Identification of FAM173B as a protein methyltransferase promoting chronic pain
Fig 8
Methyltransferase activity requirement.
WT hFAM173B but not hFAM173B-D94A expression in sensory neurons prolonged carrageenan-induced (A) mechanical and (B) thermal hypersensitivity (n = 6 mice). (C–F) Compared to WT hFAM173B, the methyltransferase-inactive mutant hFAM173B-D94A did not increase Iba1-positive area in (C) DRG and (D) dorsal horn of the spinal cord (n = 5 mice) at day 5 after intraplantar carrageenan injection, or enhance (E) ΔTMRM fluorescence in N2A in vitro (n = 115–150 cells) and (F) ROS production in small-diameter neurons in vivo (hFAM173B n = 7; hFAM173B-D94A n = 5 mice) at 5 days after intraplantar carrageenan. (G) Supernatants of 15 h–cultured sensory neurons expressing hFAM173B-D94A that were stimulated with 100 ng/ml TNFα for 6 hours and subsequently washed did not enhance IL6 release in spinal microglia in vitro (hFAM173B n = 4; hFAM173B-D94A n = 8 wells). Data are represented as mean ± SEM. * = P < 0.05; ** = P < 0.01; *** = P < 0.001. Statistical analyses were performed by unpaired two-tailed t tests (F, G), one-way ANOVA (C–E), or by a two-way ANOVA (A, B) followed by a post-hoc Holm-Sidak multiple comparison test. Underlying data can be found in S1 Data. DRG, dorsal root ganglia; Iba1, ionized calcium binding adaptor molecule 1; IL6, interleukin 6; N2A, Neuro2a; ROS, reactive oxygen species; SEM, standard error of the mean; TMRM, tetramethylrhodamine methyl ester; TNFα, tumor necrosis factor α; WT, wild-type.