CrSTR is targeted to the vacuole of C. roseus cells by an N-terminal signal peptide and a consecutive vacuolar sorting tetrapeptide (SPIL). Undifferentiated C. roseus cells were transformed to express CrSTR-GFP or deletion/fusion GFP constructs as indicated in the 1st column. Calcofluor and CFP-GUS were used to visualise the cell wall and the cytoplasm, respectively (2nd column). The merged image and the DIC morphology are presented in the 3rd and 4th columns, respectively. Note that CrSTR-GFP was localised to the vacuole and that the N-terminus signal peptide was necessary and sufficient to target the fusion protein to the secretory pathway whereas the tetrapetide SPIL was necessary and sufficient to further sort to the vacuole the fusion protein that entered the secretory pathway. sp(SPIL), N-terminus CrSTR signal peptide and SPIL vacuolar sorting tetrapeptide; spmut(SPGL), N-terminus CrSTR signal peptide and mutated SPGL vacuolar sorting tetrapeptide; Δsp(SPIL)STR, CrSTR deleted from sp(SPIL). Bar: 10 μm.