Newly identified phosphorylation sites in Ras2p and Flo8p are required for wild-type yeast invasive growth.

<p>A) Previously unreported phosphorylation sites in Ras2p are shown in red with indicated probability that the sites were correctly identified. Site-directed mutagenesis was used to generate an integrated allele of <i>ras2</i> encoding Phe and Ala substitutions at Y165 and T166, respectively. The mutant exhibits diminished invasive growth (“-”) on YPD medium. Invasive growth was scored quantitatively as mean pixel intensity of the spotted area post-washing relative to the mean pixel intensity before washing. Error bars indicate the standard deviation from three independent trials. B) Novel phosphorylation sites in Flo8p are shown in red. A mutant encoding alanine at each residue (<i>flo8-</i>S3A) yields decreased invasive growth and decreased transcriptional activity of <i>lacZ</i> reporters responsive to Ste12/Tec1p regulation (downstream of the Kss1p MAPK pathway, pFRE-<i>lacZ</i>) and Flo8p-binding (PKA-regulated, <i>P</i>_flo11-6/7), respectively.