Without capping protein, patches acquire cable-like characteristics over time. (A) Maximum intensity projections of confocal images of wild-type and cap2Δ cells expressing mNeon–Tpm1 (tropomyosin; cyan) and Arc15-RFP^mScarlet (Arp2/3 complex; magenta). In wild-type cells, mNeon–Tpm1 signal is detected in the cytosol and on actin cables (white arrow), but not patches (see merge). In cap2Δ cells, mNeon–Tpm1 signal is detected in the cytosol, on cables (white arrow), and on Arc15-RFP^mScarlet marked patches (see merge). Some of these patches have very little Arc15-RFP^mScarlet signal with bright mNeon–Tpm1 signal (yellow arrowhead). (B–G) Protein dynamics at Arc15-RFP^mScarlet marked patches were monitored in TIRF. Graphed data are the 20-point moving average signal (solid lines) and the 95% CI (shaded regions). Images of example patches from a time series are shown in Fig. S5. (B) Average relative mNeon–Tpm1 signal (cyan) and Arc15-RFP^mScarlet (magenta) signal at 33 wild-type patches (left) and 33 cap2Δ patches (right). In wild-type cells, patches exclude mNeon–Tpm1; however, mNeon–Tpm1 signal from cables near patches gives rise to some overlap in the signals for mNeon–Tpm1 and Arc15-RFP^mScarlet. (C) Quantification of the time between the peak Arc15-RFP^mScarlet signal and peak mNeon–Tpm1 signal at the same patches measured in B. (D) Average relative Abp140-mNeon (cyan) and Arc15-RFP^mScarlet (magenta) signals at 37 wild-type patches (left) and 43 cap2Δ patches (right). (E) Quantification of the time between the peak Arc15-RFP^mScarlet signal and peak Abp140-3xmNeon signal at the same patches shown in D. (F) Average relative Sac6-GFP^ENVY (cyan) and Arc15-RFP^mScarlet (magenta) signals at 40 wild-type patches (left) and 40 cap2Δ patches (right). (G) Quantification of the time between the peak Arc15-RFP^mScarlet signal and peak Sac6-GFP^ENVY signal at the same patches shown in F. In graphs (C, E, and G), a line is drawn at the mean and error bars represent the SD. Statistical significance is calculated by Students t test (***, P ≤ 0.001; ****, P ≤ 0.0001). (H) Model showing acquisition of cable-like features as patches mature in cap2Δ cells. Actin patch networks in both wild-type and cap2Δ cells are first nucleated by the Arp2/3 complex. In the absence of Cap1/2 (red), actin filaments (black) grow abnormally long and recruit proteins normally associated with actin cables, formins (blue) and tropomyosin (green), while proteins associated with actin patches, Arp2/3 (gray) and Fimbrin/Sac6 (yellow), are lost.