A Switch from a Gradient to a Threshold Mode in the Regulation of a Transcriptional Cascade Promotes Robust Execution of Meiosis in Budding Yeast

Tight regulation of developmental pathways is of critical importance to all organisms, and is achieved by a transcriptional cascade ensuring the coordinated expression of sets of genes. We aimed to explore whether a strong signal is required to enter and complete a developmental pathway, by using meiosis in budding yeast as a model. We demonstrate that meiosis in budding yeast is insensitive to drastic changes in the levels of its consecutive positive regulators (Ime1, Ime2, and Ndt80). Entry into DNA replication is not correlated with the time of transcription of the early genes that regulate this event. Entry into nuclear division is directly regulated by the time of transcription of the middle genes, as premature transcription of their activator NDT80, leads to a premature entry into the first meiotic division, and loss of coordination between DNA replication and nuclear division. We demonstrate that Cdk1/Cln3 functions as a negative regulator of Ime2, and that ectopic expression of Cln3 delays entry into nuclear division as well as NDT80 transcription. Because Ime2 functions as a positive regulator for premeiotic DNA replication and NDT80 transcription, as well as a negative regulator of Cdk/Cln, we suggest that a double negative feedback loop between Ime2 and Cdk1/Cln3 promotes a bistable switch from the cell cycle to meiosis. Moreover, our results suggest a regulatory mode switch that ensures robust meiosis as the transcription of the early meiosis-specific genes responds in a graded mode to Ime1 levels, whereas that of the middle and late genes as well as initiation of DNA replication, are regulated in a threshold mode.

[1]  Chao Zhang,et al.  Control of landmark events in meiosis by the CDK Cdc28 and the meiosis-specific kinase Ime2. , 2003, Genes & development.

[2]  Sheetal Raithatha,et al.  The Saccharomyces cerevisiae CLB5 promoter contains two middle sporulation elements (MSEs) that are differentially regulated during sporulation , 2008, Yeast.

[3]  L. Dirick,et al.  Regulation of meiotic S phase by Ime2 and a Clb5,6-associated kinase in Saccharomyces cerevisiae. , 1998, Science.

[4]  Naama Barkai,et al.  Threshold responses to morphogen gradients by zero-order ultrasensitivity , 2005, Molecular systems biology.

[5]  Y. Kassir,et al.  Monitoring meiosis and sporulation in Saccharomyces cerevisiae. , 1991, Methods in enzymology.

[6]  D. Botstein,et al.  The transcriptional program of sporulation in budding yeast. , 1998, Science.

[7]  Y. Kassir,et al.  Positive and negative feedback loops affect the transcription of IME1, a positive regulator of meiosis in Saccharomyces cerevisiae. , 1995, Developmental genetics.

[8]  Aviv Regev,et al.  Dissecting Timing Variability in Yeast Meiosis , 2007, Cell.

[9]  A. Yoshimura,et al.  SOCS1 Deficiency Causes a Lymphocyte-Dependent Perinatal Lethality , 1999, Cell.

[10]  Y. Kassir,et al.  Ime2, a Meiosis-Specific Kinase in Yeast, Is Required for Destabilization of Its Transcriptional Activator, Ime1 , 2002, Molecular and Cellular Biology.

[11]  Naama Barkai,et al.  Elucidating mechanisms underlying robustness of morphogen gradients. , 2004, Current opinion in genetics & development.

[12]  Y. Kassir,et al.  Human Cdk2 is a functional homolog of budding yeast Ime2, the meiosis-specific Cdk-like kinase , 2009, Cell cycle.

[13]  G. Braus,et al.  Inhibition of APC-mediated proteolysis by the meiosis-specific protein kinase Ime2 , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[14]  A. Varma,et al.  Partial deprivation of GTP initiates meiosis and sporulation in Saccharomyces cerevisiae , 2004, Molecular and General Genetics MGG.

[15]  Donna K Slonim,et al.  The homeodomain protein PAL-1 specifies a lineage-specific regulatory network in the C. elegans embryo , 2005, Development.

[16]  D. Koshland,et al.  An amplified sensitivity arising from covalent modification in biological systems. , 1981, Proceedings of the National Academy of Sciences of the United States of America.

[17]  E. Davidson,et al.  Transcriptional regulatory cascades in development: Initial rates, not steady state, determine network kinetics , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[18]  Jing Chen,et al.  The Goldbeter-Koshland Switch in the First-Order Region and Its Response to Dynamic Disorder , 2008, PloS one.

[19]  K. Benjamin,et al.  The Cdk-Activating Kinase Cak1p Promotes Meiotic S Phase through Ime2p , 2003, Molecular and Cellular Biology.

[20]  J. Ferrell.  Self-perpetuating states in signal transduction: positive feedback, double-negative feedback and bistability. , 2002, Current opinion in cell biology.

[21]  Martin Kupiec,et al.  11 Meiosis and Sporulation in Saccharomyces cerevisiae , 1997 .

[22]  K. Struhl,et al.  Histone deacetylase activity of Rpd3 is important for transcriptional repression in vivo. , 1998, Genes & development.

[23]  Gerald R. Fink,et al.  Methods in Yeast Genetics: A Laboratory Course Manual , 1987 .

[24]  K. Benjamin,et al.  Sum1 and Ndt80 Proteins Compete for Binding to Middle Sporulation Element Sequences That Control Meiotic Gene Expression , 2003, Molecular and Cellular Biology.

[25]  Ronald W. Davis,et al.  The core meiotic transcriptome in budding yeasts , 2000, Nature Genetics.

[26]  K. Struhl,et al.  Repression by Ume6 Involves Recruitment of a Complex Containing Sin3 Corepressor and Rpd3 Histone Deacetylase to Target Promoters , 1997, Cell.

[27]  A. Vershon,et al.  Sum1 and Hst1 repress middle sporulation‐specific gene expression during mitosis in Saccharomyces cerevisiae , 1999, The EMBO journal.

[28]  G. Fink,et al.  Methods in yeast genetics , 1979 .

[29]  Amir Rubinstein,et al.  Faithful modeling of transient expression and its application to elucidating negative feedback regulation , 2007, Proceedings of the National Academy of Sciences.

[30]  Y. Kassir,et al.  A meiosis-specific protein kinase, Ime2, is required for the correct timing of DNA replication and for spore formation in yeast meiosis , 1996, Molecular and General Genetics MGG.

[31]  George M Church,et al.  The Isw2 Chromatin Remodeling Complex Represses Early Meiotic Genes upon Recruitment by Ume6p , 2000, Cell.

[32]  E. Garí,et al.  Recruitment of Cdc28 by Whi3 restricts nuclear accumulation of the G1 cyclin–Cdk complex to late G1 , 2004, The EMBO journal.

[33]  Sheetal Raithatha,et al.  Phosphorylation and Maximal Activity of Saccharomyces cerevisiae Meiosis-Specific Transcription Factor Ndt80 Is Dependent on Ime2 , 2002, Molecular and Cellular Biology.

[34]  H. Friesen,et al.  NDT80 and the Meiotic Recombination Checkpoint Regulate Expression of Middle Sporulation-Specific Genes in Saccharomyces cerevisiae , 1998, Molecular and Cellular Biology.

[35]  J. Pak,et al.  Role of Ndt80, Sum1, and Swe1 as Targets of the Meiotic Recombination Checkpoint That Control Exit from Pachytene and Spore Formation in Saccharomyces cerevisiae , 2002, Molecular and Cellular Biology.

[36]  G. Roeder,et al.  The pachytene checkpoint. , 2000, Trends in genetics : TIG.

[37]  Enrique Herrero,et al.  G1 cyclins block the Ime1 pathway to make mitosis and meiosis incompatible in budding yeast , 1999, The EMBO journal.

[38]  J. Pak,et al.  Regulation of the Premiddle and Middle Phases of Expression of the NDT80 Gene during Sporulation of Saccharomyces cerevisiae , 2002, Molecular and Cellular Biology.

[39]  S. Chu,et al.  Gametogenesis in yeast is regulated by a transcriptional cascade dependent on Ndt80. , 1998, Molecular cell.

[40]  Galit Shenhar,et al.  A Positive Regulator of Mitosis, Sok2, Functions as a Negative Regulator of Meiosis in Saccharomyces cerevisiae , 2001, Molecular and Cellular Biology.

[41]  K. Nasmyth.  At the heart of the budding yeast cell cycle. , 1996, Trends in genetics : TIG.

[42]  Y. Kassir,et al.  Cdc28 and Ime2 possess redundant functions in promoting entry into premeiotic DNA replication in Saccharomyces cerevisiae. , 2001, Genetics.

[43]  Fred Winston,et al.  Methods in Yeast Genetics: A Laboratory Course Manual , 1990 .

[44]  C. Wittenberg,et al.  CLB5 and CLB6 are required for premeiotic DNA replication and activation of the meiotic S/M checkpoint. , 1998, Genes & development.

[45]  D. Stuart,et al.  Meiosis-Specific Regulation of the Saccharomyces cerevisiae S-Phase Cyclin CLB5 Is Dependent on MluI Cell Cycle Box (MCB) Elements in Its Promoter but Is Independent of MCB-Binding Factor Activity , 2005, Genetics.

[46]  J. Collins,et al.  Construction of a genetic toggle switch in Escherichia coli , 2000, Nature.

[47]  E. Garí,et al.  The Cln3 cyclin is down‐regulated by translational repression and degradation during the G1 arrest caused by nitrogen deprivation in budding yeast , 1997, The EMBO journal.

[48]  Karen Schindler,et al.  Phosphorylation of Ime2 Regulates Meiotic Progression in Saccharomyces cerevisiae* , 2006, Journal of Biological Chemistry.

[49]  M. Freeman.  Feedback control of intercellular signalling in development , 2000, Nature.

[50]  G. Roeder,et al.  The pachytene checkpoint in S. cerevisiae depends on Swe1-mediated phosphorylation of the cyclin-dependent kinase Cdc28. , 1999, Molecular cell.

[51]  Galit Shenhar,et al.  Transcriptional regulation of meiosis in budding yeast. , 2003, International review of cytology.