Dataset Information

Unrestrained Gα_i2 Signaling Disrupts Neutrophil Trafficking, Aging, and Clearance.

ABSTRACT: Neutrophil trafficking, homeostatic and pathogen elicited, depends upon chemoattractant receptors triggering heterotrimeric G-protein Gα_iβγ signaling, whose magnitude and kinetics are governed by RGS protein/Gα_i interactions. RGS proteins typically limit Gα_i signaling by reducing the duration that Gα_i subunits remain GTP bound and able to activate downstream effectors. Yet how in totality RGS proteins shape neutrophil chemoattractant receptor activated responses remains unclear. Here, we show that C57Bl/6 mouse neutrophils containing a genomic knock-in of a mutation that disables all RGS protein-Gα_i2 interactions (G184S) cannot properly balance chemoattractant receptor signaling, nor appropriately respond to inflammatory insults. Mutant neutrophils accumulate in mouse bone marrow, spleen, lung, and liver; despite neutropenia and an intrinsic inability to properly mobilize from the bone marrow. In vitro they rapidly adhere to ICAM-1 coated plates, but in vivo they poorly adhere to blood vessel endothelium. Those few neutrophils that cross blood vessels and enter tissues migrate haphazardly. Following Concanavalin-A administration fragmented G184S neutrophils accumulate in liver sinusoids leading to thrombo-inflammation and perivasculitis. Thus, neutrophil Gα_i2/RGS protein interactions both limit and facilitate Gα_i2 signaling thereby promoting normal neutrophil trafficking, aging, and clearance.

SUBMITTER: Yan SL

PROVIDER: S-EPMC8202015 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Unrestrained Gαi2 Signaling Disrupts Neutrophil Trafficking, Aging, and Clearance.

Yan Serena Li-Sue SL Hwang Il-Young IY Kamenyeva Olena O Kabat Juraj J Kim Ji Sung JS Park Chung C Kehrl John H JH

Frontiers in immunology 20210531

Neutrophil trafficking, homeostatic and pathogen elicited, depends upon chemoattractant receptors triggering heterotrimeric G-protein Gαiβγ signaling, whose magnitude and kinetics are governed by RGS protein/Gαi interactions. RGS proteins typically limit Gαi signaling by reducing the duration that Gαi subunits remain GTP bound and able to activate downstream effectors. Yet how in totality RGS proteins shape neutrophil chemoattractant receptor ac ...[more]

PMID: 34135907

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Project description:ObjectivesTypically, obesity results from an inappropriate balance between energy uptake from nutrient consumption and burning of calories, which leads to a pathological increase in fat mass. Obesity is a major cause of insulin resistance and diabetes. Inhibitory G proteins (Gαi) form a subfamily that is involved in the regulation of adipose tissue function. Among the three Gαi members, i.e. Gαi1, Gαi2, Gαi3, the Gαi2, protein is predominantly expressed in adipose tissue. However, the functions of the Gαi2 isoform in adipose tissue and its impact on the development of obesity are poorly understood.MethodsBy using AdipoqCreERT2 mice, we generated adipocyte-specific Gnai2-deficient mice to study Gαi2 function, specifically in white and brown adipocytes. These mice were fed either a control diet (CD) or a high fat diet (HFD). Mice were examined for obesity development, insulin resistance and glucose intolerance. We examined adipocyte morphology and the development of inflammation in the white adipose tissue. Finally, intracellular cAMP levels as an indicator of Gαi signaling and glycerol release as an indicator of lipolysis rates were measured to verify the impact of Gαi2 on the signaling pathway in brown and white adipocytes.ResultsAn adipocyte-specific deficiency of Gαi2 significantly reduced diet-induced obesity, leading to decreased fat masses, smaller adipocytes and decreased inflammation in the white adipose tissue relative to littermate controls. Concurrently, oxygen consumption of brown adipocytes and in vivo measured energy expenditure were significantly enhanced. In addition, glucose tolerance and insulin sensitivity of HFD-fed adipocyte-specific Gnai2-deficient mice were improved compared to the respective controls. In the absence of Gαi2, adrenergic stimulation of intracellular adipocyte cAMP levels was increased, which correlated with increased lipolysis and energy expenditure.ConclusionWe conclude that adipocyte Gαi2 is a major regulator of adipocyte lipid content in diet-induced obesity by inhibiting adipocyte lipolysis in a cAMP-dependent manner resulting in increased energy expenditure.

Project description:Neutrophils are not only involved in immune defense against infection but also contribute to the exacerbation of tissue damage after ischemia and reperfusion. We have previously shown that genetic ablation of regulatory Gαi proteins in mice has both protective and deleterious effects on myocardial ischemia reperfusion injury (mIRI), depending on which isoform is deleted. To deepen and analyze these findings in more detail the contribution of Gαi2 proteins in resident cardiac vs circulating blood cells for mIRI was first studied in bone marrow chimeras. In fact, the absence of Gαi2 in all blood cells reduced the extent of mIRI (22,9% infarct size of area at risk (AAR) Gnai2-/- → wt vs 44.0% wt → wt; p < 0.001) whereas the absence of Gαi2 in non-hematopoietic cells increased the infarct damage (66.5% wt → Gnai2-/- vs 44.0% wt → wt; p < 0.001). Previously we have reported the impact of platelet Gαi2 for mIRI. Here, we show that infarct size was substantially reduced when Gαi2 signaling was either genetically ablated in neutrophils/macrophages using LysM-driven Cre recombinase (AAR: 17.9% Gnai2fl/fl LysM-Cre+/tg vs 42.0% Gnai2fl/fl; p < 0.01) or selectively blocked with specific antibodies directed against Gαi2 (AAR: 19.0% (anti-Gαi2) vs 49.0% (IgG); p < 0.001). In addition, the number of platelet-neutrophil complexes (PNCs) in the infarcted area were reduced in both, genetically modified (PNCs: 18 (Gnai2fl/fl; LysM-Cre+/tg) vs 31 (Gnai2fl/fl); p < 0.001) and in anti-Gαi2 antibody-treated (PNCs: 9 (anti-Gαi2) vs 33 (IgG); p < 0.001) mice. Of note, significant infarct-limiting effects were achieved with a single anti-Gαi2 antibody challenge immediately prior to vessel reperfusion without affecting bleeding time, heart rate or cellular distribution of neutrophils. Finally, anti-Gαi2 antibody treatment also inhibited transendothelial migration of human neutrophils (25,885 (IgG) vs 13,225 (anti-Gαi2) neutrophils; p < 0.001), collectively suggesting that a therapeutic concept of functional Gαi2 inhibition during thrombolysis and reperfusion in patients with myocardial infarction should be further considered.

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Unrestrained Gα_i2 Signaling Disrupts Neutrophil Trafficking, Aging, and Clearance.

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Unrestrained Gα<sub>i2</sub> Signaling Disrupts Neutrophil Trafficking, Aging, and Clearance.

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