Pak1 dynamics at the cells ends are slower than that of Cdc42, Scd1 and Scd2. (A) Simulations of our models show a delay or phase shift between the peak accumulations of Cdc42-GTP and Pak1 at the growing end. (B) Pairs of positive-feedback polarity proteins (CRIB-3xGFP and Scd1-tdTomato, CRIB-3xGFP and Scd2-mCherry) were simultaneously observed at the cell end. (C) Each positive-feedback protein (Scd1-tdTomato, CRIB-mCherry, and Scd2-mCherry) was observed independently with the negative-feedback protein, Pak1-mEGFP. (D) The shift in time required to get the best correlation coefficient between signals from each pair of proteins was quantified using cross-correlation analysis (n ≥ 29 cell ends). (E) The Hilbert transform was applied to the smoothened traces of Cdc42-GTP and Scd1 for their phase reconstruction and phase shift (top). The Hilbert transform was applied to the smoothened traces of Pak1 and Scd1 for their phase reconstruction and phase shift (bottom). The thinnest traces represent normalized raw data, the thickest traces represent smoothened traces based on the normalized data, and the darkest traces represent the phase progression achieved through the Hilbert transform. (F) FRAP analysis shows the half-life of recovery for Scd1-mNG, Scd2-GFP, and Pak1-mEGFP (n ≥ 10 cells). n.s., not significant; P value, *<0.05, **<0.005, ****<0.0001, one-way ANOVA followed by Tukey’s multiple comparison test.