Figure 2.

Residues 10–27 and 28–52 are important for distinct trafficking steps. (A) Multiple sequence alignment of several Chs3 homologs. Residues are shaded to highlight similarity (gray) or identity (black). (B) The indicated plasmids were transformed into both a chs3ΔSEC7-Mars strain and a chs3ΔSEC7-Mars apl2Δ strain and plated on indicated media. Plates were imaged after 2 days at 30°C. See also Figure S2 for additional CW concentrations. (C) Protein expression level of mutated Chs3-GFP constructs relative to wild type as calculated from the α-GFP Western blots in Figure S1, normalized to loading control. (D) Chs3-GFP and Sec7-Mars localization in chs3Δ cells for WT Chs3-GFP and Chs3(Δ10–27)-GFP, at equivalent light levels. Scale bar, 1 μm. (E) Line profile of intensity along dotted lines in (D) of unbudded cells. Asterisks mark higher localization of Chs3(Δ10–27)-GFP to the plasma membrane at cell boundaries. (F) Chs3(Δ28–52)-GFP localization examined as in (C). Additional “Bright GFP” image with light levels scaled to improve visibility is presented. (G) Line profile as in (E) for small-budded cells. Asterisk indicates lower bud neck localization of Chs3(Δ28–52)-GFP.