality in mice with an alveolar epithelial specific loss of HOIP (HOIP-KO), IAV-infected wild-type (WT) and HOIP-KO mice were subjected to bronchoalveolar lavage (BAL) at 5 days post infection to obtain a cell pellet of inflammatory cells from which we then isolated RNA using a specialized RNA-binding silica membrane. Eluted purified RNA was then converted into cDNA using a reverse transcriptase and used in an qRT-PCR reaction to quantify TRAIL expression in BAL. We found that the absence of HOIP correlated with increased TRAIL expression, which was detected at nearly sixfold expression levels in cells from HOIP-KO mice as compared to those from WT mice. These results suggest the LUBAC plays a critical role in regulating the total levels of TRAIL expression in the mouse lung during IAV infection.
Session 3H: LUBAC-Dependent Epithelial Signaling Regulates TRAIL Expression During Influenza A Virus Infection
Room A117
The linear ubiquitin chain assembly complex (LUBAC), consists of three proteins; HOIP, the catalytic unit whose E3-ligase activity is necessary for the formation of “head-to-tail” linear ubiquitin chains, and SHARPIN and HOIL, which provide stability to the complex (Tokunaga et al. 2009). Downstream of inflammatory stimuli, LUBAC is required for the robust activation of NF-κB and subsequent upregulation of NF-kB-regulated inflammatory cytokines (Tokunaga and Iwai, 2012). During Influenza A Virus (IAV) infection, alveolar epithelial cells (AEC) are targeted for viral replication and orchestrate the inflammatory response by producing cytokines, which recruit inflammatory cells (Short et al. 2014). While infiltrating immune cells are necessary for viral clearance, they contribute to the excessive production of cytokines, which promotes AEC death, damages the alveolar epithelial barrier, and impairs proper lung function (Short et al. 2013). One of the cytokines produced, TNF-related apoptosis-inducing ligand (TRAIL), is expressed by macrophages in response to IAV-induced epithelial signaling (Herold et al. 2008). TRAIL interacts with its receptor, Death Receptor 5 (DR5), on AEC to induce death and enhance morbidity and mortality during influenza infection (Herold et al. 2008). To investigate whether TRAIL expression is driving influenza-induced mortality in mice with an alveolar epithelial specific loss of HOIP (HOIP-KO), IAV-infected wild-type (WT) and HOIP-KO mice were subjected to bronchoalveolar lavage (BAL) at 5 days post infection to obtain a cell pellet of inflammatory cells from which we then isolated RNA using a specialized RNA-binding silica membrane. Eluted purified RNA was then converted into cDNA using a reverse transcriptase and used in an qRT-PCR reaction to quantify TRAIL expression in BAL. We found that the absence of HOIP correlated with increased TRAIL expression, which was detected at nearly sixfold expression levels in cells from HOIP-KO mice as compared to those from WT mice. These results suggest the LUBAC plays a critical role in regulating the total levels of TRAIL expression in the mouse lung during IAV infection.