Similar to Tau 255–441, Tau 151–254 can form droplets in vitro. (A) Representative bright-field images of Tau 151–254/polyU droplets. Scale bar is 50 µm. (B) Direct comparison of Tau 151–254 (red) and Tau 255–441(black) shows phase separation occurs for both protein segments with addition of PolyU or heparin. In contrast to the Tau 255–441 heparin complex, Tau 151–254 does not show the capacity to form fibrils with either PolyU or heparin. Turbidity was evaluated at 500 nm (filled bar); fibrilization was evaluated by ThT fluorescence (striped bar). Protein concentration was 50 µM, PolyU concentration (where applicable) was 125 µg/ml, and heparin concentration was 12.5 µM. All data were normalized by the largest measured value or absorbance or fluorescence. n = 3; error bar in SD. (C) Predicted Tau 151–254/RNA binodal phase coexistence points modeled using FTS. Calculations performed for Tau 151–254 (red dots) are compared with our previously reported (Lin et al., 2019) phase diagram for Tau 255–441 (black dots), showing that both sequences have a stable two-phase region under similar electrostatic environments and solvent conditions. The conditions for stable binodal phase coexistence depend on the strength of the electrostatic interaction, characterized in our model by the l_B as the input parameter. The region defined by the coexistence points is the region at which the system forms a thermodynamically stable droplet phase in coexistence with a solution phase. Areas corresponding to one-phase or two-phase are shown in the diagram.