Nucleosome structure of the yeast CHA1 promoter: analysis of activation‐dependent chromatin remodeling of an RNA‐polymerase‐II‐transcribed gene in TBP and RNA pol II mutants defective in vivo in response to acidic activators

The Saccharomyces cerevisiae CHA1 gene encodes the catabolic L‐serine (L‐threonine) dehydratase. We have previously shown that the transcriptional activator protein Cha4p mediates serine/threonine induction of CHA1 expression. We used accessibility to micrococcal nuclease and DNase I to determine the in vivo chromatin structure of the CHA1 chromosomal locus, both in the non‐induced state and upon induction. Upon activation, a precisely positioned nucleosome (nuc‐1) occluding the TATA box and the transcription start site is removed. A strain devoid of Cha4p showed no chromatin alteration under inducing conditions. Five yeast TBP mutants defective in different steps in activated transcription abolished CHA1 expression, but failed to affect induction‐dependent chromatin rearrangement of the promoter region. Progressive truncations of the RNA polymerase II C‐terminal domain caused a progressive reduction in CHA1 transcription, but no difference in chromatin remodeling. Analysis of swi1, swi3, snf5 and snf6, as well as gcn5, ada2 and ada3 mutants, suggested that neither the SWI/SNF complex nor the ADA/GCN5 complex is involved in efficient activation and/or remodeling of the CHA1 promoter. Interestingly, in a sir4 deletion strain, repression of CHA1 is partly lost and activator‐independent remodeling of nuc‐1 is observed. We propose a model for CHA1 activation based on promoter remodeling through interactions of Cha4p with chromatin components other than basal factors and associated proteins.

[1]  Tom Maniatis,et al.  Transcriptional activation: A complex puzzle with few easy pieces , 1994, Cell.

[2]  R. Young,et al.  RNA polymerase II carboxy-terminal domain contributes to the response to multiple acidic activators in vitro. , 1991, Genes & development.

[3]  R. Kornberg,et al.  Nucleosomes inhibit the initiation of transcription but allow chain elongation with the displacement of histones , 1987, Cell.

[4]  L. Guarente,et al.  ADA5/SPT20 links the ADA and SPT genes, which are involved in yeast transcription , 1996, Molecular and cellular biology.

[5]  M. Grunstein,et al.  Nucleosome loss activates yeast downstream promoters in vivo , 1988, Cell.

[6]  K. Struhl,et al.  A new class of activation-defective TATA-binding protein mutants: evidence for two steps of transcriptional activation in vivo , 1996, Molecular and cellular biology.

[7]  K. Struhl.  Histone acetylation and transcriptional regulatory mechanisms. , 1998, Genes & development.

[8]  S. Pfaff,et al.  Chromatin Structure -element Positioning and Cis Dependent on Tfiiia Gene Transcription Is Xenopus , 1998 .

[9]  H. Xiao,et al.  Recruiting TATA-binding protein to a promoter: transcriptional activation without an upstream activator , 1995, Molecular and cellular biology.

[10]  J. Svaren,et al.  Transcription factors vs nucleosomes: regulation of the PHO5 promoter in yeast. , 1997, Trends in biochemical sciences.

[11]  L. Guarente,et al.  ADA3, a putative transcriptional adaptor, consists of two separable domains and interacts with ADA2 and GCN5 in a trimeric complex , 1995, Molecular and cellular biology.

[12]  J. Ahearn,et al.  A unique structure at the carboxyl terminus of the largest subunit of eukaryotic RNA polymerase II. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[13]  R. Kingston,et al.  Repression and activation by multiprotein complexes that alter chromatin structure. , 1996, Genes & development.

[14]  T Lagrange,et al.  The general transcription factors of RNA polymerase II. , 1996, Genes & development.

[15]  R. Kornberg,et al.  Upstream activation sequence-dependent alteration of chromatin structure and transcription activation of the yeast GAL1-GAL10 genes , 1989, Molecular and cellular biology.

[16]  J. Schmitz,et al.  Structural and functional requirements for the chromatin transition at the PHO5 promoter in Saccharomyces cerevisiae upon PHO5 activation. , 1993, Journal of molecular biology.

[17]  Michael R. Green,et al.  Binding of general transcription factor TFIIB to an acidic activating region , 1991, Nature.

[18]  A. Wolffe,et al.  A nucleosome‐dependent static loop potentiates estrogen‐regulated transcription from the Xenopus vitellogenin B1 promoter in vitro. , 1993, The EMBO journal.

[19]  Oscar M. Aparicio,et al.  Modifiers of position effect are shared between telomeric and silent mating-type loci in S. cerevisiae , 1991, Cell.

[20]  Jasper Rine,et al.  Silent information regulator protein complexes in Saccharomyces cerevisiae: a SIR2/SIR4 complex and evidence for a regulatory domain in SIR4 that inhibits its interaction with SIR3. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[21]  H. Gerber,et al.  Basal components of the transcription apparatus (RNA polymerase II, TATA ‐binding protein) contain activation domains: Is the repetitive c‐terminal domain (CTD) of RNA polymerase II a “Portable Enhance Domain”? , 1994, Molecular reproduction and development.

[22]  S. Buratowski,et al.  The basics of basal transcription by RNA polymerase II , 1994, Cell.

[23]  W. Hörz,et al.  A functional role for nucleosomes in the repression of a yeast promoter. , 1991, The EMBO journal.

[24]  S. Elgin,et al.  The role of a positioned nucleosome at the Drosophila melanogaster hsp26 promoter. , 1995, The EMBO journal.

[25]  J. Pérez-Ortín,et al.  Chromatin structure of the yeast FBP1 gene: Transcription‐dependent changes in the regulatory and coding regions , 1993, Yeast.

[26]  H. Xiao,et al.  A highly conserved domain of RNA polymerase II shares a functional element with acidic activation domains of upstream transcription factors , 1994, Molecular and cellular biology.

[27]  L. Guarente,et al.  ADA1, a novel component of the ADA/GCN5 complex, has broader effects than GCN5, ADA2, or ADA3 , 1997, Molecular and cellular biology.

[28]  K. Struhl,et al.  Mechanisms of transcriptional activation in vivo: two steps forward. , 1996, Trends in genetics : TIG.

[29]  M. Horikoshi,et al.  Recombinant yeast TFIID, a general transcription factor, mediates activation by the gene-specific factor USF in a chromatin assembly assay. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[30]  A. Dean,et al.  Yeast alpha 2 repressor positions nucleosomes in TRP1/ARS1 chromatin , 1990, Molecular and cellular biology.

[31]  S. Elgin,et al.  Protein/DNA architecture of the DNase I hypersensitive region of the Drosophila hsp26 promoter. , 1988, The EMBO journal.

[32]  C. Allis,et al.  Transcription-linked acetylation by Gcn5p of histones H3 and H4 at specific lysines , 1996, Nature.

[33]  R. Young,et al.  RNA polymerase II C-terminal repeat influences response to transcriptional enhancer signals , 1990, Nature.

[34]  A. Wolffe,et al.  Architectural specificity in chromatin structure at the TATA box in vivo: nucleosome displacement upon beta-phaseolin gene activation. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[35]  L. Franco,et al.  Fine analysis of the chromatin structure of the yeast SUC2 gene and of its changes upon derepression. Comparison between the chromosomal and plasmid-inserted genes. , 1987, Nucleic acids research.

[36]  P. Schjerling,et al.  Comparative amino acid sequence analysis of the C6 zinc cluster family of transcriptional regulators. , 1996, Nucleic acids research.

[37]  L. Verdone,et al.  Chromatin remodeling during Saccharomyces cerevisiae ADH2 gene activation , 1996, Molecular and cellular biology.

[38]  C. Ingles,et al.  Direct and selective binding of an acidic transcriptional activation domain to the TATA-box factor TFIID , 1990, Nature.

[39]  P. Schjerling,et al.  Cha4p of Saccharomyces cerevisiae activates transcription via serine/threonine response elements. , 1996, Genetics.

[40]  J. Wiame,et al.  Occurrence of a catabolic L-serine (L-threonine) deaminase in Saccharomyces cerevisiae. , 2005, European journal of biochemistry.

[41]  C. Brandl,et al.  Identification of Native Complexes Containing the Yeast Coactivator/Repressor Proteins NGG1/ADA3 and ADA2* , 1997, The Journal of Biological Chemistry.

[42]  S. Berger,et al.  Characterization of Physical Interactions of the Putative Transcriptional Adaptor, ADA2, with Acidic Activation Domains and TATA-binding Protein (*) , 1995, The Journal of Biological Chemistry.

[43]  M. Sheldon,et al.  Transcriptional Activation: Tuning-up transcription , 1995, Current Biology.

[44]  M. Espelund,et al.  A simple method for generating single-stranded DNA probes labeled to high activities. , 1990, Nucleic acids research.

[45]  D. Lohr,et al.  The relationship of regulatory proteins and DNase I hypersensitive sites in the yeast GAL1-10 genes. , 1985, Nucleic acids research.

[46]  K. Struhl,et al.  The TBP-TFIIA interaction in the response to acidic activators in vivo. , 1995, Science.

[47]  J. Rine,et al.  Replication and segregation of plasmids containing cis-acting regulatory sites of silent mating-type genes in Saccharomyces cerevisiae are controlled by the SIR genes , 1987, Molecular and cellular biology.

[48]  Pamela Reinagel,et al.  Contact with a component of the polymerase II holoenzyme suffices for gene activation , 1995, Cell.

[49]  D. Reinberg,et al.  Specific interaction between the nonphosphorylated form of RNA polymerase II and the TATA-binding protein , 1992, Cell.

[50]  C. Vincenz,et al.  The nucleoprotein hybridization method for isolating active and inactive genes as chromatin. , 1991, Methods in cell biology.

[51]  K. Struhl,et al.  Increased recruitment of TATA-binding protein to the promoter by transcriptional activation domains in vivo. , 1994, Science.

[52]  A. Berk,et al.  A class of activation domains interacts directly with TFIIA and stimulates TFIIA-TFIID-promoter complex assembly , 1995, Molecular and cellular biology.

[53]  M. Strubin,et al.  Stimulation of RNA polymerase II transcription initiation by recruitment of TBP in vivo , 1995, Nature.

[54]  P. Schjerling,et al.  A regulatory element in the CHA1 promoter which confers inducibility by serine and threonine on Saccharomyces cerevisiae genes. , 1993, Molecular and cellular biology.

[55]  J. O. Thomas.  Chromatin structure. , 1977, Biochemical Society symposium.

[56]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[57]  J. T. Kadonaga,et al.  Role of nucleosomal cores and histone H1 in regulation of transcription by RNA polymerase II. , 1991, Science.

[58]  F. Winston,et al.  Yeast SNF/SWI transcriptional activators and the SPT/SIN chromatin connection. , 1992, Trends in genetics : TIG.

[59]  S. Holmberg,et al.  Molecular genetics of serine and threonine catabolism in Saccharomyces cerevisiae. , 1988, Genetics.

[60]  A. Wolffe,et al.  The amino-terminal tails of the core histones and the translational position of the TATA box determine TBP/TFIIA association with nucleosomal DNA. , 1995, Nucleic acids research.

[61]  James T Kadonaga,et al.  SWI2/SNF2 and Related Proteins: ATP-Driven Motors That Disrupt-Protein–DNA Interactions? , 1997, Cell.

[62]  C. Peterson,et al.  Role for ADA/GCN5 products in antagonizing chromatin-mediated transcriptional repression , 1997, Molecular and cellular biology.

[63]  C. Allis,et al.  Tetrahymena Histone Acetyltransferase A: A Homolog to Yeast Gcn5p Linking Histone Acetylation to Gene Activation , 1996, Cell.

[64]  R. Young,et al.  Functional redundancy and structural polymorphism in the large subunit of RNA polymerase II , 1987, Cell.

[65]  U. Venter,et al.  [8] In Vivo Analysis of nucleosome structure and transcription factor binding in Saccharomyces cerevisiae , 1995 .

[66]  M. Ptashne,et al.  RNA Polymerase II Holoenzyme Recruitment Is Sufficient to Remodel Chromatin at the Yeast PHO5 Promoter , 1997, Cell.

[67]  M. Demma,et al.  Interaction with RAP74 subunit of TFIIF is required for transcriptional activation by serum response factor , 1995, Nature.

[68]  B. Cairns,et al.  Chromatin remodeling machines: similar motors, ulterior motives. , 1998, Trends in biochemical sciences.

[69]  J. Knezetic,et al.  The presence of nucleosomes on a DNA template prevents initiation by RNA polymerase II in vitro , 1986, Cell.

[70]  W Hörz,et al.  Nuclease hypersensitive regions with adjacent positioned nucleosomes mark the gene boundaries of the PHO5/PHO3 locus in yeast. , 1986, The EMBO journal.

[71]  J. Workman,et al.  Activation domains of stably bound GAL4 derivatives alleviate repression of promoters by nucleosomes , 1991, Cell.

[72]  S. Elgin,et al.  The chromatin structure of specific genes: II. Disruption of chromatin structure during gene activity , 1979, Cell.

[73]  J. Workman,et al.  Binding of transcription factor TFIID to the major late promoter during in vitro nucleosome assembly potentiates subsequent initiation by RNA polymerase II , 1987, Cell.

[74]  Michael Shales,et al.  Extensive homology among the largest subunits of eukaryotic and prokaryotic RNA polymerases , 1985, Cell.

[75]  M. Lee,et al.  Transcription‐induced nucleosome ‘splitting’: an underlying structure for DNase I sensitive chromatin. , 1991, The EMBO journal.

[76]  Michael R. Green,et al.  Facilitated binding of TATA-binding protein to nucleosomal DNA , 1994, Nature.