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. 2013 Feb 26;11(2):e1001495. doi: 10.1371/journal.pbio.1001495

Figure 3. Gps1 regulates Rho1 at the cell division site.

Figure 3

(A) Time-lapse series of GFP-Rho1 and Myo1-3Cherry at the bud neck in gps1Δ and wild-type cells. Red arrowhead indicates the new polarization site. (B) Quantification of (A) showing GFP-Rho1 fluorescence levels in wild-type (n = 6) and gps1Δ (n = 6) cells. Cytokinesis events are indicated. Chs2, Fks1, and Cts1 are cargos of the secretory pathway. (C) Quantification of the levels of bud-neck-associated Bni1-GFP (wild-type [n = 6], gps1Δ [n = 7], gps1Δ lrg1Δ [n = 8]), Sec3-GFP (wild-type [n = 5], gps1Δ [n = 5], gps1Δ lrg1Δ [n = 5]), and Fks1-GFP (wild-type [n = 5], gps1Δ [n = 4], gps1Δ lrg1Δ [n = 5]). (D) Continuous tethering of GFP-Rho1 to the bud neck with Shs1-GBP. Kymographs of the mother (m)–daughter (d) cell axis (yellow line) of the indicated genotypes are shown. The red arrowhead indicates the time of cell splitting, as determined by the loss of fluorescence signal between mother and daughter cell. (E) The immunoblot shows GFP-Rho1 levels in GPS1, GPS1 SHS1-GBP, gps1Δ, and gps1Δ SHS1-GBP cells. An unspecific (unsp.) signal was used as a loading control. (F) Serial dilutions of the indicated cell types carrying GPS1 on an URA3-based plasmid. (G) Septum thickness of GFP-Rho1-containing GPS1 (n = 34), GPS1 SHS1-GBP (n = 25), gps1Δ (n = 45), and gps1Δ SHS1-GBP (n = 46) cells. (H) Septum thickness of wild-type (n = 42), gps1Δ (n = 48), lrg1Δ (n = 26), and gps1Δ lrg1Δ (n = 32) cells. (I) Serial dilutions of the indicated cell types carrying GPS1 on an URA3-based plasmid. Error bars in (B) and (C) show the standard deviation. “**” indicates p<0.0001. Scale bars: 5 µm. a.u., arbitrary units; fl. intensity, fluorescence intensity; PS, primary septum; SC, synthetic complete medium; SS, secondary septum.