Rab11 localizes to PI4P-containing organelles and its localization during cytokinesis requires fwd. (A and B) Phase–contrast (phase) and corresponding fluorescence micrographs of live squashed spermatocytes expressing GFP-Rab11 and RFP-PH-FAPP. Colocalization (arrows) of GFP-Rab11 (green) and RFP-PH-FAPP (magenta) appears white (overlay). Arrows indicate the midzone. Bars, 20 µm. (A) GFP-Rab11 and RFP-PH-FAPP (PI4P) colocalize in spermatocytes (top panels) and in dividing cells (bottom panels). Rab11 and RFP-PH-FAPP colocalize on organelles near the midzone of dividing cells (bottom panels). (B) In fwd mutant cells (fwd/Df), GFP-Rab11 and RFP-PH-FAPP colocalize to small puncta in spermatocytes (top panels) and show diffuse localization during cytokinesis (bottom panels). Note that pictures in A and B were taken on different days and the images in B were adjusted for brightness and contrast to reveal weak signals in fwd mutant. (C) Fluorescence micrographs of wild-type and fwd mutant (fwd/Df) male germ cells expressing GFP-Rab11. Arrowheads indicate elongating spermatids. GFP-Rab11 is uniformly distributed in wild type (left), but localizes to linear structures in fwd mutant (right). Bar, 20 µm.