Extended Data Figure 7: Moesin is a target of PP1-87B–Sds22 and controls cortical relaxation at anaphase. | Nature

Extended Data Figure 7: Moesin is a target of PP1-87B–Sds22 and controls cortical relaxation at anaphase.

From: Kinetochore-localized PP1–Sds22 couples chromosome segregation to polar relaxation

Extended Data Figure 7

ac, The effect of constitutively active moesin on anaphase polar relaxation. a, An SOP cell (1 of 13 cells) expressing constitutively active moesin (moesin(T559D)) imaged in metaphase and anaphase (top panel) and a kymograph of a cross-section over time (yellow box). Only the anterior pole is indicated. b, Plot of the DNA-to-cortex distance over time for a representative moesin–GFP-expressing cell (cell X) and a moesin(T559D)–GFP-expressing cell (cell Y in a). mD, mean distance during anaphase. c, Box-and-whisker plot (median and 10–90 percentiles) of mD in moesin–GFP and moesin(T559D)–GFP-expressing cells. This shows that the DNA comes into close apposition to the cortex in cells expressing constitutively active moesin as the result of failure to trigger efficient polar relaxation, as it does in cells depleted for PP1-87B or Sds22. df, The effect of moesin(T559D) expression on pre-furrow elongation in the same experiment as ac. d, Images show representative SOP cells expressing moesin–GFP or moesin(T559D)–GFP transgenes at metaphase and anaphase (out of 13 cells in each case). e, Outlines of the boundary of cells shown in d at different times during anaphase. f, Bar graph of cell elongation in these two backgrounds showing mean and s.d. As observed in PP1-87B- or Sds22-depleted cells, moesin(T559D)–GFP-expressing cells show aberrant cell elongation at anaphase. g, Immunoprecipitation (IP) assays showing moesin dephosphorylation by PP1-87B–Sds22. Calyculin A (CalA) is an inhibitor of PP1 activity (left panel). Upon addition of CalA, PP1-87B activity is suppressed, leading to higher levels of phosphorylated moesin than in the absence of the compound (see p-Flag–moesin immunoblotting). PP1-87B acts with Sds22 to dephosphorylate active moesin (right panel). Upon Sds22 depletion, PP1-87B is less efficient in inactivating moesin. Red arrows indicate PP1-87B–GFP band. Results in g were replicated three times. h, Scheme of PP1–Sds22-dependent inactivation of moesin. In a and d, scale bar, 5 µm. Significance was assessed using a two-tailed unpaired t-test. PM, plasma membrane.

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