Fig 5.

Kch1 and Kch2 promote K⁺ uptake and K⁺-dependent activation of HACS. (A) A trk1 trk2 double mutant and a trk1 trk2 mid1 triple mutant (CS30 and CS163) bearing a control plasmid or plasmids overexpressing KCH1-HA₃ or KCH2-HA₃ genes (pCS42, pCS43, and pCS44) were serially diluted and then spotted onto SDAP pH 4.5 agar medium lacking methionine but containing 100 or 50 mM KCl as indicated. Colonies were photographed after 3 days incubation at 30°C. (B) ⁴⁵Ca²⁺ uptake into log-phase cultures of kch1 kch2 double mutants and cch1 kch1 kch2 triple mutants (CS03 and CS08) transformed with control plasmid or plasmids overexpressing KCH1-HA₃ and KCH2-HA₃ genes, in SC-100 medium supplemented with 1 μg of FK506/ml and incubated for 1.5 h. (C) ⁴⁵Ca²⁺ uptake into the kch1 kch2 double mutants expressing KCH1-HA₃ and KCH2-HA₃ genes was performed in SC-100 media that had been supplemented with 0.2 μg of FK506/ml plus the indicated concentrations of KCl, RbCl, or NaCl. The data points were normalized to corresponding plasmid controls to represent Kch1- and Kch2-dependent calcium uptake. Plots depict averages (± the SEM).