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RGS21, a regulator of taste and mucociliary clearance?

ABSTRACT: Motile cilia of airway epithelial cells help to expel harmful inhaled material. Activation of bitterant-responsive G protein-coupled receptors (GPCRs) is believed to potentiate cilia beat frequency and mucociliary clearance. In this study, we investigated whether regulator of G protein signaling-21 (RGS21) has the potential to modulate signaling pathways connected to airway mucociliary clearance, given that RGS proteins modulate GPCR signaling by acting as GTPase-accelerating proteins (GAPs) for the G? subunits of heterotrimeric G proteins.This is a pilot investigation to determine if RGS21, a potential tastant specific RGS gene, is expressed in sinonasal mucosa, and to determine its specific G? substrate using in vitro biochemical assays with purified proteins.Rgs21 expression in sinonasal mucosa was determined using quantitative, real-time PCR and a transgenic mouse expressing RFP from the Rgs21 promoter. Rgs21 was cloned, over-expressed, and purified using multistep protein chromatography. Biochemical and biophysical assays were used to determine if RGS21 could bind and accelerate the hydrolysis of GTP on heterotrimeric G? subunits.Rgs21 was expressed in sinonasal mucosa and lingual epithelium. Purified recombinant protein directly bound and accelerated GTP hydrolysis on G? subunits.Rgs21 is expressed in sinonasal mucosa, is amenable to purification as a recombinant protein, and can bind to G?(i/o/q) subunits. Furthermore, RGS21 can accelerate the hydrolysis rate of GTP on G?i subunits. This provides evidence that RGS21 may be a negative regulator of bitterant responses. Future studies will be needed to determine the physiological role of this protein in mucociliary clearance.

SUBMITTER: Kimple AJ

PROVIDER: S-EPMC3985343 | biostudies-literature | 2014 Mar

REPOSITORIES: biostudies-literature

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RGS21, a regulator of taste and mucociliary clearance?

Kimple Adam J AJ Garland Alaina L AL Cohen Staci P SP Setola Vincent V Willard Francis S FS Zielinski Thomas T Lowery Robert G RG Tarran Robert R Siderovski David P DP

The Laryngoscope 20131002 3

<h4>Objectives/hypothesis</h4>Motile cilia of airway epithelial cells help to expel harmful inhaled material. Activation of bitterant-responsive G protein-coupled receptors (GPCRs) is believed to potentiate cilia beat frequency and mucociliary clearance. In this study, we investigated whether regulator of G protein signaling-21 (RGS21) has the potential to modulate signaling pathways connected to airway mucociliary clearance, given that RGS proteins modulate GPCR signaling by acting as GTPase-ac ...[more]

PMID: 23908053

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Project description:Background: Infants undergoing congenital cardiac surgery with cardiopulmonary bypass are at high risk for respiratory complications. As impaired airway mucociliary clearance (MCC) can potentially contribute to pulmonary morbidity, our study objective was to measure airway clearance in infants undergoing congenital cardiac surgery and examine correlation with clinical covariables that may impair airway clearance function. Materials and Methods: Airway clearance in infants was measured over 30 min using inhaled nebulized Technetium 99m sulfur colloid administered either via a nasal cannula or the endotracheal tube in intubated infants. This was conducted bedside with a portable gamma camera. No difficulty was encountered in positioning the gamma camera over the patient, and neither the camera nor the MCC scan interfered with routine medical care or caused any adverse events. Patient and perioperative variables were examined relative to the MCC measurements. Results: We prospectively enrolled 57 infants undergoing congenital cardiac surgery and conducted a single MCC scan per patient. MCC data from 42 patients were analyzable, including five pre-operative, 15 (40.5%) in the immediate post-operative period (days 1-2), and 22 (59.5%) were later post-operative (≥3 days). Pre-operative MCC was inversely proportional to days requiring post-operative mechanical ventilation (p = 0.006) and non-invasive positive pressure ventilation (p = 0.017). MCC was higher at later post-operative days (p = 0.002) with immediate post-operative MCC being lower (3%; 0-13%) than either pre-operative (21%; 4-25%) (p = 0.091) or later post-operative MCC (18%; 0-29%) (p = 0.054). Among the infants with low post-operative MCC, significantly more were pre-mature [5/19 (26%) vs. 0/18 (0%); p = 0.046], were intubated [14/19 (75%) vs. only 7/18 (39%); p = 0.033] and were receiving higher FiO2 (40%, 27-47% vs. 26%, 21-37%; p = 0.015). Conclusions: This is the first study to show that infants undergoing congenital cardiac surgery have impaired MCC. MCC appeared lowest in the immediate post-operative period. Worse MCC was associated with pre-maturity, mechanical ventilation, or receiving higher FiO2. These findings suggest MCC scans should be further explored for informing clinical decision making to improve post-surgical respiratory outcomes. The possible therapeutic benefit of airway clearance maneuvers for infants with poor MCC function should also be investigated.

Project description:BackgroundAirway tuft cells, formerly called brush cells have long been described only morphologically in human airways. More recent RNAseq studies described a chemosensory cell population, which includes tuft cells, by a distinct gene transcription signature. Yet, until which level in the tracheobronchial tree in native human airway epithelium tuft cells occur and if they function as regulators of innate immunity, e.g., by regulating mucociliary clearance, remained largely elusive.MethodsWe performed immunohistochemistry, RT-PCR and immunoblotting analyses for various tuft cell markers to confirm the presence of this cell type in human tracheal samples. Immunohistochemistry was conducted to study the distribution of tuft cells along the intrapulmonary airways in humans. We assessed the influence of bitter substances and the taste transduction pathway on mucociliary clearance in mouse and human tracheal samples by measuring particle transport speed.ResultsTuft cells identified by the expression of their well-established marker POU class 2 homeobox 3 (POU2F3) were present from the trachea to the bronchioles. We identified choline acetyltransferase in POU2F3 expressing cells as well as the transient receptor potential melastatin 5 (TRPM5) channel in a small population of tracheal epithelial cells with morphological appearance of tuft cells. Application of bitter substances, such as denatonium, led to an increase in mucociliary clearance in human tracheal preparations. This was dependent on activation of the TRPM5 channel and involved cholinergic and nitric oxide signalling, indicating a functional role for human tuft cells in the regulation of mucociliary clearance.ConclusionsWe were able to detect tuft cells in the tracheobronchial tree down to the level of the bronchioles. Moreover, taste transduction and cholinergic signalling occur in the same cells and regulate mucociliary clearance. Thus, tuft cells are potentially involved in the regulation of innate immunity in human airways.

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RGS21, a regulator of taste and mucociliary clearance?

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RGS21, a regulator of taste and mucociliary clearance?

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