Rhes’s SUMO E3-like domain promotes TNT-like Rhes tunnels, but only a Rhes WT travels efficiently from cell to cell. (A) Diagram of Rhes domains. (B) Representative confocal image of striatal neuronal cells coexpressing either GFP-tagged Rhes WT or its mutants with mCherry-Farnesyl 5. Arrows indicate TNT-like process. (C–F) Bar graphs show the quantification of all protrusions (C), all TNT-like protrusions (D), single TNT-like process/cell (E), and more than one TNT-like process/cell (F). Data mean ± SEM are shown in Table 1; one-way ANOVA, **, P < 0.05; ***, P < 0.001 (compared with GFP); ^##, P < 0.01; ###, P < 0.001 (compared with Rhes WT); n = 210–230 cells/group, from three independent experiments. (G) Experimental design for H. (H) FACS plots for GFP-tagged Rhes WT and its mutant or mCherry alone expressing striatal neuronal cells (20,000 cells were recorded). (I) Bar graph shows ± SEM; one-way ANOVA (***, P < 0.001, compared with Rhes WT), n = 3. Quantification of GFP and mCherry double-positive population (%) for Rhes-FL, Rhes S33N, Rhes C263S, Rhes 1–171, and Rhes 171–266.