Asian Program

Japanese

Data

Former Fellows

Dissertation Abstracts

Philippines

  Calcineurin, a protein phosphatase required for Ca²⁺ signaling in many cell types, is a heterodimer composed of catalytic and regulatory subunits. The fission yeast genome encodes a single set of catalytic (Ppb1) and regulatory (Cnb1) subunits, providing an ideal model system to study these subunits' functions in vivo. Here, we cloned the cnb1⁺ gene, and showed that cnb1 knockout (Δcnb1) exhibits identical phenotypes with Δppb1, and that overexpression of Ppb1 failed to suppress the phenotypes of Δcnb1. Interestingly, overexpression of C-terminal-deleted Ppb1 (Ppb1ΔC), the constitutively active form of Ppb1, also failed to suppress the phenotypes of Δcnb1. FK506 caused MgCl2 sensitivity to the wild-type cells in an FKBP12-dependent manner. Co-overexpression of Ppb1 and Cnb1 suppressed the FK506-induced MgCl2 sensitivity, but the suppression was only partial, suggesting that an excess amount of Ppb1/Cnb1 complex cannot titrate out the FKBP12/FK506 complex. Although overexpression of Ppb1ΔC alone had little effect on cell growth, co-overexpression of Ppb1ΔC and Cnb1 caused distinct growth defect. FK506 suppressed the growth defect when Cnb1 was co-expressed using attenuated nmt1 promoter, but it failed to suppress the defect when Cnb1 was co-expressed using wild-type nmt1 promoter. Knockout of prz1+ gene, encoding a downstream target transcription factor of calcineurin, suppressed the growth defect irrespective of the promoter potency. These results suggest that Cnb1 is essential for the activation of calcineurin, and that the activated calcineurin is the pharmacological target of the FKBP12/FK506 complex in vivo.