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. 2012 Oct 1;23(19):3814–3826. doi: 10.1091/mbc.E12-05-0400

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© 2012 Chen et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 7: — The Gic2p N-terminus can replace the Bni1p regulatory regions. (A) The Gic2p N-terminal Cdc42p-binding domain is sufficient for Bni1p regulation in rewired cells (DLY12621, 12626). Function of the indicated constructs was tested as in Figure 3A. (B) 3HA-tagged Bni1p and the GIC2^N-FH1-FH2 construct are expressed at comparable levels. Blot probed with anti-HA and anti-Cdc11p (loading control). (C) GIC2^N-FH1FH2 can function as the sole formin in rewired cells. Spore viability of cells with the indicated genotypes was deduced from tetrad dissection of heterozygous diploids (DLY 12848, 12594, 12744). (D) DIC images of rewired cells expressing full-length Bni1p (left; DLY12888) or Gic2^N-FH1-FH2 (middle, right; DLY12892, 12887) in place of endogenous Bni1p (left, middle) or both Bni1p and Bnr1p (right). (E) GFP-Gic2^N-FH1-FH2 (DLY13430) localizes to the bud tip (top) and to the polarization site (marked with Bem1p-tdTomato) in the absence of F-actin (bottom). (F) F-actin distribution in cells containing the indicated construct as the sole formin (DLY13703, 13446, 13444, 13452). (G) GFP-Sec4p (vesicle marker) localization in the same strains as in F. (H) GFP-Sec4p localization categories (cartoon) were scored in small- and medium-budded cells of the strains in F and G. Scale bar, 5 μm.