TY - JOUR
T1 - Sfl1 functions via the co-repressor Ssn6-Tup1 and the cAMP-dependent protein kinase Tpk2
AU - Conlan, R. Steven
AU - Tzamarias, Dimitris
N1 - Funding Information:
We thank S. Kuhara, L. Robertson and G. Fink for yeast strains, and S. Alexaki for plasmid construction. This work was supported by grants to D.T. from the Greek Ministry of Development and the EU-TMR network programme, and to R.S.C. from The Nuffield Foundation (NUF-NAL 00).
Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2001/6/22
Y1 - 2001/6/22
N2 - Ssn6 (Cyc8) is a component of the yeast general corepressor Ssn6-Tup1 that inhibits the transcription of many diversely regulated genes. The corepressor does not interact directly with DNA but is recruited to different promoters through interactions with distinct pathway-specific, DNA-binding repressor proteins. Using yeast two-hybrid and GST chromatography interaction experiments, we have determined that Sfl1, a novel repressor protein, interacts directly with Ssn6, and in vivo repression data suggest that Sfl1 inhibits transcription by recruiting Ssn6-Tup1 via a specific domain in the Sfl1 protein. Sin4 and Srb10, components of specific RNA polymerase II sub-complexes that are required for Ssn6-Tup1 repression activity, are found to be required for Sfl1 repression function. These results indicate a possible mechanism for Sfl1-mediated repression via Ssn6-Tup1 and specific subunits of the RNA polymerase II holoenzyme. Electrophoretic mobility shift and chromatin immunoprecipitation assays demonstrate that Sfl1 is present at the promoters of three Ssn6-Tup1-repressible genes; namely, FLO11, HSP26, and SUC2. Sill is known to interact with Tpk2, a cAMP-dependent protein kinase that negatively regulates Sfl1 function. Consistently, we show that phosphorylation by protein kinase A inhibits Sfl1 DNA binding in vitro, and that a tpk2Δ mutation increases the levels of Sfl1 protein associated with specific promoter elements in vivo. These data indicate a possible mechanism for regulating Sfl1-mediated repression through modulation of DNA binding by cAMP-dependent protein kinase-dependent phosphorylation. Taken together with previous data, these new observations suggest a link between cAMP signaling and Ssn6-Tup1-mediated transcriptional repression.
AB - Ssn6 (Cyc8) is a component of the yeast general corepressor Ssn6-Tup1 that inhibits the transcription of many diversely regulated genes. The corepressor does not interact directly with DNA but is recruited to different promoters through interactions with distinct pathway-specific, DNA-binding repressor proteins. Using yeast two-hybrid and GST chromatography interaction experiments, we have determined that Sfl1, a novel repressor protein, interacts directly with Ssn6, and in vivo repression data suggest that Sfl1 inhibits transcription by recruiting Ssn6-Tup1 via a specific domain in the Sfl1 protein. Sin4 and Srb10, components of specific RNA polymerase II sub-complexes that are required for Ssn6-Tup1 repression activity, are found to be required for Sfl1 repression function. These results indicate a possible mechanism for Sfl1-mediated repression via Ssn6-Tup1 and specific subunits of the RNA polymerase II holoenzyme. Electrophoretic mobility shift and chromatin immunoprecipitation assays demonstrate that Sfl1 is present at the promoters of three Ssn6-Tup1-repressible genes; namely, FLO11, HSP26, and SUC2. Sill is known to interact with Tpk2, a cAMP-dependent protein kinase that negatively regulates Sfl1 function. Consistently, we show that phosphorylation by protein kinase A inhibits Sfl1 DNA binding in vitro, and that a tpk2Δ mutation increases the levels of Sfl1 protein associated with specific promoter elements in vivo. These data indicate a possible mechanism for regulating Sfl1-mediated repression through modulation of DNA binding by cAMP-dependent protein kinase-dependent phosphorylation. Taken together with previous data, these new observations suggest a link between cAMP signaling and Ssn6-Tup1-mediated transcriptional repression.
KW - cAMP signaling
KW - Corepressor
KW - Phosphorylation
KW - Transcription repression
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U2 - 10.1006/jmbi.2001.4742
DO - 10.1006/jmbi.2001.4742
M3 - Article
C2 - 11399075
AN - SCOPUS:0035933333
VL - 309
SP - 1007
EP - 1015
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
SN - 0022-2836
IS - 5
ER -