Figure 5.

Cell wall degradation enzymes focalize at the fusion focus. (A) Crosses of heterothallic h⁻ and h⁺ myo52-tdTomato strains expressing GFP-tagged glucanases as indicated. All glucanases localize to the fusion focus. (B) Fusion efficiency of crosses between h⁺ fus1Δ strains and h⁻ single, double, and triple glucanase deletion strains. n > 100. (C) Calcofluor images of nonfusing pairs in heterothallic crosses of h⁺ fus1Δ, myoVΔ, and agn2Δ eng2Δ to h⁻ fus1Δ, myoVΔ, and agn2Δ eng2Δ, as indicated. This shows presence of cell wall at the cell–cell contact. (D) Time-averaged projections over 10 s of crosses of heterothallic h⁻ bgs1-GFP (top) and bgs4-GFP (bottom) to h⁺ myo52-tdTomato, imaged every second. Cell wall synthases localize as a crescent at the fusion site. (E) Homothallic h90 wild type (left) and fus1Δ (right) myo52-tdTomato strains coexpressing sfGFP-tagged versions of Agn2, Eng2, and Exg3. Cell wall glucanases colocalize at fusion site with Myo52 either as a dot in wild type or as a crescent in fusion-deficient fus1Δ cells. Cell outlines are shown with dotted lines. (F) Homothallic h90 myo51Δ myo52Δ (myoVΔ) strains coexpressing sfGFP-tagged versions of Agn2, Eng2, and Exg3. Glucanases are not detected at fusion site. (G) Heterothallic h⁺ myo51^Δtail-12Myc (left) and h⁺ myo52^Δtail-tdTomato (right) strains coexpressing sfGFP-tagged versions of Agn2, Eng2, and Exg3 crossed to h⁻ myo52-tdTomato cells. Cell wall glucanases localize at the fusion site in myo51^Δtail mutants but not, or are highly reduced, in myo52^Δtail. WT, wild type. Error bars are standard deviations. Cell outlines are shown with dotted lines. Bars, 1 µm.