Fig. 2. Subcellular localization of ApsB. (A) Both ApsB-GFP (SDM1000) (a) and GFP-ApsB (SEa3) (b) localized to the nuclear SPB (green dots at nuclei) and to the cytoplasm (arrowheads). Cytoplasmic ApsB was always bound to MTs and moved fast up and down the filaments (see text and supplementary material, Movies 8-10a). Nuclei were stained with red fluorescent protein (DsRedT4) in (a) or with DAPI in (b). (B) C-terminally tagged ApsB as well as N-terminally tagged ApsB (C) was detected at the poles of the mitotic spindle. In (B) ApsB-GFP and GFP-TubA is visible (SDM40), while a red signal was used to set apart the mRFP1-ApsB (C,b) from the green GFP-TubA (C,a) (SDV1B). In (C,c) an overlay of (Ca,b) is shown. (D) In addition, N-terminally tagged GFP-ApsB localized to septa [arrows in (D,a,b)], while C-terminally tagged ApsB-GFP (E) did not [arrowheads in (E,a,b)]. Septa were not stained completely, but GFP-ApsB is seen as dots near the septal hole, as insert in (D, a) shows. The septa in (E) were negatively stained by weak cytoplasmic GFP background [insert in (E,a)]. Bar, 3 µm.