C-terminus of Inn1 interacts with Chs2 (original) (raw)

2016

Abstract

(A) Truncated allele of Chs2 lacking transmembrane domain (Chs2-1-629) and fragment containing the tail of Chs2 (1–215) interact in a yeast two-hybrid assay with C-terminus of Inn1 (134–409). (B) Control cells (YAD382) and INN1-C-terminus-TAP cells (YMF82) were grown at 24°C in YPD medium, arrested in G1 phase and released for 105 minutes. Inn1 C-terminus-TAP was immunoprecipitated from cell extracts on IgG beads before the detection of the indicated proteins by immunoblotting. (C) C-terminus of Inn1 inhibits the catalytic activity of Chs2. The protein levels of overexpressed Chs2 and C-terminus (i) and percentage of active chitin synthase (ii) in control and cells lacking Chs3 and overexpressing either GAL-CHS2 (YMF687) or GAL-CHS2 GAL-Cterminus (YMF673) were determined in membranes isolated from asynchronous cultures (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005864#sec011" target="_blank">Materials and Methods</a>). (D) The protein levels of overexpressed Chs2 and Inn1 (i) and percentage of active chitin synthase (ii) in control and cells lacking Chs3 and overexpressing either GAL-CHS2 (YMF687) or GAL-CHS2 GAL-INN1 (YMF561) were determined in membranes isolated from asynchronous cultures (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005864#sec011" target="_blank">Materials and Methods</a>).</p

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