FIG. 6.
Inactivation of a temperature-sensitive allele of NUT2 allows Swi4p-independent expression. (A) Expression of the UASGAL-URS2R-lacZ reporter was assayed in wild-type (RT688; lanes 1 and 2), swi4Δ (RT689; lanes 3 and 4), swi4Δ nut1Δ nut2-1 (RT718; lanes 5 and 6), and swi4Δ NUT1 nut2-ts70 (RT748; lanes 7 and 8) strains. RNA was isolated from these strains after growth in YEP galactose liquid medium at 25°C (odd-numbered lanes) or 37°C (even-numbered lanes) for 3 h. RNA was analyzed by Northern blotting and hybridized with probes for lacZ, CLN2, PCL1, and TCM1. The latter serves as an RNA loading control. The TCM1 signal indicates that lanes 2, 3, and 4 contain slightly more RNA than other lanes. All strains are isogenic. (B) Expression of the SUC2 gene was assayed in wild-type (RT688; lanes 1 and 5), swi4Δ (RT689; lane 2), swi4Δ nut1Δ nut2-1 (RT718; lane 3), and SWI4 nut1Δ nut2-1 (RT720; lane 4) strains by Northern analysis. Strains were grown in dextrose (D; lanes 1 to 4) or in galactose (G; lane 5) medium. (C) STE2 expression was assayed in MATα cells (lanes 1 and 3) and MATa cells (lanes 2 and 4) that were either wild type (lanes 1 and 2) or nut1Δ nut2-1 mutants (lanes 3 and 4).