Ags1 coincides spatially and temporally with Bgs1 in the growing sites: CAR, septum, and poles. (A) Ags1 and Bgs1 colocalize in the medial zone before septum synthesis. ags1⁺-RFP GFP-bgs1⁺ cells were imaged by time-lapse microscopy. Arrowhead: appearance of Ags1 and Bgs1 in the cell middle; arrow: appearance of septum (CW staining). (B) Ags1 and Bgs1 colocalize in the CAR and septum membrane. (C) Ags1 and Bgs1 colocalize in the old end before cytokinesis completion. Elapsed time is shown in minutes. (D) Ags1 and Bgs1 colocalize in the aberrant septa of the medial ring mutants. mid1-366 ags1⁺-RFP GFP-bgs1⁺ cells were shifted to 32°C for 6 h. (E) Bgs1 coimmunoprecipitates with Ags1. Solubilized membrane proteins (MP; see Materials and methods) from the indicated strains were immunoprecipitated (IP) with anti-GFP serum. Solubilized MP (left) and IP (right) were transferred to the same membrane and blotted with monoclonal anti-GFP or anti-HA antibodies. Bars, 2.5 µm.