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. 2016 Jun 28;5(5):802–818. doi: 10.1002/mbo3.371

Figure 4.

Figure 4

Inactivation of the folC2 gene resulted in repression of the hdc gene cluster. (A) Quantitative real‐time PCR yielded evidence of decreased expression of hdcA and hdcP genes in the L. reuteri 6475::folC2 mutant compared to wild‐type L. reuteri 6475. The gene narI, which was suggested by the comparative transcriptomics analysis to be down‐regulated in the folC2 mutant, was also repressed in 6475::folC2. Expression ratios of each gene (folC2 mutant vs. wild‐type) were calculated, and results represent the mean ± SD, n = 3, **P < 0.01, *P < 0.05 compared to the theoretical mean of 1.0. (B) Quantitative real‐time PCR demonstrated increased expression of all three hdc genes, hdcA, hdcB, and hdcP, when L. reuteri 6475 was grown in medium supplemented with l‐histidine compared to unsupplemented medium. Expression ratios of each gene (l‐histidine‐supplemented vs. unsupplemented) were calculated. Results represent the mean ± SD, n = 3, ***P < 0.005, **P < 0.01, *P < 0.05 compared to the theoretical mean of 1.0. (C) Quantitative real‐time PCR demonstrated no significant changes in expression of all three hdc genes when L. reuteri 6475::folC2 was grown in medium supplemented with l‐histidine compared to unsupplemented medium. (D). Inactivation of the folC2 gene resulted in decreased histamine production. Quantification of secreted L. reuteri‐derived histamine by a histamine‐specific ELISA demonstrated decreased histamine production in the folC2 mutant compared to wild‐type L. reuteri 6475 even when grown in l‐histidine‐supplemented medium. Data were analyzed by two‐way ANOVA. Results represent the mean ± SD, n = 3, P < 0.0001 compared to wild‐type 6475.