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Calcineurin Regulatory Subunit Calcium-Binding Domains Differentially Contribute to Calcineurin Signaling in Saccharomyces cerevisiae.

Abstract：

:The protein phosphatase calcineurin is central to Ca2+ signaling pathways from yeast to humans. Full activation of calcineurin requires Ca2+ binding to the regulatory subunit CNB, comprised of four Ca2+-binding EF hand domains, and recruitment of Ca2+-calmodulin. Here we report the consequences of disrupting Ca2+ binding to individual Cnb1 EF hand domains on calcineurin function in Saccharomyces cerevisiae Calcineurin activity was monitored via quantitation of the calcineurin-dependent reporter gene, CDRE-lacZ, and calcineurin-dependent growth under conditions of environmental stress. Mutation of EF2 dramatically reduced CDRE-lacZ expression and failed to support calcineurin-dependent growth. In contrast, Ca2+ binding to EF4 was largely dispensable for calcineurin function. Mutation of EF1 and EF3 exerted intermediate phenotypes. Reduced activity of EF1, EF2, or EF3 mutant calcineurin was also observed in yeast lacking functional calmodulin and could not be rescued by expression of a truncated catalytic subunit lacking the C-terminal autoinhibitory domain either alone or in conjunction with the calmodulin binding and autoinhibitory segment domains. Ca2+ binding to EF1, EF2, and EF3 in response to intracellular Ca2+ signals therefore has functions in phosphatase activation beyond calmodulin recruitment and displacement of known autoinhibitory domains. Disruption of Ca2+ binding to EF1, EF2, or EF3 reduced Ca2+ responsiveness of calcineurin, but increased the sensitivity of calcineurin to immunophilin-immunosuppressant inhibition. Mutation of EF2 also increased the susceptibility of calcineurin to hydrogen peroxide inactivation. Our observations indicate that distinct Cnb1 EF hand domains differentially affect calcineurin function in vivo, and that EF4 is not essential despite conservation across taxa.

journal_name

Genetics

journal_title

Genetics

authors

Connolly S,Quasi-Woode D,Waldron L,Eberly C,Waters K,Muller EM,Kingsbury TJ

doi

10.1534/genetics.118.300911

subject

Has Abstract

pub_date

2018-07-01 00:00:00

pages

801-813

issue

eissn

0016-6731

issn

1943-2631

pii

genetics.118.300911

journal_volume

209

pub_type

杂志文章

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Calcineurin Regulatory Subunit Calcium-Binding Domains Differentially Contribute to Calcineurin Signaling in Saccharomyces cerevisiae.