Supplementary Materials

RNA buffers the phase separation behavior of prion-like RNA binding proteins

Shovamayee Maharana, Jie Wang, Dimitrios K. Papadopoulos, Doris Richter, Andrey Pozniakovsky, Ina Poser, Marc Bickle, Sandra Rizk, Jordina Guillén-Boixet, Titus Franzmann, Marcus Jahnel, Lara Marrone, Young-Tae Chang, Jared Sterneckert, Pavel Tomancak, Anthony A. Hyman, Simon Alberti

Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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  • Materials and Methods
  • Figs. S1 to S24
  • Captions for Movies S1 to S4
  • References

Images, Video, and Other Media

Movie S1
RNA degradation induces phase separation of nuclear FUS. This movie shows the formation of nuclear FUS assemblies by phase separation immediately after RNase A microinjection in a HeLa cell expressing FUS-GFP. Initially the FUS is localized diffusely in the nucleus. At 0.28 min microinjection is performed, after which condensation of FUS assemblies can be seen. Two arrow heads indicate the region where FUS assemblies fuse to form larger spherical assemblies. The cell at the bottom right was not injected with RNase and shows a diffuse FUS localization pattern.
Movie S2
Nuclear FUS assemblies induced through RNA degradation harden with time. This movie shows nuclear FUS assemblies after 30 min of RNase A microinjection in HeLa cells expressing FUS-GFP. The shown FUS assemblies stick to each other and do not undergo fusion anymore.
Movie S3
Aged FUS has solid-like properties. This movie shows the optical trap assisted fusion of FUS droplets formed in vitro. The drop in the focus is held in place by one optical trap and the second droplet is then trapped with another optical trap and the two droplets are then brought into close proximity. Once the droplets are close to each other fusion occurs. Left: Fusion of freshly phase-separated FUS droplets (young). Right: Fusion of 3 hours old, aged FUS droplets.
Movie S4
RNA degradation induces TDP43 phase separation in the nucleus. HeLa cells transiently expressing TDP43-GFP were microinjected with RNase A. Note that RNase injection triggers TDP43 phase separation. The resulting TDP43 droplets are initially dynamic and grow in size through fusion events, but they harden with time and show less fusion events.