FIGURE 2.

Orm2 protein level is slightly increased by calcofluor white-induced cell wall stress. Cells were grown overnight to mid-log phase in SC-uracil medium at 30 °C and then shifted to YPD for 6 h before treatment with tunicamycin (1 μg/ml), DTT (1 mm), or calcofluor white (40 μg/ml) for an additional 2 h. Lysates were assayed for both Orm2-TAP levels and activation of cell wall stress signaling. A, top, Western blot for Orm2-TAP in cells treated with or without tunicamycin (T), DTT (D), or calcofluor white (C). Bottom, phosphoglycerate kinase (Pgk) blotted as a loading control. B, β-galactosidase activity (μmol/min/mg) of an MPK1-lacZ reporter to quantitate cell wall stress signaling. Cell wall stress was maximally induced by calcofluor white. C, diagram showing that ER stress-induced Orm2 increase is dependent on calcineurin activation, whereas cell wall stress-induced Orm2 increase is signaled through Mpk1 to calcineurin activation.