Small‐Interfering RNA‐Mediated Identification and Regulation of the Ternary SNARE Complex Mediating RBL‐2H3 Mast Cell Degranulation

Abstract

Dysregulation of mast cell function contributes to allergic and autoimmune disease that affects more than 70 million persons in the United States alone. Identifying novel mast cell targets that mediate disease or disease progression is required for the development of innovative therapeutics for the treatment of allergy/asthma and autoimmune disease. RNA interference technologies offer hope both as basic research tools for target identification and as potential, novel, specific therapeutics. Soluble N‐ethylmaleimide‐sensitive factor attachment protein receptors (SNAREs) are a family of evolutionarily conserved proteins that have been postulated to mediate the transport and fusion of inflammatory mediator‐laden vesicles to the membrane in mast cells leading to their subsequent exocytosis. The functional role(s) of specific SNARE family member complexes in mast cell degranulation has not been fully elucidated. Here, we characterize the functional importance of SNARE complexes in FcεRI receptor‐mediated degranulation of RBL‐2H3 cells utilizing RNA interference. We demonstrate that ternary SNARE complexes of synaptosomal‐associated protein‐23, Syntaxin 4 and vesicle‐associated membrane protein‐7 (VAMP‐7) or VAMP‐8 are directly involved in mast cell degranulation. Additionally, we evaluate the siRNAs directed against these molecules as potential therapeutic agents for disease intervention. These studies have identified specific SNARE proteins and complexes that serve as novel targets for the development of siRNA therapies to treat allergic and autoimmune disease.

SNAP-25 Contains Non-Acylated Thiol Pairs that can Form Intrachain Disulfide Bonds: Possible Sites for Redox Modulation of Neurotransmission

“…The SNAP-25 family consists of multiple isoforms, including SNAP-25a, SNAP-25b, and SNAP-23. It is noteworthy in this regard that a recent study reported that the specific SNAP-25a/b antibody used here did not detect the abundant SNAP-23 in mast cells (Woska and Gillespie 2011). This finding demonstrates that this antibody exhibits no or very weak cross-reactivity with SNAP-23 and argues that the ''SNAP-25'' detected in our experiments was a composite of the neuron-specific SNAP-25a and SNAP25b isoforms only.…”

Intracellular trafficking and secretion of inflammatory cytokines

“…VAMP8 can control various stages of granule-mediated secretion. Mast cells undergo piecemeal degranulation, which is regulated through the VAMP8 and VAMP7 as R-SNAREs located on secretory granules and Stx4/SNAP23 on the cell surface [92,96,[98][99][100]. In pancreatic acinar cells VAMP8 regulates granule-to-granule, but not granule-to-surface, fusion during compound exocytosis, whether this is the case for mast cell is unclear [101].…”

CD80 and CD86 knockdown in dendritic cells regulates Th1/Th2 cytokine production in asthmatic mice

“…The ability of RNAi to inhibit virulent gene expression has been widely used to treat a variety of diseases (33)(34)(35). Also, a number of studies have reported that RNAi can be used in DCs to diagnose and treat bronchial asthma (36)(37)(38)(39). Darcan-Nicolaisen et al (40) discovered that the two major signs of allergic asthma in the OVA-induced asthma mouse model, which are airway inflammation and hyperactivity, were significantly ameliorated by signal transducer and activator of transcription 6 (STAT6) silencing in airway epithelial cells using the administration of siRNA nose drops.…”