Extended Data Fig. 5: Stp-mCherry fusion proteins are functional and localize to speckles surrounding fungal hyphae after plant colonization.
From: A cell surface-exposed protein complex with an essential virulence function in Ustilago maydis
a, Seven-day-old maize seedlings were infected with stp deletion strains complemented with the respective mCherry fusion alleles. For stp5 complementation strains with single (s) as well as two (d) copies of the construct are shown. Dashed lines separate independently tested groups of strains. The rightmost panel presents infections with strain SG200stp1-YFP stp3-mCherry, where both fusion proteins are expressed from genes in their native loci. 12 d.p.i. disease symptoms were scored using the scores depicted below. Data represent mean of n = 3 biologically independent experiments. Total numbers of infected plants are indicated above the respective columns. b, Anti-HA western blot of the total extract obtained from leaves 3 d.p.i. with strain SG200Δstp5 HAmCherry-stp5 (s). c, d, Anti-HA western blot after HA-specific immunoprecipitations from total maize leaf extracts prepared 3 d.p.i. with the indicated U. maydis strains. b-d, Signals corresponding to full-length fusion proteins are marked with an asterisk. e, Hyphal tips of SG200 strains expressing the indicated mCherry fusion proteins in the plant epidermis observed 2 d.p.i. mCherry signal: red. The images represent maximum projections of confocal z-stacks. Scale bars, 5 µm. f, Constitutive expression of all seven complex members changes surface topology of hyphae. Scanning electron micrographs (SEM) of AB33 and AB33Δ6complex show smooth hyphae with single scattered structures on the surface. In AB33complexcon a large number of small surface exposed structures are seen in addition. Scale bars, 500 nm.
