R01AI155426

Regulation of TLR Signaling, Inflammation, and Antigen Presentation by VPS33B - Project Summary

Initiation of innate immune responses depends on cognate interaction between germline-encoded pattern recognition receptors (PRRs) and their ligands expressed by microbes. Following recognition, the receptors activate downstream signal transduction pathways that often involve recruitment of downstream adapters and kinases. The Toll-like receptor (TLR) family of PRRs, which are the subject of current investigation, are expressed both on the plasma membrane and in the endosomes.

Several recent studies have demonstrated that endocytosis of the plasma membrane TLRs, especially TLR4, plays a critical role in regulating both the quality and magnitude of inflammatory responses in responding macrophages. Other studies have shown that TLR signaling enhances phagocytosis of microbial cargo but not apoptotic cell cargo, suggesting a degree of specificity that is not yet understood. Additionally, while endocytosis of TLR4 and the events following endocytosis that influence signal transduction are well studied, it is not entirely clear if and how endocytosis influences signaling downstream of other plasma membrane and endosomal TLRs.

In our studies, we have found that a protein called VPS33B regulates the handling of phagocytic and endocytic cargo following pattern recognition receptor activation. More importantly, VPS33B directly influences the outcome of signaling downstream of TLRs in mice and Toll- and IMD pathways in Drosophila. Mutations in the genes VPS33B and VPS16B are linked to a rare human disease called ARC (Arthrogryposis-Renal Dysfunction-Cholestasis) syndrome. Both of these ARC genes encode paralogs of HOPS complex subunits, suggesting a role in membrane fusions. However, how perturbation of the function of these proteins results in a diverse spectrum of disease symptoms in ARC patients is not entirely clear. It has been documented that ARC patients suffer from sepsis and recurrent bacterial infections, so we investigated the role of these proteins in influencing immune responses.

We found that in the absence of VPS33B, Drosophila respond vigorously to microbial insult. Exaggerated immune responses are generated in response to live or dead bacteria and purified ligands of the Toll and IMD pathways result in death of VPS33B mutant flies, but not wild-type flies. This function of VPS33B is conserved in vertebrates, and we find that mouse macrophages lacking VPS33B secrete very high quantities of inflammatory cytokines when stimulated by either plasma membrane or endosomal TLR ligands. Therefore, we hypothesize that activation of pattern recognition receptors, specifically TLRs, leads to the formation of specialized endosomes that depend on VPS33B for lysosomal fusion. The lack of VPS33B is likely to affect several aspects of innate and adaptive immunity.

To test this hypothesis, we propose to:

1. Define the molecular events that regulate VPS33B function in endosomal maturation.
2. Define the role of VPS33B-regulated TLR trafficking and signaling.
3. Investigate the role of VPS33B in regulating cargo handling by dendritic cells.
4. Investigate the role of VPS33B in regulating antigen presentation and adaptive immunity.

Childrens Hospital Medical Center

TO ASSIST PUBLIC AND PRIVATE NONPROFIT INSTITUTIONS AND INDIVIDUALS TO ESTABLISH, EXPAND AND IMPROVE BIOMEDICAL RESEARCH AND RESEARCH TRAINING IN INFECTIOUS DISEASES AND RELATED AREAS, TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS. TO ASSIST PUBLIC, PRIVATE AND COMMERCIAL INSTITUTIONS TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS, TO PROVIDE RESEARCH SERVICES AS REQUIRED BY THE AGENCY FOR PROGRAMS IN INFECTIOUS DISEASES, AND CONTROLLING DISEASE CAUSED BY INFECTIOUS OR PARASITIC AGENTS, ALLERGIC AND IMMUNOLOGIC DISEASES AND RELATED AREAS. PROJECTS RANGE FROM STUDIES OF MICROBIAL PHYSIOLOGY AND ANTIGENIC STRUCTURE TO COLLABORATIVE TRIALS OF EXPERIMENTAL DRUGS AND VACCINES, MECHANISMS OF RESISTANCE TO ANTIBIOTICS AS WELL AS RESEARCH DEALING WITH EPIDEMIOLOGICAL OBSERVATIONS IN HOSPITALIZED PATIENTS OR COMMUNITY POPULATIONS AND PROGRESS IN ALLERGIC AND IMMUNOLOGIC DISEASES. BECAUSE OF THIS DUAL FOCUS, THE PROGRAM ENCOMPASSES BOTH BASIC RESEARCH AND CLINICAL RESEARCH. SMALL BUSINESS INNOVATION RESEARCH (SBIR) PROGRAM EXPANDS AND IMPROVES PRIVATE SECTOR PARTICIPATION IN BIOMEDICAL RESEARCH. THE SBIR PROGRAM INTENDS TO INCREASE AND FACILITATE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, TO INCREASE SMALL BUSINESS PARTICIPATION IN FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. THE SMALL BUSINESS TECHNOLOGY TRANSFER (STTR) PROGRAM STIMULATES AND FOSTERS SCIENTIFIC AND TECHNOLOGICAL INNOVATION THROUGH COOPERATIVE RESEARCH AND DEVELOPMENT CARRIED OUT BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO FOSTER TECHNOLOGY TRANSFER BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO INCREASE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. RESEARCH CAREER DEVELOPMENT AWARDS SUPPORT THE DEVELOPMENT OF SCIENTISTS DURING THE FORMATIVE STAGES OF THEIR CAREERS. INDIVIDUAL NATIONAL RESEARCH SERVICE AWARDS (NRSAS) ARE MADE DIRECTLY TO APPROVE APPLICANTS FOR RESEARCH TRAINING IN SPECIFIED BIOMEDICAL SHORTAGE AREAS. IN ADDITION, INSTITUTIONAL NATIONAL RESEARCH SERVICE AWARDS ARE MADE TO ENABLE INSTITUTIONS TO SELECT AND MAKE AWARDS TO INDIVIDUALS TO RECEIVE TRAINING UNDER THE AEGIS OF THEIR INSTITUTIONAL PROGRAM.

93.855 - Allergy and Infectious Diseases Research

National Institute of Allergy and Infectious Diseases (NIAID) [HHS - NIH]

Cincinnati, Ohio 45229 United States

Single Zip Code

NIH Research Project Grant (Parent R01 Clinical Trial Not Allowed) (PA-20-185)

Amendment Since initial award the total obligations have increased 463% from $605,972 to $3,409,595.