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. 2013 Oct 14;8(10):e76424. doi: 10.1371/journal.pone.0076424

Figure 6. cdc48 mutations can cause a Bur- phenotype.

Figure 6

A) A heterozygous CDC48–12xMYC::KAN integrant diploid (GY2476) was induced to undergo meiosis and tetrads dissected. Two representative tetrads are shown, with spores labeled A-D. Perfect linkage was observed between slow growth, G418-resistance, and the Bur- phenotype in 30 four-spored tetrads. B) A plasmid shuffle screen for cdc48 Bur- mutants uncovered an arginine-to-lysine change at amino acid 369, located in the ATPase D1 domain. This mutation confers both Bur- and cold-sensitive phenotypes (bottom panel). The strain being tested here contains the suc2Δuas-HIS3 reporter, with growth on SC-His plate being indicative of the Bur- phenotype. C) The cold sensitivity of cdc48-R369K is complemented by CDC48 on a CEN plasmid. Strains GY2387 (cdc48Δ::TRP1 <pAR54  =  CEN URA3 CDC48> and GY2406 (cdc48Δ::TRP1 <pAR64  =  CEN URA3 cdc48-R369K>) were transformed with empty vector or wild-type CDC48 on a LEU2-marked CEN plasmid (pAR56). Cold sensitivity was assayed at 16°C. D) The cdc48-R369K mutation does not affect the ability of Cdc48 to co-immunoprecipitate with Irc20.