JCS026716 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Anillin localization is unaltered after RNAi depletion of spectrins. Cells were treated for 72 hours with dsRNAs directed against GFP (control), α-spectrin, β-spectrin, β_H-spectrin, α- + β-spectrin, or α- + β_H-spectrin, and then fixed and stained to detect Anillin (red in merged panels), tubulin (green in the merged panels) and DNA (blue in the merged panels). Bar, 10 µm.

• Supplemental Figure S2 -

  Fig. S2. RacGAP50C localization in telophase cells after Anillin RNAi. Cells were treated for 48 hours with dsRNAs directed against either Anillin (Ani RNAi) or GFP (control) and then fixed and stained to detect tubulin (green in the merged panels), RacGAP50C (red in merged panels) and DNA (blue in the merged panels). Judging by the size of the nuclei, the Anillin RNAi cells are most likely tetraploid, indicating that cytokinesis already failed once because of Anillin depletion. See also the western blot in Fig. 4 for the level of Anillin depletion. Bar, 10 µm.

• Supplemental Figure S3 -

  Fig. S3. F-actin and Anillin localization during cell division. Cells were fixed and stained to detect Anillin (red in merged panels), F-actin (green in the merged panels) and DNA (blue in the merged panels). Bar, 10 µm.

• Supplemental Figure S4 -

  Fig. S4. Latranculin-A treatment promotes microtubule stabilization. Cells were treated with DMSO (control) or Latranculin A (Lat-A) for 1 hour and then fixed and stained to detect Anillin (red in merged panels), tubulin (green in the merged panels) and DNA (blue in the merged panels).

• Supplemental Figure S5 -

  Fig. S5. Subcellular localization of GFP-tagged Anillin fragments. (A) Cells stably expressing the GFP-tagged Anillin fragment lacking only the PH domain (see Fig. 3) were fixed and stained to detect tubulin (red in merged panels), GFP (green in the merged panels) and DNA (blue in the merged panels). (B) Cells stably expressing the GFP-tagged Ani_410-1104 fragment lacking the Ac, My and PH domains (see Fig. 3) were fixed and stained to detect tubulin (red in merged panels), GFP (green in the merged panels) and DNA (blue in the merged panels). (C) Cells stably expressing the GFP-tagged Ani_410-766 fragment lacking the Ac, AHD, My and PH domains (see Fig. 3) were fixed and stained to detect tubulin (red in merged panels), GFP (green in the merged panels) and DNA (blue in the merged panels). Bars, 10 µm.

• Supplemental Figure S6 -

  Fig. S6. GFP-tagged Ani_410-1104 co-localizes with RacGAP50C in telophase. Top: Cells were fixed and stained to detect tubulin (red in merged panels), Anillin (green in the merged panels) and DNA (blue in the merged panels). Bottom: Cells stably expressing the GFP-tagged Ani_410-1104 fragment lacking the Ac, My and PH domains (see Fig. 3) were fixed and stained to detect GFP (green in the merged panels), RacGAP50C (red in merged panels) and DNA (blue in the merged panels). Bars, 10 µm.

• Supplemental Figure S7 -

  Fig. S7. Pnut and Sep2 do not localize to the cortex in metaphase. Cells were fixed and stained to detect tubulin (green in the merged panels), DNA (blue in the merged panels) and either Pnut or Sep2 (red in merged panels). Bars, 10 µm.