Role of Surfactant Protein A and its Receptor SP-R210 in Influenza Responses in Macrophages
Restricted (Penn State Only)
- Author:
- Yau, Eric
- Graduate Program:
- Biomedical Sciences (PHD)
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- February 23, 2022
- Committee Members:
- Jong Yun, Outside Unit & Field Member
Nicholas Buchkovich, Major Field Member
John Wills, Major Field Member
Zissis Chroneos, Chair & Dissertation Advisor
Ralph Keil, Program Head/Chair - Keywords:
- Surfactant Protein
SP-A
Macrophages
Alveolar Macrophages
Influenza
IAV
Trafficking
Pulmonary Immunity
Pulmonary Inflammation
Surfactant Protein
SP-A
Alveolar Macrophage
Macrophage
AM
Influenza
Influenza A
IAV
Pulmonary Immunity
Pulmonary Inflammation
Trafficking
Virus Trafficking
IAV Trafficking
IAV Endocytosis
Surfactant Protein A - Abstract:
- Influenza A virus (IAV) is a yearly public health concern due to the recurrent respiratory tract infection it causes in healthy and immunocompromised individuals. IAV infections typically cause self-limiting infections in the upper respiratory tract that are cleared with the help of mucus trapping of the pathogen, subsequent mucociliary clearance, as well as infiltrating immune cells like macrophages, neutrophils, and dendritic cells. However, IAV can also cause severe infections in the lower respiratory tract, especially in individuals with compromised immune systems. In the lower respiratory tract, proper balance is required between inflammatory responses to clear the pathogen and the subsequent anti-inflammatory process to initiated repair and limit immunopathology. Oftentimes, severe influenza infections can result in pneumonia, acute respiratory distress, or subsequent infections such as SARS-CoV2 and Streptococcus pneumoniae. This balance is maintained with the help of alveolar macrophages, the resident innate immune cell in the lung, and surfactant proteins, specifically surfactant protein A (SP-A). SP-A is a member of the collectin family, and both it and its receptor, SP-R210, have been shown to modulate macrophage phenotype and functions critical to the maintenance of immune homeostasis, life-long immunosurveillance of the airway, and precise regulation of innate host responses to airborne infection. The overarching concern of the present work is understand how the SP-A/SP-R210 pathway can affect acute and chronic inflammatory responses of macrophages to pathogens in the lung. Better understanding of the local-host pathogen interactions can identify novel therapeutic targets to arrest development of inflammatory sequelae and limit pathogen replication during such respiratory infections. The present work focused on understanding extrinsic and intrinsic interactions of IAV with SP-A and SP-R210 in macrophages. Studies in chapter 2 sought to define the direct interaction of SP-A with influenza A virus hemagglutinin (HA). Previous studies report that SP-A presents decoy glycoconjugates for binding of HA in Ca++-dependent manner, infection of host cells. This current work, however, determined that SP-A and HA interact through novel protein-protein interactions independent of glycosylation of either protein and Ca++, and that this interaction reduces replication of the viral genome in macrophages. Experiments also revealed that prior exposure of macrophages to SP-A, similar to the pulmonary environment in vivo, inhibited viral replication. This led to studies to closely examine the effect of SP-A and SP-R210 deficiency on viral trafficking. Experiments in chapter 2 revealed that SP-A delays endosomal exit of the virus genome to nucleus. Studies in chapter 3 show that blockade of SP-R210 using antibodies recognizing the both isoforms, SP-R210L and SP-R210S, suppresses replication of the virus genome, suggesting a role for the receptor in IAV infection. To investigate the role of each SP-R210 isoform, studies revealed that depletion of the SP-R210L isoform renders macrophages highly resistant to infection, while depletion of both isoforms delayed viral genome replication. Furthermore, this was associated with delayed viral acidification in the context of altered SP-R210 isoform expression; this was interestingly in contrast to faster endosomal acidification, suggesting an alternative trafficking mechanism for IAV in macrophages lacking SP-R210 isoforms. Significantly, endosomal arrest and activation of the antiviral response was traced to recruitment of interferon inducible transmembrane protein 3 (IFITM3) in SP-R210L-deficient cells. IFITM3 is a known locus of genetic polymorphism and susceptibility to viral infection in humans. These findings led to studies in Chapter 4 to further understand how differential expression of SP-R210 isoforms affect macrophage function. These experiments revealed discrete upregulation of inflammatory and antiviral pathways such as TLR signaling, RIG-I signaling, and NLR-signaling with depletion SP-R210L, while cells lacking SP-R210 completely only showed upregulation of RIG-I and NLR signaling. Furthermore, these changes were reflected at an epigenetic level with alterations in histone H3 methylation and binding of the pioneer transcription factor PU.1. These findings support the idea that the shape and severity of the inflammatory response depends on differential expression of SP-R210 isoforms in resident and exudative macrophages, with SP-R210 and SP-A altering early IAV uptake and trafficking events in macrophages. This altered viral uptake and processing by macrophages may lead to altered antigen processing and presentation, affecting activation of the adaptive immune response. Dysregulation of the SP-A/SP-R210 pathway in macrophages may lead to excessive inflammation that perturbs the capacity of the lung’s immune system to restore homeostasis following clearance of IAV infection.