Fig. 4: NRE1 deletion reduces NAMPT induction and LPS/IFNγ inflammatory responses. | Nature Communications

Fig. 4: NRE1 deletion reduces NAMPT induction and LPS/IFNγ inflammatory responses.

From: A Stat1 bound enhancer promotes Nampt expression and function within tumor associated macrophages

Fig. 4

a NRE1 Cre-lox knockout scheme: diagram indicating inserted loxP sites bounding NRE1 relative to STAT1 consensus sites, CRISPR deletion alleles, and loxP detection primers, in both the floxed and deleted alleles. Representative agarose gel showing PCR products from macrophages using the indicated primers for the presence of each loxP site relative to WT (non-floxed) NRE1, and generation of a deletion junction product from Vav-Cre NRE1 fl/fl (NRE1-KO) primary bone marrow macrophages. Size markers are in bp. b RT-qPCR from the indicated macrophage RNAs shows the absence of NRE1 signal in the deletion. c Mature Nampt mRNA expression measured by RT-qPCR from WT and NRE1-KO macrophage RNAs treated with IFNγ. d Western analysis of multiple IFNγ or untreated macrophage cultures from mice of the indicated genotypes showing lowered inducibility of NAMPT protein relative to ACTIN controls. e Mature Nampt mRNA expression in the presence of LPS/IFNγ is blunted in the absence of NRE1 in Vav-Cre NRE1 fl/fl (NRE1-KO) macrophages. f Ccr1 mRNA expression measured by RT-qPCR from WT and NRE1-KO macrophage RNAs treated with IFNγ. g Inflammatory and glycolytic pathway genes’ mRNA expression measured by RT-qPCR from WT and NRE1-KO macrophage RNAs treated with IFNγ and LPS. For b, c, eg, results presented are the average of N = 3 biological replicates, and are representative of at least three independent experiments. p values were determined by a two-tailed unpaired t-test: *p < 0.05, **p < 0.005, ***p < 0.0005, ****p < 0.00005. Error bars represent SEM. Uncropped images and raw scans provided in Source Data file in Supplementary information.

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