Supplemental Material

This article contains the following supporting material:

• Figure 1 - Design of mutations used in this study.
  Alignment of Aip1 primary sequences from S. cerevisiae (P46680), H. Sapiens (AF020056), M. musculus (AF020055), X. laevis (AF124140), A. thaliana (AAD14533), S. pombe (O14301), and C.elegans (Q11176). Arrows mark ·-strand secondary structure in S. cerevisiae (top) and C.elegans Aip1 (bottom). Highly conserved amino acids are shaded, and mutated residues in each aip1 allele are marked.

• Figure 2 - (A) Wild type (AIP1) and aip1· cells were fixed and co-stained with Alexa-488 phalloidin and anti-cofilin antibodies. Cofilin-decorated actin cables in aip1· cells were detected by phalloidin. Arrowhead points to actin cables.

• Figure 3 - Growth defects of aip1 cof1-22 double mutant strains.
  Haploid strains with the cof1-22 mutation combined with different aip1 mutations were compared for growth by two assays. (A) Cells were grown to log phase, then serially diluted, plated on YPD, and grown for 2 days at 25 °C. (B) Cells were streaked on YPD plates and grown for 2 days at 25 °C.

• Figure 4 - Defects in actin patch turnover in aip1 mutant cells.
  (A) Wild type (AIP1) and mutant (aip1·, aip1-108, aip1-109, aip1-119) yeast cells were treated with 50 μM Latrunculin A (Lat-A). Samples of cells were removed at the indicated time points, fixed, and stained with alexa-488 phalloidin. (B) Cells from (A) were scored for visible actin patches and graphed. n >200 cells were counted for each time point and the columns are the average of two independent experiments. Error bars, standard deviation.

• Figure 5 - Comparison of actin binding surfaces defined by different approaches.
  Two actin-binding surfaces defined by this study are colored red and yellow, and surfaces defined in very recent study (Clark et al. 2006) are labeled in blue.

• Table 1
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• Table 3