Cdc14 Early Anaphase Release, FEAR, Is Limited to the Nucleus and Dispensable for Efficient Mitotic Exit.

Christopher M Yellman,G Shirleen Roeder,Maurizio Del Poeta

doi:10.1371/journal.pone.0128604

Christopher M Yellman, G Shirleen Roeder + Show 1 more

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https://doi.org/10.1371/journal.pone.0128604

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Journal: PloS one	Publication Date: Jun 19, 2015
Citations: 20	License type: CC BY 4.0

Affiliation: Howard Hughes Medical Institute, Yale University

Abstract

Cdc14 phosphatase is a key regulator of exit from mitosis, acting primarily through antagonism of cyclin-dependent kinase, and is also thought to be important for meiosis. Cdc14 is released from its sequestration site in the nucleolus in two stages, first by the non-essential Cdc Fourteen Early Anaphase Release (FEAR) pathway and later by the essential Mitotic Exit Network (MEN), which drives efficient export of Cdc14 to the cytoplasm. We find that Cdc14 is confined to the nucleus during early mitotic anaphase release, and during its meiosis I release. Proteins whose degradation is directed by Cdc14 as a requirement for mitotic exit (e.g. the B-type cyclin, Clb2), remain stable during mitotic FEAR, a result consistent with Cdc14 being restricted to the nucleus and not participating directly in mitotic exit. Cdc14 released by the FEAR pathway has been proposed to have a wide variety of activities, all of which are thought to promote passage through anaphase. Proposed functions of FEAR include stabilization of anaphase spindles, resolution of the rDNA to allow its segregation, and priming of the MEN so that mitotic exit can occur promptly and efficiently. We tested the model for FEAR functions using the FEAR-deficient mutation net1-6cdk. Our cytological observations indicate that, contrary to the current model, FEAR is fully dispensable for timely progression through a series of anaphase landmarks and mitotic exit, although it is required for timely rDNA segregation. The net1-6cdk mutation suppresses temperature-sensitive mutations in MEN genes, suggesting that rather than activating mitotic exit, FEAR either inhibits the MEN or has no direct effect upon it. One interpretation of this result is that FEAR delays MEN activation to ensure that rDNA segregation occurs before mitotic exit. Our findings clarify the distinction between FEAR and MEN-dependent Cdc14 activities and will help guide emerging quantitative models of this cell cycle transition.

Highlights

Release of Cdc14 phosphatase from the nucleolus is a critical event during mitotic exit in the budding yeast Saccharomyces cerevisiae [1]
We find that Cdc14 released by the Fourteen Early Anaphase Release (FEAR) pathway is limited to the nucleus in both mitosis and meiosis I, and that even prolonged FEAR activation does not lead to the protein degradation characteristic of mitotic exit
Since Mitotic Exit Network (MEN)-activated release of Cdc14 can supplant FEAR, FEAR is usually examined in MEN-defective mutants that have been synchronized and released into the cell cycle

Summary

Introduction

Release of Cdc phosphatase from the nucleolus is a critical event during mitotic exit in the budding yeast Saccharomyces cerevisiae [1]. Cdc counteracts mitotic cyclin-dependent kinase (Cdk) activity by dephosphorylating and thereby activating three substrates: Sic, a direct Cdk inhibitor, Swi, a transcription factor for Sic, and Cdh, a substrate-specificity factor for the ubiquitin ligase APCCdh1 [2, 3]. APCCdh subsequently targets key mitotic proteins for degradation, including the major mitotic cyclin Clb and the mitotic kinase Cdc, permitting the transition to G1 phase. From G1 through metaphase, Cdc is bound by the protein Net in a nucleolar complex called RENT (Regulator of Nucleolar silencing and Telophase exit). Release of Cdc from the RENT complex and from the nucleolus is generally equated with activation [4,5,6], but at least one report suggests that Cdc can be active while bound to the RENT complex [7]

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