NK cells are a subpopulation of lymphocytes whose unique receptors facilitate the detection of infected,transformed, or `stressed' cells. This immune recognition subsequently leads to the development of NK cell-mediated cytotoxicity or the generation of cytokines and chemokines that activate other components of theimmune system. Patients with hyper-IgE syndrome (HIES) have immune dysregulation and can be affected byrecalcitrant cutaneous herpes virus and papillomavirus infections. A major genetic abnormality found in thesepatients is deletion or loss-of-function mutations in the gene encoding Dedicator of Cytokinesis 8 (DOCK8), aguanine nucleotide exchange factor (GEF) for Cdc42. The high incidence of recurrent cutaneous viralinfections in DOCK8-deficient patients is suggestive of defects in natural killer (NK) cell function. In fact, othersand we have recently shown that NK cells deficient in DOCK8 have reduced lytic function, decreasedadhesion, F-actin accumulation at the cytotoxic synapse and an inability to polarize lytic granules toward thetarget cell. However, how DOCK8 can regulate so many critical steps in the development of NK cell killing isnot known. It is our central hypothesis that the DOCK8-interactome coordinates the regulation of the actinand microtubule cytoskeletons to facilitate NK cell polarization and effect NK cellular cytotoxicity.Based on the preliminary data included in this proposal, we hypothesize that: (a) DOCK8 activation of CDC42is critical to the development of NK cell killing; (b) DOCK8 interaction with WASP is critical for its localizationand F-actin generation at the NKIS; (c) talin recruitment by DOCK8 mediates NK cell ? target adhesion; (d)HkRP3, a hematopoietically expressed protein that interacts with DOCK8 is involved in MTOC polarization andlytic granule clustering and affects lytic granule clustering through its interactions with tubulin and the dyneinmotor complex; (e) septins are DOCK8 interacting proteins that regulate NK cell killing; (f) BORG stabilizationof septin filaments is impaired by active Cdc42 in order to regulate NK cell killing. In order to test thesehypotheses we will: (1) Determine the mechanism by which DOCK8 regulates F-actin dynamics and celladhesion; (2) Define the mechanism by which HkRP3 regulates lytic granule convergence and MTOCpolarization; (3) Determine the role of septins in the regulation of the NK cell cytotoxicity. The outcome of theproposed experiments will provide an experimental basis for understanding the molecular events that areinvolved in the regulation of NK cell effector functions, and will, in a broader context, advance ourunderstanding of fundamental processes in cellular activation leading to F-actin regulation, MTOC polarizationand granule exocytosis. Moreover, the information obtained through these studies will likely instruct howDOCK8 is functioning in other immune cells.
|Effective start/end date||5/17/16 → 4/30/21|
- National Institutes of Health: $397,500.00