Figure 2. Sexual differentiation in PKA mutants after Tor2 inactivation.

(A and B) h⁹⁰ tor2-51, h⁹⁰ tor2-51 pka1Δ and h⁹⁰ tor2-51 cgs1Δ cells were incubated for 48 hours at 32°C in the presence of nitrogen (YES medium). The percentage of zygotes was determined and represented as a plot (A) and spores were stained with iodine vapour (B). h⁹⁰ tor2-51 pka1Δ cells showed a higher mating efficiency than h⁹⁰ tor2-51 cells after Tor2 inactivation. In contrast, h⁹⁰ tor2-51 cgs1Δ cells were unable to mate after Tor2 inactivation. (C) tor2-51, tor2-51 pka1Δ and tor2-51 cgs1Δ cells were grown to mid-exponential phase in minimal medium, washed several times and incubated in minimal medium lacking nitrogen. Samples were collected at the indicated times to extract RNA and Northern blot was performed and hybridized with a probe against the ste11+ gene. rRNA stained with methylene blue was used as a loading control. In tor2-51 pka1Δ cells, ste11 expression levels increased earlier and to a higher level than in tor2-51. In the tor2-51 cgs1Δ double mutant there was no relevant activation of ste11+ transcription after Tor2 inactivation.