EE motility drives ER, PO, and LD motility independently. (A) ER in U. maydis. 2D-deconvolved maximum projection of a Z-axis stack, adjusted in brightness, contrast, and gamma settings. Bar, 5 µm. (B) Contrast-inverted kymographs showing ER motility (arrowheads) in control and Δhok1. Bars, 2 s, 2 µm. See Video 6. (C) Frequency of ER motility in control and Δhok1 mutants. Bars are mean ± SEM (n = 42 cells, 2 experiments). ***, P < 0.0001 versus control (Student’s t test). (D) Examples of co-motility of EEs (mCh-Rab5a) and ER (ER-GFP). Note that EEs lead the ER during co-motility. Top kymographs contrast inverted. Bars, 3 s, 2 µm. (E) ER-independent motility of POs (mCh-SKL) and LDs (Erg6-GFP). ER was labeled by ER-mCherry (ER-mCh) or ER-GFP. Note that right kymographs show transient interaction of ER (arrowheads) with a moving LD. Upper kymographs contrast inverted. Bars: (left) 3 s, 2 µm; (right) 2 s, 1 µm. (F) Graph showing the degree of co-motility among ER, LDs, and POs. Bars are mean ± SEM (n = 3 experiments, 45–53 cells). ***, P < 0.0001 versus control (Student’s t test). (G) Motility of POs (mChSKL) and LDs (Erg6G) in the same cell. Top kymographs contrast inverted. Bars, 3 s, 2 µm. (H) Proposed role of EEs in motility of POs, LDs and ER. ER interacts with EEs independently of POs and LDs, which appear to exclude each other during motility. The mode of interaction with EEs is unknown, but linker proteins are likely to exist (?₁ and ?₂). Most EEs are constantly moving because of the activity of dynein and kinesin-3, bound to the Hok1 adapter. Transient interaction with the organelles helps PO, LD, and ER distribution and may foster interorganelle communication.