Fig. 2: YAP-chromatin dwell times are sensitive to protein charge.

A YAP contains two IDRs (A, top) that are negatively charged (alpha-fold prediction of YAP protein with surface charge annotation, A bottom), raising the possibility of electrostatic interactions for condensate formation. B Schematic depiction of YAP’s behavior in the presence of a negatively charged tag (net charge: −30), which would be expected to impair electrostatic-based interactions of YAP in condensates. C Dual Halo-YAP labeling for simultaneous visualization of total YAP (Halo-tag ligand JFX650) and single YAP molecules (Halo-tag ligand JFX549). Comparing cells with similar total YAP levels (dashed magenta outlines, left panel), the negative charge tag (−30) shows decreased YAP-chromatin interaction events (kymographs, right panel) as compared to a neutral charge tag (GFP(0)). Scale bar, 10 s. D Quantitation of YAP single-molecule dwell times from conditions shown in (C). Shown are the average fold-change for different dwell time populations (unspecific binding events, <1.2 s; specific binding events, >= 1.2 s, >= 7 s, >= 10 s, as determined in Fig. 1C). Only the specific YAP binding events (>= 1.2 s dwell time) are impaired by the negative charge tag. Shown are mean +/− SEM from N = 4 independent experiments. p values from two-sided unpaired Student’s t test: GFP(0)-Halo-YAP vs GFP(−30)-Halo-YAP for dwell times <1.2 s, p = 0.02; dwell times >= 1.2 s, p = 0.005; dwell times >= 7 s, p = 0.02; dwell times >= 10 s; p = 0.004.