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. 2015 Dec 4;1(11):e1500615. doi: 10.1126/sciadv.1500615

Fig. 3. Lysine 107–dependent PRMT8 induces neurite branching in NGF-stimulated PC12 cells.

Fig. 3

(A) Alignment of the amino acids in the HKD motifs from human PRMT8 and PLD2 and the mitochondrial PLD isoform MitoPLD. The amino acids that were mutated in the study are marked with a red asterisk. (B) Schematic representation of the K107R mutation generated in human PRMT8. (C) Translocation of GFP-PABD (Spo2051–91), a PA sensor, to the plasma membrane upon PRMT8-mediated PA generation. PC12 cells were cotransfected with GFP-PABD and pmCherry vector (control), WT PRMT8-mCherry, or the PRMT8 K107R mutant. Controls were stimulated with or without NGF (100 ng ml−1) for 5 min (left panel). Scale bar, 50 μm. Neurite development of NGF-differentiated PC12 cells. (D) PC12 cells were coexpressed with the pVenus vector and pmCherry vector (control), WT PRMT8-mCherry, or the PRMT8 K107R mutant, and stimulated with NGF (100 ng ml−1) for 24 hours. Scale bars, 30 μm. (E to H) Cells bearing neurites that were twofold longer than their cell body lengths were scored in terms of (E) total neurite length, (F) the number of branching points per cell, (G) the number of neurite roots, and (H) the number of extremities. Results are shown as mean ± SEM of five independent experiments with at least 50 cells scored in each experiment; *P < 0.05, **P < 0.01, ***P < 0.0001, or NS as compared to the control (CON).