Yeast Enhancer of Polycomb defines global Esa1-dependent acetylation of chromatin

Boudreault, Alexandre A.; Cronier, Dominique; Selleck, William; Lacoste, Nicolas; Utley, Rhea T.; Allard, Stéphane; Savard, Julie; Lane, William S.; Tan, Song; Côté, Jacques

doi:10.1101/gad.1056603

Cited by 203 publications

(342 citation statements)

References 38 publications

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“…We have similarly identified the Piccolo NuA4 complex of Epl1, Yng2 and Esa1 as the catalytic core of the megadalton NuA4 complex [13,14]. Interestingly, Piccolo NuA4 possesses even greater HAT activity on nucleosomes than the whole NuA4 complex [14].…”

Section: Introductionmentioning

confidence: 83%

“…However, in contrast to SAGA which is apparently directed to the promoter through its Tra1 subunit, the Ada2/Ada3/Gcn5 complex (which may be identical to the HAT-A2 complex [10,11]) may act as a global, untargeted nucleosome acetyltransferase in the cell [12]. We have similarly identified the Piccolo NuA4 complex of Epl1, Yng2 and Esa1 as the catalytic core of the megadalton NuA4 complex [13,14]. Interestingly, Piccolo NuA4 possesses even greater HAT activity on nucleosomes than the whole NuA4 complex [14].…”

Section: Introductionmentioning

confidence: 93%

“…Furthermore, if the Esa1 subunit is HIS-tagged, it is possible that the Piccolo NuA4 prep will be contaminated with some Esa1 polypeptide that is not associated with the Epl1 and Yng2 subunits of Piccolo NuA4. This could complicate quantitative measurements of the nucleosomal HAT activity of Piccolo NuA4 relative to its naked histone HAT activity since both Esa1 and Piccolo acetylate histones but Piccolo possesses a much higher specific activity on nucleosomes than Esa1 does [13,14]. Therefore, it may be advisable to further purify the Piccolo NuA4 complex by additional chromatography steps.…”

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confidence: 99%

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Expression and purification of recombinant yeast Ada2/Ada3/Gcn5 and Piccolo NuA4 histone acetyltransferase complexes

Barrios

Selleck

Hnatkovich

et al. 2007

Methods

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Acetylation of histone tails by histone acetyltransferase (HAT) enzymes is a key post-translational modification of histones associated with transcriptionally active genes. Acetylation of the physiological nucleosome substrate is performed in cells by megadalton complexes such as SAGA and NuA4. To understand how HAT enzymes specifically recognize their nucleosome and not just histone tail substrates, we have identified the catalytic SAGA and NuA4 subcomplexes sufficient to act on nucleosomes. We describe here expression and purification procedures to prepare recombinant yeast Ada2/Ada3/Gcn5 subcomplex of SAGA which acetylates histones H3 and H2B on nucleosomes, and the Piccolo NuA4 complex which acetylates histones H4 and H2A on nucleosomes. We demonstrate an unexpected benefit of using the BL21-CodonPlus strain to enhance the purity of metal affinity purified Ada2/Ada3/Gcn5 complex. We also identify E. coli EF-Tu as a contaminant that copurifies with both complexes over multiple chromatographic steps and use of hydrophobic interaction chromatography to remove the contaminant from the Piccolo NuA4 complex. The methods described here will be useful for studies into the molecular mechanism of these enzymes and for preparing the enzymes as reagents to study the interplay of nucleosome acetylation with other chromatin modification and remodeling enzymes.

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Section: Introductionmentioning

confidence: 83%

Section: Introductionmentioning

confidence: 93%

mentioning

confidence: 99%

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Expression and purification of recombinant yeast Ada2/Ada3/Gcn5 and Piccolo NuA4 histone acetyltransferase complexes

Barrios

Selleck

Hnatkovich

et al. 2007

Methods

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“…Within NuA4, Epl1 serves as a scaffold protein bridging Yng2, Eaf6 and Esa1 to form the acetyltransferase core (Boudreault et al, 2003). Similarly, Epc1/2 bridges the interaction of TIP60 with ING3 and EAF6 to form the tetrameric acetyltransferase core , so an interesting possibility is whether BRPFs play a similar role in assembling and modulating MOZ and MORF complexes (Doyon et al, 2006).…”

Section: Moz and Morf As Catalytic Subunits Of Quartet Complexesmentioning

confidence: 99%

MOZ and MORF, two large MYSTic HATs in normal and cancer stem cells

2007

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Genes of the human monocytic leukemia zinc-finger protein MOZ (HUGO symbol, MYST3) and its paralog MORF (MYST4) are rearranged in chromosome translocations associated with acute myeloid leukemia and/or benign uterine leiomyomata. Both proteins have intrinsic histone acetyltransferase activity and are components of quartet complexes with noncatalytic subunits containing the bromodomain, plant homeodomain-linked (PHD) finger and proline-tryptophan-tryptophan-proline (PWWP)-containing domain, three types of structural modules characteristic of chromatin regulators. Although leukemia-derived fusion proteins such as MOZ-TIF2 promote self-renewal of leukemic stem cells, recent studies indicate that murine MOZ and MORF are important for proper development of hematopoietic and neurogenic progenitors, respectively, thereby highlighting the importance of epigenetic integrity in safeguarding stem cell identity.

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“…Esa1p is the catalytic component of the 1.4-MDa NuA4 HAT complex that is important for transcriptional activation both in vitro and in vivo at specific target loci, including many ribosomal protein genes (reviewed in Doyon and Côté, 2004). Esa1p also functions in the highly active, smaller picNuA4 complex (Boudreault et al, 2003).The NuA4 complex itself is broadly conserved (Doyon and Côté, 2004) and the human catalytic subunit Tip60 functions in nontranscriptional components of apoptosis and signaling processes (Ikura et al, 2000;Sheridan et al, 2001). Tip60 has been linked to DNA repair (Ikura et al, 2000) and a role for Esa1p and the Yng2p NuA4 subunit was found in replication-coupled DNA double-stranded break repair (Bird et al, 2002;Choy and Kron, 2002).…”

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confidence: 99%

Distinct Roles for the Essential MYST Family HAT Esa1p in Transcriptional Silencing

Clarke

Samal

Pillus

2006

MBoC

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Among acetyltransferases, the MYST family enzyme Esa1p is distinguished for its essential function and contribution to transcriptional activation and DNA double-stranded break repair. Here we report that Esa1p also plays a key role in silencing RNA polymerase II (Pol II)-transcribed genes at telomeres and within the ribosomal DNA (rDNA) of the nucleolus. These effects are mediated through Esa1p's HAT activity and correlate with changes within the nucleolus. Esa1p is enriched within the rDNA, as is the NAD-dependent protein deacetylase Sir2p, and the acetylation levels of key Esa1p histone targets are reduced in the rDNA in esa1 mutants. Although mutants of both ESA1 and SIR2 have enhanced rates of rDNA recombination, esa1 effects are more modest yet result in distinct structural changes of rDNA chromatin. Surprisingly, increased expression of ESA1 can bypass the requirement for Sir2p in rDNA silencing, suggesting that these two enzymes with seemingly opposing activities both contribute to achieve optimal nucleolar chromatin structure and function. INTRODUCTIONHistone modifications such as phosphorylation, methylation, acetylation, and deacetylation provide a mechanism by which chromatin structure is modulated to affect gene expression both positively and negatively (for reviews see Strahl and Allis, 2000;Iizuka and Smith, 2003;Kurdistani and Grunstein, 2003). The acetylation of lysine residues on histone N-terminal tails is classically linked to increases in gene expression and more directly to roles in transcriptional activation. By contrast, deacetylation of histones is most frequently correlated with transcriptionally silent chromatin. As more is learned about the enzymes catalyzing these modifications, the histone acetyltransferases (HATs) and histone deacetylases (HDACs), simple functional distinctions between acetylation and deacetylation of histones do not hold (Iizuka and Smith, 2003). For example, in several organisms, mutations in RPD3, a gene encoding a deacetylase, lead to increases in silencing rather than the expected decreases (DeRubertis et al., 1996;Rundlett et al., 1996). This brings forward the possibility that histone acetylation may play an important part in both silent and active chromatin. In addition, coactivator proteins, such as the histone acetyltransferases p300 and CBP and nuclear hormone receptors, appear to have roles in transcriptional repression through acetylation and association with particular binding partners (for examples, see Chen and Li, 1998;Waltzer and Bienz, 1998;Baluchamy et al., 2003;Girdwood et al., 2003;Rajabi et al., 2005). Thus, understanding of multiple roles for acetylation in controlling gene expression is expanding.In Saccharomyces cerevisiae, there are three regions of the genome controlled by chromatin-mediated transcriptional silencing: telomeres, the silent mating-type loci HMR and HML, and the rDNA array. Many factors are important for transcriptional silencing at subsets of these loci including the four core histones, the repressor activator protein Rap1...

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Yeast Enhancer of Polycomb defines global Esa1-dependent acetylation of chromatin

Cited by 203 publications

References 38 publications

Expression and purification of recombinant yeast Ada2/Ada3/Gcn5 and Piccolo NuA4 histone acetyltransferase complexes

Expression and purification of recombinant yeast Ada2/Ada3/Gcn5 and Piccolo NuA4 histone acetyltransferase complexes

MOZ and MORF, two large MYSTic HATs in normal and cancer stem cells

Distinct Roles for the Essential MYST Family HAT Esa1p in Transcriptional Silencing

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