{"CAPTION FIG1.png": "'Figure 1: Actin filament Severing by Collins Observed by Evanescent Wave Fluorescence Microscopy Conditions were as follows: 50 mM KCl, 10 mM imidazole, 1 mM MgCl2, 1 mM EGTA, 0.2 mM ATP, 0.25% methyl ceHulose, 100 mM DTT, 15 mM glucose, 20 mg/mL catalase, and 100 mg/mL glucose oxidase at 22\u00b0 C and pH 7.0. (A) Fluorescence micrographs of entire microscopic fields of 25% Oregon Green-labeled actin filaments polymerized for 6 min taken at indicated time points after wash in from time-lapse movies of actin filament severing. First row, actin filaments washed with S. pombe coffin at the optimal severing concentration of 10 nM. Filaments were attached to the glass surface at their barbed ends by S. pombe GST-CdCl21[FH1FH2]p. Second row, actin filaments washed with actophorin at the optimal severing concentration of 100 nM. Filaments were attached to the glass surface by NEM-inactivated muscle myosin II. (B) Detailed mortgage of one actin filament being sewered by 100 nM actophorin over time.\\n\\n'", "CAPTION FIG2.png": "'Figure 2: Analysis of Actin Filament Severing by Collins (A) Dependence of severing activity on the concentration of _S. pombe_ cofflin under the conditions of Figure 1. Degree of saturation of the subunits in the filaments was calculated by using Equation. 2. The units of severing activity are severing events per um of actin filament per second. Optimal severing activity occurs at 10 nM _S. pombe_ cofflin with cofflin bound to 0.12% of the actin subunits. No severing occurred without cofflin or with 1 \u03bcM S. pombe cofflin, which saturates 49% of the actin subunits.\\n\\n'", "CAPTION FIG4.png": "'Figure 4: Effect of Collins on Actin Filament Elongation and Nucleation (A\u2013C) Elongation was observed by evanescent wave fluorescence microscopy with 50% Oregon Green-labeled ATP-actin. Conditions as in Figure 3. Dependence of the elongation rate on the concentration of unlabeled actin: (A) actin alone, (B) actin with 32 \u03bcM human coffin, and (C) actin with 24 \u03bcM _M_S_._pombe coffin. Filled circles represent barbed-end rates and filled triangles represent pointed-and rates. Slope = k_v_-int_ = k_v_-, and x-int = Cc. (D\u2013F) nucleation of actin filaments by coffins. Evanescent wave fluorescence micrographs of filaments formed by 0.8 \u03bcM 50% Oregon Green-labeled ATP-actin after 400 s in polymerization buffer. Conditions as in Figure 1. (D) Actin alone, (E) actin with 32 \u03bcM human coffin, and (F) actin with 24 \u03bcM _M_S_. pombe coffin. (G\u2013I) Quantitation of barbed ends formed by nucleation by dilution of samples into 2 \u03bcM 25% pyrenyl ATP-actin monomers and measurement of the rate of elongation. Conditions for nucleation and subsequent elongation were as in Figures 2C and 2D. (G) Time course of barbed-end formation by 6 \u03bcM ATP-actin with 10 \u03bcM _M_S_._pombe coffin fit with a hyperbola. Inset, time course of ends formed by 6 \u03bcM ATP-actin with 10 \u03bcM _M_S_. pombe coffin (filled circles), or with 10 \u03bcM _M_S_._pombe coffin and 10 \u03bcM _M_S_._pombe profilin (filled triangles). (H) Dependence of and formation on actin monomer concentration. A range of ATP-actin concentrations was polymerized for 30 s with 10 \u03bcM _M_S_._pombe coffin. (I) Dependence of the rate of end formation by 10 \u03bcM _M_S_._pombe coffin concentration. Samples were polymerized for 60 s before dilution.\\n\\n'", "CAPTION FIG5.png": "'Figure 5: Concentration Dependence of Codlin Activity\\n\\nVery low concentrations of codlin (blue ovals) do not bind actin filaments (green). At an optimal low concentration of codlin, single codlins bind and sever actin filaments. Capping of severed filament barbed ends promotes dissociation of actin monomers (green circles) from pointed ends, leading to filament disassembly. Higher concentrations of codlin bind cooperatively to actin filaments and promote the release of inorganic phosphate (P\\\\({}_{\\\\beta}\\\\)) but do not sever them. Very high concentrations of codlin bind actin monomers and stimulate nucleation, leading to actin filament assembly.\\n\\n'", "CAPTION FIG7.png": "'Figure 3: Depolymerization of Actin Filaments in the Presence of Collins Observed by Evanescent Wave Fluorescence Microscopy Conditions as in Figure 1. ATP-actin filaments were grown for 6 min from 1.5 \u03bcM 50% Oregon Green-labeled actin alone, or with 32 \u03bcM human coffin, or with 24 \u03bcM S, pombe coffin. These samples were washed with buffer alone or with 10 \u03bcM human coffin or with 10 \u03bcM S, pombe coffin, and observed. The data are presented in two ways: (A)\u2013(F), time course of length versus time, where dark gray lines are the global fit rates with dark dotted lines at 95% confidence intervals and light gray lines are the average individual filament rates with light dotted lines at minimum and maximum slopes and dashed lines indicating standard deviations, and (B)\u2013(L), histograms of the distribution of measured length changes during each 9 s interval. (A\u2013C) Barbed ends. (A) Actin filaments alone, (B) actin with 10 \u03bcM human coffin, and (C) actin with 10 \u03bcM S, pombe coffin. (D\u2013F), Pointed ends. (D) Actin filaments alone, (E) actin with 10 \u03bcM human coffin, and (F) actin with 10 \u03bcM S, pombe coffin. (G\u2013L) Barbed ends. (G) Actin filaments alone, (F) actin with 10 \u03bcM human coffin, and (I) actin with 10 \u03bcM S, pombe coffin. (J\u2013L) Pointed ends. (J) Actin filaments alone, (K) actin with 10 \u03bcM human coffin, and (L) actin with 10 \u03bcM S, pombe coffin.\\n\\n'", "CAPTION TAB1.png": "'\\n\\n\\\\begin{table}\\n\\\\begin{tabular}{l l} \\\\hline \\\\hline \\\\multicolumn{1}{c}{**Table 1.**} & \\\\multicolumn{1}{c}{**S**over**} \\\\\\\\ \\\\multicolumn{1}{c}{**and**} & \\\\multicolumn{1}{c}{**Optimal**} & \\\\multicolumn{1}{c}{**S**over**} \\\\\\\\ \\\\multicolumn{1}{c}{**and**} & \\\\multicolumn{1}{c}{**Optimal**} & \\\\multicolumn{'", "CAPTION TAB2.png": "'* [19]'", "SUPP CAPTION FIGS1.png": "'\\n\\n**Fig. S1.** X-ray crystal structure of S. pombe cofilin. (A) Stereo ribbon diagram of S. pombe cofilin with residues R78 and K80 highlighted, and sulfate ion bound to N-terminus. Residues 2-123 shown. (B) Model and electron density for residues surrounding the bound sulfate.\\n\\n'", "SUPP CAPTION FIGS2-1.png": "'\\n\\n**Fig. S2.** Inhibition of nucleotide exchange on actin monomer by S. pombe cofllin. Conditions: 10 mM imidazole, 1 mM MgCl\\\\({}_{2}\\\\), 1 mM EGTA, 0.5 mM DTT, 1 mM NaN\\\\({}_{3}\\\\) at 22 degC and pH 7.0. Actin concentrations were'", "SUPP CAPTION FIGS2-2.png": "'0.2 uM. Observed rate constant for etheno-ATP or etheno-ADP dissociation from actin monomer upon addition of 0.5 mM ATP or ADP plus S. pombe cofilin is plotted versus S. pombe cofilin concentration. Black circles, dashed line: ADP-actin. Open circles, dashed line: ATP-actin. Grey triangles, solid line: ADP-actin plus 50 mM KCl. Open triangles, solid line, ATP-actin plus 50 mM KCl.\\n\\n'", "SUPP CAPTION FIGS2.png": "''", "SUPP CAPTION FIGS3.png": "'\\n\\n**Fig. S3.** Time course of fluorescence change following the mixing of 0.2 \\\\(\\\\upmu\\\\)M S. pombe cofilin with 6 \\\\(\\\\upmu\\\\)M pyrene-labeled skeletal muscle ADP-actin filaments. The inset shows the initial rapid decrease in fluorescence on a faster time scale, fit to a single-exponential. Conditions: 10 mM imidazole, 2 mM Tris-HCl pH 8.0, 50 mM KCl, 1 mM MgCl\\\\({}_{2}\\\\), 1 mM EGTA, 0.2 mM ATP, 0.5 mM DTT, 1 mM NaN\\\\({}_{3}\\\\) at 22 \\\\({}^{\\\\circ}\\\\)C and pH 7.0'", "SUPP CAPTION FIGS4-1.png": "'Fig. S4. Raw data of 2 \\\\(\\\\mu\\\\)M pyrene actin elongating from actin nucleated in the presence and absence of human, S. pombe, and R78E K80E S. pombe cofilin mutant. Conditions: 10 mM imidazole, 2 mM Tris-HCl pH 8.0, 50 mM KCl, 1 mM MgCl2, 1 mM EGTA, 0.2 mM ATP, 0.5 mM DTT, 1 mM NaN3 at 22 \\\\({}^{\\\\circ}\\\\)C and pH 7.0. The slopes of the fluorescence traces indicate the number of ends formed under each condition: no seeds (heavy black line), 6 mM actin seeds (light black line), 6 mM actin/10 mM S. pombe cofilin seeds (light dotted line), 6 mM actin/10 mM Human cofilin'", "SUPP CAPTION FIGS4-2.png": "'seeds (grey line), 6 uM actin/10 uM R78E K80E S. pomhe cofilin mutant seeds (heavy dotted line).\\n\\n'", "SUPP CAPTION FIGS4.png": "'Figure 44: Raw data of 2 \\\\(\\\\mu\\\\)M prime actin elongating from actin nucleated in the presence and absence of human, S. pombe, and R79E K80E S. pombe for actin mutant seeds (heavy dotted line). 6 mM actin most seeds (light black line), 6 mM actin most seeds (light black line), 6 mM actin/10 mM S. pombe cofflin mutant. Conditions: 10 mM imidazole, 2 mM Tris-HCl pH 8.0, 50 mM KCl, 1 mM MgCl2, 1 mM EGTA, 0.2 mM ATP, 0.5 mM DTT, 1 mM NaN, at 22 \u00b0C and pH 7.0. The slopes of the fluorescence traces indicate the number of ends formed under each condition: no seeds (heavy black line), 6 mM actin seeds (light black line), 6 mM actin most seeds (light black line), 6 mM actin/10 mM S. pombe cofflin seeds (light dotted line), 6 mM actin/10 mM Human cofflin\\n\\n'", "SUPP CAPTION FIGS5.png": "'\\n\\n**Fig. S5.** Predicted spatial compartmentalization of ADF/cofilin activities.\\n\\n'", "SUPP CAPTION TABS1.png": "'\\n\\n**Table S1.** Data Collection and Refinement Statistics'"}