Figure 6.

Mutant FUS Forms Poorly Reversible Gels, which Fail to Release Cargo such as SMN and STAU-1

(A) When recombinant GFP-tagged SMN (or STAU-1) protein is mixed with FUS(LC) protein and gel assembly is induced by cooling, little SMN (or STAU-1) is released into the buffer (as measured by fluorescence) regardless of whether the FUS gel is derived from wild-type or ALS/FTD mutant FUS(LC) (black bars). When the gel is reversed by warming to 23°C (white bars), significant SMN (or STAU-1) protein is released into the buffer from FUS(WT) hydrogels. Very little SMN (or STAU-1) is released by the irreversible ALS/FTD mutant FUS(LC) gels (gray bars;^∗∗p < 0.001).

(B) Fluorescent molecule tracking in liquid, reversible gel, and irreversible gel assemblies reveals that in liquid state, SMN and STAU-1 diffuse rapidly at 1–4 μm/s in both mutant and wild-type samples (left two columns). On formation of a gel (center two columns), movement of SMN and STAU-1 is constrained (<1 μm²/s). On rewarming, reversible FUS(WT) gels disassemble and release SMN and STAU-1. The irreversible mutant gels continue to constrain movement of SMN and STAU-1. Image stacks are captured at 29 Hz; pixel width corresponds to 160 nm on the specimen. Representative areas are boxed and visualized in a 1.3 μm² plot, color coded to the diffusion coefficient fitted to each track.

Error bars are SEM.