Fig. 2: YARS1 binds to F-actin and organizes actin filaments in vitro.

a Coomassie-stained gels of supernatant (S) and pellet (P) fractions from high-speed pelleting (see schematic) of 0.5 µM YARS1^WT or YARS1^E196K upon increasing F-actin concentrations. b Graphical representation of the YARS1 fraction in the pellet obtained in three independent experiments, fitted into Michaelis-Menten nonlinear curve. c F-actin bundling capacity of YARS1 proteins tested upon low-speed pelleting (see schematic) of increasing concentrations of YARS1 and preassembled 2 µM F-actin. Graphs in d and e show the fraction of bundled F-actin and YARS1 in the pellets from five independent experiments; error bars in b, d, and e—SEM; *p = 0.0187, two-sided unpaired t test. Supernatant and pellet samples derive from the same experiment and gels were processed in parallel. f A schematic representation of the TIRF experiment. g Representative time-lapse images from a TIRF microscopy movie of 10% OG-actin labeled filaments alone (CTRL) or in the presence of YARS1 demonstrate stronger bundling capacity of YARS1^E196K at 0.5 µM (areas of bundle formation depicted with yellow arrowhead) and pronounced cable formation at 2 µM. h Graph representing the fold change of mean gray value of OG-actin in control and upon flow-in of 0.5 µM YARS1, measured in areas of cable formation. n = 3 represents randomly selected, non-overlapping fields of view, imaged within one TIRF experiment per condition (CTRL, YARS1^WT, and YARS1^E196K). Presented data derive from one of two independently performed experiments. Error bars—SD; **p = 0.0011, ***p = 0.0003, two-sided unpaired t-test. Source data are provided as a Source data file.