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. 2009 Jun 15;20(12):2820–2830. doi: 10.1091/mbc.E09-01-0069

Figure 1.

Figure 1.

Identification and characterization of the Gpa1-Kar3 interaction. (A) Identification of Kar3 as a Gpa1 interactor in pheromone-treated cells. Proteins that copurify with GST-Gpa1 were run on Immobulin two-dimensional gels. The thin white line indicates the positions of GST-Gpa1, which runs as multiple spots, and the white arrows indicate the two spots identified as Kar3 by peptide mass fingerprinting. The gel fragments encompass the region from molecular weight 60–100 kDa and pI of 7–9. Left, input. Middle, GST-Gpa1 pull-down from untreated cells. Right, GST-Gpa1 pull-down from pheromone-treated cells. (B) Reciprocal copurification of Gpa1 and Kar3 from yeast lysates. Left, pull-down of Kar3-HA with GST-Gpa1. Right, immunoprecipitation of Gpa1 with Kar3-HA. (C) Pheromone-stimulation of the Gpa1–Kar3 interaction does not depend on the induction of KAR3 transcription. Pull-down of Kar3-GST expressed from the CUP1 promoter. (D) The Gpa1–Kar3 interaction does not depend on polymerized microtubules. Left, pull-down of Kar3-HA with GST-Gpa1. Right, immunoprecipitation of Gpa1 with Kar3-HA.