ros1 overexpression inhibits b-induced filamentation and triggers septation and nuclear divisions.

(A) Strain AB33_Pcrg1Ros1 expressing bE1 and bW2 from the nitrate-inducible nar1 promoter and ros1 from the arabinose-inducible crg1 promoter was grown in CM supplemented with glucose to an OD₆₀₀ of 0.6 and then shifted to NM medium containing either glucose (left panel) or arabinose (right panel). In NM + glucose only the b genes are induced whereas in NM + arabinose expression of the b genes and ros1 is induced. Samples were collected 6 h after induction. Nuclei and cell walls were stained with DAPI and calcofluor, respectively, and the morphology of the cells was assessed by DIC and fluorescence microscopy. In the presence of nitrate and glucose a functional bE1/bW2 heterodimer is produced and cells switch to filaments. In NM + arabinose, bE1, bW2 and ros1 are expressed simultaneously. In this case, cells fail to form filaments and become swollen and septated with one nucleus in each section. Bar = 10 μM. (B) AB33_Pcrg1Ros1 was pre-incubated for 6 h in NM + glucose to allow cells to filament and then transferred to either NM + glucose (left panel) or NM + arabinose (right panel) for another 6 h. In NM + glucose the cells remained filamentous and contained one nucleus (red arrow). In NM+ arabinose, when ros1 is induced, filaments became septated (yellow arrows) and each section contained one nucleus (red arrows). Nuclei and cell walls were stained as in A. Bar = 10 μm.