Regulators Of G Protein Signaling 2 Research Articles

Effects of hypoxia on regulators of G‐protein signaling 2 (RGS2) regulation of pulmonary arterial constriction

RGS2 protein inhibits Gq‐coupled G protein coupled receptor (GPCR) signaling. Our goal was to determine whether hypoxia reduces RGS2 in pulmonary arterial smooth muscle causing enhanced GPCR agonist calcium (Ca) signaling and constriction. Human pulmonary arterial smooth muscle cells (SMC) and RGS2 KO and WT mice were used. RGS2 protein was knocked down by RGS2 siRNA. SMC were incubated in hypoxia (1% O2) or normoxia (21% O2) for 1–48 hrs. We measured RGS2 expression by western blotting, Ca mobilization with Ca dyes and constriction of pulmonary arteries and SMC by video morphometry. RGS2 siRNA caused knockdown (KD) of RGS2 by 68%±1 in SMC. Ca mobilization for U46619 and endothelin1 (ET‐1) was 2‐fold higher for RGS2 KD compared to control and constriction by ET‐1 and U46619 was increased by 21%±2 and 14%±2, respectively. Pulmonary arteries of lung slices from RGS2 KO mice had increased constriction to serotonin (26%±4) and U46619 (20%±4) compared to WT. Hypoxia significantly reduced RGS2 at 8 (54%±9), 24 (59%±8) and 48 (71%±6) hours in SMC. 48 hours of hypoxia increased constriction for ET‐1 and U46619 by 19%±3 and 21%±2, respectively while Ca was increased 2‐fold for both agonists. We found that RGS2 plays a role in GPCR constriction of SMC and pulmonary arteries. Hypoxia downregulates RGS2 causing augmented Ca signaling and constriction which may contribute to pulmonary hypertension. Support: Am Heart Asso; Am Asthma Fdn

Cloning and Characterization of Inducible Genes at the Beginning of Adipocyte Differentiation

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Cloning and characterization of inducible genes at the beginning of adipocyte differentiation

Adipocyte differentiation takes place via a complex series of steps. While PPAR gamma and C/EBP alpha are known to be master regulators, the events at the earliest stage of adipocyte differentiation are not yet known. In this study, we cloned the genes that are induced at the beginning of the differentiation of 3T3-L1 preadipocyte cells. Of 102 clones obtained, only several clones were already reported as genes that are expressed differentially during adipocyte development. The expression of TCL/TC10 beta L (TC10-like/TC10 beta Long) and RGS2 (regulators of G protein signaling 2) genes isolated here rapidly increased after the addition of inducers (insulin, dexemethasone, 3-isobutyl-1-methylxanthine, fetal bovine serum [FBS]). Further, the antisense TCL/TC10 beta L inhibited the adipogenesis of mouse 3T3-L1 preadipocyte cells, prevented cytoplasmic triglyceride accumulation, and decreased the expression of PPAR gamma and C/EBP alpha. Moreover, the constitutive overexpression of TCL/TC10 beta L or RGS2 in the mouse fibroblast cell line NIH-3T3 results in efficient adipocyte conversion when stimulated with 10% FBS, insulin, 3-isobutyl-1-methylxanthine, dexamethasone, and PPAR gamma ligand BRL49653. These results strongly suggest that TCL/TC10 beta L and RGS2 have crucial roles in the program of adipocyte differentiation, probably linked to the PPAR gamma pathway. Using a subtraction protocol, the genes specifically regulated by TCL/TC10 beta L were also isolated. The expression pattern of some was similar to TCL/TC10 beta L expression in adipogenesis, suggesting that these genes are regulated by TCL/TC10 beta L.

RGS2 Promotes Adipocyte Differentiation in the Presence of Ligand for Peroxisome Proliferator-activated Receptor γ

The events at the earliest stage of adipocyte differentiation are yet to be fully elucidated. Previously, we cloned the genes that are induced at the beginning of the differentiation of mouse 3T3-L1 preadipocyte cells. We found that the gene expression of regulators of G protein signaling-2 (RGS2) rapidly increased after the addition of inducers and decreased at 3-12 h. The expression pattern of RGS2 mRNAs differed among growth-arrested and proliferating 3T3-L1 cells and NIH-3T3 cells, indicating a specificity for adipogenesis. Here we report that the ectopic expression of RGS2 using a retroviral system in mouse NIH-3T3 cells promotes adipogenesis only in the presence of BRL49653, which is a ligand for the peroxisome proliferator-activated receptor gamma (PPARgamma). These results strongly suggest that RGS2 play a crucial role in the program of adipocyte differentiation and may contribute to the function of PPARgamma.