Cooperation of Calcineurin and Vacuolar H-ATPase in Intracellular CaHomeostasis of Yeast Cells

doi:10.1074/jbc.270.17.10113

Cooperation of Calcineurin and Vacuolar H-ATPase in Intracellular CaHomeostasis of Yeast Cells (*)

From the (1) Department of Plant Sciences, Graduate School of Science, University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113, Japan,
(2) Tonen Corporation, Corporate Research and Development Laboratory, 1-3-1, Nishi-tsurugaoka, Iruma-gun, Ohi-machi, Saitama 356, Japan, and the
(3) Division of Cell Proliferation, National Institute for Basic Biology, 38 Nishigonaka, Myodaijicho, Okazaki 444, Japan

^§ To whom correspondence should be addressed. Tel.: 81-3-3812-2111 (ext. 4461); Fax: 81-3-3812-4929.

Abstract

Saccharomyces cerevisiae VMA genes, encoding essential components for the expression of vacuolar membrane H-ATPase activity, are involved in intracellular ionic homeostasis and vacuolar biogenesis. We report here that the immunosuppressants FK506 and cyclosporin A cause general growth inhibition of the vma3 mutant. Upon addition of the drugs, the mutant grew neither in the presence of more than 5 mM Canor above pH 6.0. The action of the immunosuppressants is dependent on their binding proteins and ascribable to inhibition of calcineurin activity; a mutation of a calcineurin subunit (cnb1) shows synthetic lethal interaction with the vma mutation. The addition of FK506 decreases the cytosolic free concentration of Cain the vma3 mutant cells. Consequently, FK506 induces an 8.9-fold elevation of a nonexchangeable Capool. These results suggest that calcineurin controls calcium homeostasis by repression of Caflux into a cellular compartment(s) and that the vacuolar H-ATPase is essential for cell growth cooperating with calcineurin to regulate the cytosolic free concentration of Ca.

Footnotes

↵* This work was supported in part by Grants-in-aid for Scientific Research on Priority Areas 06740565 and 04266103 (to Y. O. and Y. A.) from the Ministry of Education, Science, and Culture of Japan and a grant from the International Human Frontier Science Program Organization (to Y. A.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

[Ca]

cytosolic free Caconcentration

PCR

polymerase chain reaction

FKBP-12

FK506 binding protein

Cyp-18

cyclophilin A

FOA

5-fluoroorotic acid

kb

kilobase(s)

MES

4-morpholineethanesulfonic acid.
↵² Y. Takita, Y. Ohya, and Y. Anraku, unpublished observations.