Extended Data Figure 2. GFP:Mud localizes at TCJ from G2 interphase to mitosis.

(a) Scheme depicting the accumulation of the Drosophila FUCCI reporters during the cell cycle. ECFP:E2F1 accumulates during G1 phase, G2 phase and mitosis, whereas mRFP1:CycB accumulates during S phase, G2 phase and mitosis⁵⁵.

(b) Localization of GFP:Mud (green left column and white in the second column panels), mRFP1:CycB (red in the left column and white in the third column panels) and ECFP:E2F1 (blue in the left column and white in the right column panels) in epithelial cells of the pupal notum tissue. Confocal sections at the level of septate junctions are shown. Cells in G1 (n=21), S (n=6), G2 (n=35) phases and mitosis (n=6) are indicated in the left panels. During both G1 and S phase (upper two row panels), GFP:Mud is weakly localized at the nuclear envelope membrane, weakly localized at the cortex and at the apically localized centrioles (not shown). During G2 phase GFP:Mud becomes prominently localized at the TCJ (one cell in the 1^st row panels and 2 cells in the 3^rd row panels). Arrows indicate examples of TCJ GFP:Mud accumulation. During mitosis GFP:Mud remains localized at the TCJ and accumulates on the spindle and the spindle pole (bottom row panels). Similar results were obtained on fixed tissue for which the cell cycle phases were determined using the PCNA S-phase marker and the nucleus size to distinguish cells in G1 or G2 interphases (not shown).

(c) GFP:Mud (green arrows) and ChFP:Mud (red arrows) in adjacent tissue patches in G2 (n=31) and mitotic (n=8) cells. The FLP/FRT system was used to generate adjacent groups of cells labelled with either GFP:Mud or ChFP:Mud. By analysing the distribution of GFP:Mud in dividing cells adjacent to ChFP:Mud interphasic cells, we found that GFP:Mud was localized at the TCJs of the dividing cell from G2 through mitosis.

Scale bars: 1μm (b, c).