Figure 4.

Genetic evidence that ACA2-GFPp retains normal functional properties when expressed in yeast. The yeast mutant K616 was transformed with different constructs and tested for its ability to grow on synthetic medium supplemented with 10 mm CaCl₂ (control) or on synthetic medium supplemented with 10 mm EGTA. Growth of K616 harboring different constructs is shown. Both tagged and untagged versions of a deregulated pump (ACA2–8 and ACA2–2, respectively) were able to restore growth of the K616 mutant on EGTA. The constructs shown are: Vector, the parent vector pYX-112; GFP, pYX-GFP-JFH1, which encodes a GFP(S65/T) (J.F. Harper, unpublished data); ACA2–2, pYX-ACA2–2, which encodes a deregulated ACA2p (Harper et al., 1998); and ACA2–8, pYX-ACA2–8, which encodes the same deregulated ACA2–2p with an additional C-terminal GFP tag. The tagged and untagged full-length versions of ACA2p (respectively encoded by pYX-ACA2–7 and pYX-ACA2–1) did not complement and are not shown.