Fig 4. Growth complementation of ammonium uptake-defective yeast strain by mutants of AtTIP2;1.

The 31019b yeast strain (Δmep1–3) was transformed with the empty vector pYeDP60u or with pYeDP60u-carrying cDNA encoding the positive controls HsAQP8 or HsAQP1, or AtTIP2;1 or its mutants. The five amino acid residues of the extended selectivity filter in each construct are indicated in one letter code to the left in the order H2^P, LC^P, H5^P, LE^P, and HE^P, showing substitutions in bold. Complementation and failure to complement are indicated by + and −, respectively. Transformants were spotted at an OD₆₀₀ of 1 (right column) and 0.01 (left column) on plates containing 0.2% proline or the indicated concentrations of ammonium as a sole nitrogen source and growth was recorded after 13 d at 28°C. Each panel showing growth at a specific concentration is compiled by individual pictures of each spot taken from a distinct growth condition. All single pictures were treated in the same manner.