Fission of melanosomal tubules requires the Arp2/3 complex activity. (A and B) SR-IFM on MNT-1 cells costained for endogenous cortactin (A) or p34 (B) and TYRP1. Cortactin and p34 (A and B; arrowheads, 3× of boxed regions) localize as discrete puncta associated with TYRP1⁺ melanosomes. (C and D) Percentage of TYRP1⁺ melanosomes positive for cortactin (C) or p34 (D). (E and F) Linear pixel values across n melanosomes show cortactin (E) or p34 (F) fluorescence enrichment at TYRP1⁺ melanosomal subdomain (cortactin, n = 102; p34, n = 94). (G and H) Relative number of cortactin (G) and p34 (H) fluorescent spots per n melanosome (C, D, G, and H; cortactin, n = 441; p34, n = 420). (I) Live imaging frame of mCh-VAMP7 expressing MNT-1 treated with DMSO or 50 µM CK-666. Magnified areas (4×) of boxed regions show tubules (arrowheads) associated with melanosomes (arrows). (J) Relative number (n) of mCh-VAMP7⁺ melanosomal tubules during 40-s movies per 256-µm² area of cells treated as in I (DMSO, n = 5 independent experiments; CK-666, n = 3 independent experiments). (K) Percentage of mCh-VAMP7⁺ tubules detaching from melanosomes in DMSO- or CK-666–treated cells (J and K; DMSO, n = 5 independent experiments; CK-666, n = 3 independent experiments). Data are presented as the mean ± SEM. Bars: (A, B, and I) 10 µm; (A, B, and I, magnifications) 1 µm. **, P < 0.01; *, P < 0.05 (unpaired t test).