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,
Cristina Popescu,2,
Laura Torroja,1,#
Daniel Ortuño-Sahagún,1,
Imre Boros,2,3 and
Alberto Ferrús1*
Department of Cellular, Molecular, and Developmental Neurobiology, Cajal Institute, C.S.I.C., Ave. Dr. Arce 37, Madrid 28002, Spain,1 Institute of Biochemistry, Biological Research Center, Temesvári krt. 62, H-6726 Szeged, Hungary,2 Department of Biochemistry HAS-USZ Chromatin Structure and Gene Expression Research Group, University of Szeged, Közép Fasor 52, H-6726 Szeged, Hungary3
Received 20 July 2007/ Returned for modification 25 September 2007/ Accepted 17 October 2007
| ABSTRACT |
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| INTRODUCTION |
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In yeast and higher eukaryotes, several GCN5-containing HAT complexes also share ADA2- and ADA3-type adaptors (1, 38, 45). Two Drosophila melanogaster HAT complexes, SAGA and ATAC, exhibit different specificities in the targeted histone lysines, and these complexes differ in the adaptor type, ADA2a or ADA2b, present (15, 30). A further ADA-type adaptor, ADA3, is present in both SAGA and ATAC complexes (29, 37); its role in vivo, however, so far has not been addressed. ADA3 was first isolated as a suppressor of GAL4-VP16-induced toxicity in yeast (53). Yeast ADA3 mutants grow slowly in minimal medium, reduce the efficiency of some transcriptional activation domains, and alter the selection of initiation sites in basal transcription (53). Based on these observations, ADA3 was proposed to act as a coactivator bridging acidic activation domains with the basic transcriptional machinery. A human ADA3-like protein was isolated from the PCAF complex, a HAT complex related to yeast SAGA (48), but little is known about its precise function in transcriptional regulation. In addition to being modified by acetylation, histones can be modified by phosphorylation. H3, in particular, exhibits acetylation of lysine 9 and phosphorylation of serine 10 (47). Several factors playing a role in H3 S10 phosphorylation have been identified, but the relationship between complexes depositing these two adjacent histone marks is not known (38).
In Drosophila melanogaster and other species, it is well documented that gene transcription can be affected by chromatin structure changes in the vicinity of the initiation site, a phenomenon referred to as position effect variegation (PEV). Defective histone deacetylation in RPD3 mutants enhances PEV (17), and a lack of phosphorylation by JIL-1 kinase on H3 affects PEV (34). SAGA and TFIID share TAFII components, illustrating the coupling between the alteration of chromatin structure and transcription initiation (28, 46). Selective gene expression, however, requires that, in addition to functional coupling, chromatin remodeling and transcription initiation complexes act on selective genome sites and tissues and during developmental times. Hence, there is a need to identify the elements involved in the site specificity of HAT complex activity and to demonstrate their role in vivo.
| MATERIALS AND METHODS |
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6 and
9 were kindly provided by Pilar Carrera (IGBMC, Strasbourg, France). These were generated by imprecise excision of P{Mae-UAS.6.11}CG7536GG01344, which is located 5' in dik (dAda3) coding sequence. Both deletions remove the 5' end of dik (dAda3) and parts of the second exon of the gene CG7536 within which dik (dAda3) is nested. The IRO-C rearrangements and the AS-C enhancer constructs are from J. Modolell (Center for Molecular Biology, Madrid, Spain). AS-C enhancers correspond to the following genomic fragments from the achaete-scute region. sc1.1 extends 1.1 kb and includes the scute start site (42); (1.2/0.0) 3.7 sc extends 3.7 kb from the same site near the scute promoter (11), and we refer to it as 3.7 sc for brevity; and, finally, AS 1.4DC18 corresponds to 1.4 kb from the scute start site, but it contains a heat shock basal promoter (21). All these constructs carried the LacZ gene as the expression reporter. Alleles dAda2a189 and dAda2b842 have been described previously (49). The upstream activation sequence-dAda3+ construct was cloned in the pUAST vector and injected into y w embryos to obtain transgenic lines. Primer sequences used for cDNA cloning are available upon request. For genomic rescue experiments, the EcoRI fragment 422E4 (3) was inserted into the germ line transformation vector pCasper (54). The proximal XbaI fragment of 422E4, which contains an additional transcription unit, HL-IV, was deleted by partial restriction digestion. The resulting construct, pCasper422E4RX, was used for P element-mediated germ line transformation. We refer to this construct as E4RX for brevity. Other deficiencies, mutants, and chromosomal rearrangements were from the Bloomington Stock Center (Fly Base [http://flybase.bio.indiana.edu]). Mosaic and phenotype analyses. Twin clones were scored on the cuticle of adult y w dik*os/f5 os females X-ray irradiated at 0 to 48 h after egg laying. A twin clone is composed of two adjacent patches generated in the same event of recombination, one marked with forked and the other with yellow, wherein the yellow clone is homozygous for the lethal mutation. The generation of germ line clones was elicited by X-ray irradiation of first-instar ovoKS1237 v/y w dAda3* larvae as described previously (20). To determine the lethality phase, dAda3*/FM6 females were crossed with wild-type Canton S males. Fertilized females were allowed to lay eggs for a 20-h period at 25°C. Groups of 20 to 30 eggs were examined at 1-day intervals; the surviving individuals were counted. The imaginal disk size was evaluated on dissected material from late-third-instar larvae. The extent of variegation was evaluated by comparing sibling individuals from crosses of males with genotype y w dAda3*; E4RX/TM3 against females with genotype In(1)wm4/FM6 and females with genotype y w dAda3*/FM6; Ki Sb Dp(1;3)JC153/TM2 against males with genotype In(1)wm4. An additional control reference was obtained from the cross, In(1)wm4/FM6 females against y w; E4RX/TM3 males. Variegation for the hairy wing (Hwv) phenotype was studied in the offspring from the cross: In(1)sc8/In(1)sc8 females against y w dAda3*; E4RX/TM3 males. The number of pleural bristles was counted in In(1)sc8/y w dAda3*; TM3/+ females and the sibling controls with the genotype In(1)sc8/y w dAda3*; E4RX/+. These values were subjected to Student's t test for statistical significance. All variegation studies were done on cultures raised at 25°C.
Gene expression studies.
For Northern blotting, poly(A)+ mRNA was isolated using a QuickPrep Micro mRNA purification kit (Pharmacia Biotech), and the probe was [
-32P]dATP radiolabeled by the random priming method (19). In situ expression was analyzed by hybridization of a cDNA probe. The corresponding cRNA was labeled with digoxigenin and hybridized to 12-µm paraffin-embedded sections by following standard procedures. In vivo experiments on directed gene expression were done using the upstream activation sequence-Gal4 system (50, 51). For a source of the Iroquois gene complex component araucan, we used the UASara29 construct (18) under Gal4-AC216 and Gal4-MD751 enhancer-trap drivers, which correspond to the spalt and u-shaped (ush) genes, respectively (M. Calleja, personal communication).
Anti-dADA3 antibodies, bacterial expression, and Western blots. The dADA3-specific polyclonal serum was generated against the peptide DSLDKDEKRQDRRK, which corresponds to amino acids 126 to 139 in the dADA3 sequence. Peptide synthesis and immunization of rabbits were performed by Sigma Co. The antibodies were purified by affinity chromatography on peptide-bound Sepharose 4 Fast Flow matrix (Amersham Bioscience) according to the manufacturer's instructions. For protein expression in bacteria, dAda3 cDNA was generated by reverse transcription-PCR using nucleotide primers based on expressed sequence tag sequences. The fragment was inserted into pGEX-4T-3 expression vector (GE Healthcare Life Sciences) for transformation. Protein expression was induced by IPTG (isopropyl-β-D-1-thiogalactopyranoside; 1 mM) and analyzed on polyacrylamide gel electrophoresis. Nuclear extracts of third-instar larvae were prepared as described elsewhere (25). For Western blotting, protein samples were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, electroblotted to nitrocellulose membrane, and incubated with affinity-purified primary antibodies. Peroxidase-conjugated goat anti-rabbit immunoglobulin (Ig) and rabbit anti-mouse (Sigma-Aldrich) specific antibody were used as secondary antibodies, and the blots were developed using Super Signal West Pico chemiluminescent substrate (Pierce).
Immunostaining of polytene chromosomes. Salivary gland polytene chromosomes were obtained from wandering third-instar larvae of dAda3, dAda2a, dAda2b, and dGcn5 mutants (52). Immunopurified anti-ADA3 polyclonal rabbit serum was used in 100-fold dilution. H3 acetyl (Ac)K14 (1:100)-, H3 AcK9 (1:100)-, H3 AcK18 (1:300)-, and H3 P-S10 (1:50)-specific antibodies (Upstate), H4 AcK5 (1:100) and H4 AcK12 (1:150) (ABCAM) antibodies, and H4 AcK16 (1:200) (Serotec) antibody were used in dilutions as indicated. Mouse anti-RNA polymerase II (Pol II; 7G5) antibody was a gift of L. Tora (IGBMC, Strasbourg, France). Secondary antibodies, AlexaFluor488-conjugated goat anti-mouse Ig, AlexaFluor555-conjugated goat anti-rabbit Ig, and AlexaFluor568-conjugated goat anti-rabbit Ig (Molecular Probes) were used in 500-fold dilutions. Images for the semiquantitative analysis of fluorescence signal were acquired under constant conditions by using a Leica TCS SP5 confocal microscope. Image analysis was performed using ImageJ (http://rsb.info.nih.gov/ij/). Mean fluorescence intensities of the different signals of the different chromosomes were calculated after thresholding the channels. Normalization of the data was avoided by applying ratios of the corresponding signal couples for the following statistics. Data were compared using Mann-Whitney U test.
Nucleotide sequence accession number. We reported the sequence of the dADA3-encoding gene to the NCBI database (GeneID accession number 32787).
| RESULTS |
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dADA3 is involved in site-specific acetylation/phosphorylation of histones. Since dADA3 is a component of several HAT complexes (30, 37), we first searched for genetic interactions between dAda3 alleles and mutants affecting subunits of HAT complexes. dAda2a, dAda2b, and dGcn5 mutations impair the function of ATAC and SAGA complexes (38). Similar to dAda3 mutants, mutants of these HAT complex components show lethality phases around metamorphosis (see Fig. S6 in the supplemental material). dAda3 and dGcn5 null mutations are lethal at early pupal and late larval stages, respectively. In contrast, dAda3-dGcn5 double mutants die at L1 and L2 stages, earlier than either single mutant (see Fig. S6 in the supplemental material). For dAda2a-dAda3 and dAda2b-dAda3 double mutants, there was no evidence of genetic interaction because there was no shift of the lethal phase (not shown). Also, we noticed that the distorted polytene chromosome structure characteristic of dAda2a and dGcn5 mutants, but not of dAda2b mutants (49), is very similar to that observed here in dAda3 mutants (Fig. 3D). Furthermore, in dGcn5 and dAda2a mutants, the dADA3 immunosignal on chromosomes is greatly reduced (Fig. 4A, B, and D). In contrast, the signal in dAda2b mutants appears with intensity not different from that of the wild type (Fig. 4C). It should be noted that the dAda2b allele used is a transcriptional null (49), which rules out the possibility that the normal dADA3 signal on chromosomes could be due to residual dADA2b protein content. These qualitative data were confirmed with Western blots from larval nuclear extracts (Fig. 4E and F).
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dADA3 regulates the expression of specific genes. While testing chromosomal deficiencies for possible phenotype interactions with dAda3 alleles, we detected a significant interaction with the Df(3L)iro2 (see Table S1 in the supplemental material). This rearrangement deletes the Iroquoi gene complex (IRO-C), composed by the homeoprotein-encoding genes araucan, caupolican, and mirror (27). Double heterozygotes for dAda3 and IRO-C [the genotype for females was dAda3/+; Df(3L)iro2/+ and that for males was dAda3; Df(3L)iro2/E4RX] exhibit, with differing penetrances and expressivities, IRO-C phenotypes (Fig. 7D and E). In a given cross, two-thirds of the adult offspring (n = 54) showed a recognizable IRO phenotype, and about half of these showed the phenotype illustrated in Fig. 7E. In a stock established for more than two years [genotype y w dAda32/FM6; Df(3L)iro2/TM3] the penetrance of IRO-C phenotypes seems stabilized at around 80% in heterozygous females. Penetrance and expressivity of this effect is temperature sensitive, since cultures raised at 27°C exhibit stronger phenotypes than those raised at 17°C. The effect was reproduced, although with somewhat lower expressivity, when the mutants with partial deletions of the IRO-C, Df(3L)iroDFM1 (with araucan and caupolican deleted) and Df(3L)iroDFM3 (with araucan, caupolican, and the promoter of mirror deleted) mutants, were assayed (n, 47 and 62, respectively). As reference controls, the described phenotypes were never observed among heterozygote deficiencies for either dAda3 or IRO-C. Since Df(3L)iro2 includes also the Gcn5 locus (59), it may be argued that this component of the SAGA and ATAC complexes may contribute, in addition to dADA3, to the observed enhancing of IRO-C phenotypes.
The IRO-C-encoded functions seem to be partially redundant because deletions, but not single gene mutations, yield mutant phenotypes (27). We took advantage of this redundancy to test if the observed phenomenon in dAda3/+; Df(3L)iro2/+ adults was due to a depletion of IRO-C expression or to some other function included in this deficiency (e.g., GCN5). To that end, we generated adults of genotype dAda3/+; Gal4-AC216/+; Df(3L)iro2/UAS-ara29 in which the expression of the araucan component of IRO-C is driven to the wing disk by Gal4-AC216, an insertion under the control of enhancers from the spalt gene. Here, IRO-C phenotypes were never observed (n = 235), demonstrating that the phenomenon results from the combined reduced expression of dAda3 and IRO-C. In the corresponding crosses, the sibling genotype dAda3/+; Df(3L)iro2/UAS-ara29 exhibit the regular IRO-C phenotypes (Fig. 7E). To further document the functional relationship between dAda3 and IRO-C genes, we assayed one of the IRO-C targets, the regulatory regions of the achaete-scute gene complex, AS-C (27). We tested the effects of dAda32 on the expression of constructs in which the LacZ reporter is under the control of several AS-C enhancers. Mutant wing disks (n = 36) were tested for the expression of enhancers sc 1.1, AS 1.4DC, and 3.7 sc. In all cases, the expression was abolished in the mutant disks, while it was maintained in their sibling controls (Fig. 7F,G). Only a small number of mutant disks (n = 6) showed a weak LacZ expression in some, but not all, of the normal wing disk expression domains. This observation is consistent with a role of dADA3 in regulating the expression of AS-C, either directly or in conjunction with members of IRO-C. As an additional demonstration of these gene transcriptional effects of dADA3, we checked the transcriptome of dAda3 mutant larvae. The microarray data indicate the expected down regulation of scute, araucan, and caupolican genes (not shown).
Following the rationale inspired by the results with IRO-C, we analyzed other genes involved in bristle patterning in the dorsal notum. The mutant amosTft produces an excess of macrobristles in the same area of the thorax where most dominant IRO-C phenotypes are found in dAda3 heterozygotes, the scutellum. We found that the amosTft phenotype, like that of IRO-C, is enhanced in a dAda3/+ background (the mean ± standard deviation of thorax macrobristles was 22.5 ± 2.1 in the mutants, compared to 18.1 ± 3.1 in controls [16 and 8 adults, respectively]). A similar enhancement effect is detected with AntennapediaHu with respect to the number of extrabristles in the humerus (not shown). It should be noted that the humerus and the scutellum are equivalent metameric structures. The observed effects seem specific, since equivalent tests with other deletions [one of the deletions, Df(2L)TW201.6, included a cluster of histone-encoding genes], genetic constructs, or mutants failed to show evidences of functional interaction (see Table S1 in the supplemental material).
| DISCUSSION |
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(4) and also functions as a coactivator for p53-dependent (36) and retinoic acid X receptor alpha-mediated transactivation of target genes (66). More recently, the presence of an ADA3-like protein in several Drosophila HAT complexes has been demonstrated by biochemical dissociation of SAGA and ATAC subunits (37). In all these multiprotein complexes, ADA3 is thought to bind ADA2 and a transcriptional activator through its carboxy and amino termini, respectively (13, 32). In turn, ADA2 associates with GCN5, the acetylase component, sustaining the activity of the whole complex. Under the assumption that these interactions occur also in Drosophila, the mutant versions analyzed here would be unable to interact with dADA2 because of their truncation towards the C terminus. The selective loss of histone H3 and H4 acetylation at specific lysine residues in dAda3 mutants provides an in vivo demonstration that dADA3 plays an essential role in HAT complexes. In Drosophila, dADA3 is present in the SAGA and ATAC complexes, both of which have a preference for H3 and H4 acetylation. Thus, the loss of selected acetylation activities in both histones indicates the functional deficiency of both complexes in dAda3 mutants. We have previously reported the involvement of fly ADA2b-containing SAGA complex in histone H3 K9 and K14 acetylation and the involvement of the dADA2a-containing ATAC complex in histone H4 K5 and K12 acetylation (15). Neither complex, however, seems involved in the acetylation of H3 K4 or K18 or H4 K8 or K16. Here, in dAda3 mutants, the acetylations of H3 K9 and K14 and H4 K12 are reduced, offering repertoires that are similar, but not identical, to those of dAda2b and dAda2a mutants. Surprisingly, we did not observe acetylation defects in H4 K5 in dAda3 mutants, while in dAda2a and dGcn5 mutants, this site is affected. This discrepancy probably reflects the relative contribution that each ADA component has to the selectivity of the HAT complex to which it belongs. Perhaps, in the absence of dADA3, a site specificity shift of the malfunctioning ATAC from H4 K5 to K12, as described for yeast, could be considered (2). The alternative possibility of additional, yet unidentified, dADA3-containing HAT complexes cannot be ruled out.
The phosphorylation effect on H3 S10 is a novel feature for an ADA3 component. It should be emphasized, however, that acetylation and phosphorylation of H3 appear as two closely related processes, since an adaptor and an acetylating enzyme affect both of them, suggesting that the two forms of histone modification are a property of HAT complexes. The TRA1 component of yeast SAGA exhibits kinase activity, but the H3 S10 phosphorylation is carried out by the SNF1 kinase (40). In Drosophila, however, the precise identity of the H3-phosphorylating complex remains to be determined. The H3-targeted JIL-1 kinase activity is known to associate with the MSL complex involved in the transcriptional modulation of male X chromosome genes (34). In addition, Jil-1 mutants show chromosome structure defects akin to those of dAda3, dGcn5, and dAda2a mutants (34, 49). However, it is still unknown if JIL-1 is a component of a HAT complex. Consistent with its role in histone modification, dADA3 is found on transcriptionally active regions of polytene chromosomes. The quantitative analysis of in situ data showed colocalization of dADA3 with RNA Pol II, albeit with variable stoichiometry according to chromosomal site. This feature is compatible with the proposal that dADA3-containing complexes could have different, probably sequential, effects on nucleosomes, for instance, phosphorylation (e.g., H3 S10 phosphorylation) followed by acetylation and finalizing with initiation of gene transcription by RNA Pol II. In the first event, dADA3 should not be expected to colocalize with RNA Pol II, while it should during initiation of gene transcription. Perhaps it is worth not considering this sequence of events the result of a single type of HAT complex activity or the ordered parade of different complexes on the same chromosomal site. It may be more appropriate to envision the whole sequence as a gradual transition of chemical interactions elicited by a complex in which its components are substituted at different rates. Under this speculative view, assigning a component to a complex becomes less relevant unless the functional context of the chromosomal site is specified. Thus, as an example, a HAT complex would become gradually transformed into a TFIID complex. A case reflecting this conceptual view is already documented for the yeast INO1 promoter (40).
The modification of histones is expected to result in structural changes of the chromatin. In this context, it is important to note that dAda3 mutant chromosomes appear abnormally structured in their polyteny. This is direct evidence that dADA3-containing complexes play a role in the maintenance of chromatin integrity, a process in which dADA2a and dGCN5 also participate. dADA2b, however, seems not to be involved in this structural role. In turn, chromatin structure is expected to result in gene expression effects. Gene variegation is a form of gene silencing elicited by changes in chromatin structure that can yield several types of position effects. Relocation of centromeric heterochromatin to the vicinity of transcription units is among the most widely studied effects (16); modification of pairing between homologous chromosome regions is another widely studied effect (26). Different types of heterochromatin, most notably that from the X chromosome in comparison to that from the Y or autosome chromosomes, exhibit differential variegation properties (39). The data reported here show that dADA3 is a modifier of variegation in a gene-dosage-dependent manner. Loss of function enhances the phenomenon, while additional gene copies reduce it. The effect, however, is dependent on the variegating rearrangement. The three X heterochromatin rearrangements tested showed the dAda3 background effect, while Y heterochromatin (bwD), euchromatic [Tp(2;2)Sco], and homologue-pairing (white and zeste) rearrangements have not. It appears, thus, that the mutant effect is selective of loci and/or heterochromatin type. This feature is compatible with targeting of the dADA3-containing complexes to selective chromatin regions. The site specificity of chromatin-binding complexes and the intervening mechanisms are still poorly defined (5, 9, 63), although some genetic functions have been described as being specific for certain types of variegation (57). Transcription factors such as GAGA and proliferation disrupter (Prod) are known to bind heterochromatin at specific times during the cell cycle (55). GAGA, in particular, binds AC-rich sequences in the heterochromatin associated with bwD. Consistent with the lack of dAda3 mutant background effect on bwD variegation, we tested the GAGA-encoding gene trithorax-like (Trl) for potential effects in double heterozygotes with dAda3 mutants, yielding negative results (data not shown). The loss of dADA3 causes a reduced transcriptional activity of specific genes, as demonstrated for the IRO-C and their targets on scute enhancers. This is direct evidence that dADA3 plays a role in gene expression and that this role is locus specific. In this context, unraveling the specificity of adaptors and other HAT components opens new avenues to target gene expression in vivo in a tissue- and time-controlled manner.
| ACKNOWLEDGMENTS |
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Research was funded by grants from the Spanish Ministry of Science (BFU2006-10180), the Hungarian Science Fund (OTKA T046414), and the European Network TAF-CHROMATIN (MRTN-CT-2004-504288).
| FOOTNOTES |
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Published ahead of print on 29 October 2007. ![]()
Supplemental material for this article may be found at http://mcb.asm.org/. ![]()
These authors have contributed equally to this report. ![]()
# Present address: Departamento de Biología, Facultad de Ciencias Biológicas, Universidad Autónoma de Madrid, Madrid, Spain. ![]()
Present address: Department of Cellular and Molecular Biology, Instituto de Neurobiología, Universidad de Guadalajara, Guadalajara, México. ![]()
| REFERENCES |
|---|
|
|
|---|
2. Balasubramanian, R., M. G. Pray-Grant, W. Selleck, P. A. Grant, and S. Tan. 2002. Role of the Ada2 and Ada3 transcriptional coactivators in histone acetylation. J. Biol. Chem. 277:7989-7995.
3. Baumann, A., I. Krah-Jentgens, R. Muller, F. Muller-Holtkamp, R. Seidel, N. Kecskemethy, J. Casal, A. Ferrus, and O. Pongs. 1987. Molecular organization of the maternal effect region of the Shaker complex of Drosophila: characterization of an I(A) channel transcript with homology to vertebrate Na channel. EMBO J. 6:3419-3429.[Medline]
4. Benecke, A., C. Gaudon, J. M. Garnier, E. vom Baur, P. Chambon, and R. Losson. 2002. ADA3-containing complexes associate with estrogen receptor alpha. Nucleic Acids Res. 30:2508-2514.
5. Berger, S. L. 1999. Gene activation by histone and factor acetyltransferases. Curr. Opin. Cell Biol. 11:336-341.[CrossRef][Medline]
6. Berger, S. L. 2007. The complex language of chromatin regulation during transcription. Nature 447:407-412.[CrossRef][Medline]
7. Bickel, S., and V. Pirrotta. 1990. Self-association of the Drosophila zeste protein is responsible for transvection effects. EMBO J. 9:2959-2967.[Medline]
8. Bingham, P. M., and Z. Zachar. 1985. Evidence that two mutations, wDZL and z1, affecting synapsis-dependent genetic behavior of white are transcriptional regulatory mutations. Cell 40:819-825.[CrossRef][Medline]
9. Bonifer, C. 2000. Developmental regulation of eukaryotic gene loci: which cis-regulatory information is required? Trends Genet. 16:310-315.[CrossRef][Medline]
10. Brown, C. E., L. Howe, K. Sousa, S. C. Alley, M. J. Carrozza, S. Tan, and J. L. Workman. 2001. Recruitment of HAT complexes by direct activator interactions with the ATM-related Tra1 subunit. Science 292:2333-2337.
11. Campuzano, S., L. Carramolino, C. V. Cabrera, M. Ruiz-Gomez, R. Villares, A. Boronat, and J. Modolell. 1985. Molecular genetics of the achaete-scute gene complex of D. melanogaster. Cell 40:327-338.[CrossRef][Medline]
12. Candau, R., and S. L. Berger. 1996. Structural and functional analysis of yeast putative adaptors. Evidence for an adaptor complex in vivo. J. Biol. Chem. 271:5237-5245.
13. Candau, R., J. X. Zhou, C. D. Allis, and S. L. Berger. 1997. Histone acetyltransferase activity and interaction with ADA2 are critical for GCN5 function in vivo. EMBO J. 16:555-565.[CrossRef][Medline]
14. Cheung, P., K. G. Tanner, W. L. Cheung, P. Sassone-Corsi, J. M. Denu, and C. D. Allis. 2000. Synergistic coupling of histone H3 phosphorylation and acetylation in response to epidermal growth factor stimulation. Mol. Cell 5:905-915.[CrossRef][Medline]
15. Ciurciu, A., O. Komonyi, T. Pankotai, and I. M. Boros. 2006. The Drosophila histone acetyltransferase Gcn5 and transcriptional adaptor Ada2a are involved in nucleosomal histone H4 acetylation. Mol. Cell. Biol. 26:9413-9423.
16. Csink, A. K., and S. Henikoff. 1996. Genetic modification of heterochromatic association and nuclear organization in Drosophila. Nature 381:529-531.[CrossRef][Medline]
17. De Rubertis, F., D. Kadosh, S. Henchoz, D. Pauli, G. Reuter, K. Struhl, and P. Spierer. 1996. The histone deacetylase RPD3 counteracts genomic silencing in Drosophila and yeast. Nature 384:589-591.[CrossRef][Medline]
18. Diez del Corral, R., P. Aroca, J. L. Gomez-Skarmeta, F. Cavodeassi, and J. Modolell. 1999. The Iroquois homeodomain proteins are required to specify body wall identity in Drosophila. Genes Dev. 13:1754-1761.
19. Feinberg, A. P., and B. Vogelstein. 1984. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal. Biochem. 137:266-267. (Addendum.)[CrossRef][Medline]
20. Ferrus, A., S. Llamazares, J. L. de la Pompa, M. A. Tanouye, and O. Pongs. 1990. Genetic analysis of the Shaker gene complex of Drosophila melanogaster. Genetics 125:383-398.[Abstract]
21. Garcia-Garcia, M. J., P. Ramain, P. Simpson, and J. Modolell. 1999. Different contributions of pannier and wingless to the patterning of the dorsal mesothorax of Drosophila. Development 126:3523-3532.[Abstract]
22. Gerasimova, T., D. Gdula, D. Gerasimov, O. Simonova, and V. Corces. 1995. A Drosophila protein that imparts directionality on a chromatin insulator is an enhancer of position-effect variegation. Cell 82:587-597.[CrossRef][Medline]
23. Geyer, P. K., and V. G. Corces. 1987. Separate regulatory elements are responsible for the complex pattern of tissue-specific and developmental transcription of the yellow locus in Drosophila melanogaster. Genes Dev. 1:996-1004.
24. Geyer, P. K., M. M. Green, and V. G. Corces. 1990. Tissue-specific transcriptional enhancers may act in trans on the gene located in the homologous chromosome: the molecular basis of transvection in Drosophila. EMBO J. 9:2247-2256.[Medline]
25. Gim, B., J. Park, J. Yoon, C. Kang, and Y. Kim. 2001. Drosophila Med6 is required for elevated expression of a large but distinct set of developmentally regulated genes. Mol. Cell. Biol. 21:5242-5255.
26. Goldsborough, A. S., and T. B. Kornberg. 1996. Reduction of transcription by homologue asynapsis in Drosophila imaginal discs. Nature 381:807-810.[CrossRef][Medline]
27. Gomez-Skarmeta, J. L., R. Diez del Corral, E. de la Calle-Mustienes, D. Ferre-Marco, and J. Modolell. 1996. Araucan and caupolican, two members of the novel iroquois complex, encode homeoproteins that control proneural and vein-forming genes. Cell 85:95-105.[CrossRef][Medline]
28. Grant, P. A., D. Schieltz, M. G. Pray-Grant, D. J. Steger, J. C. Reese, J. R. Yates III, and J. L. Workman. 1998. A subset of TAF(II)s are integral components of the SAGA complex required for nucleosome acetylation and transcriptional stimulation. Cell 94:45-53.[CrossRef][Medline]
29. Guelman, S., T. Suganuma, L. Florens, S. K. Swanson, C. L. Kiesecker, T. Kusch, S. Anderson, J. R. Yates III, M. P. Washburn, S. M. Abmayr, and J. L. Workman. 2006. Host cell factor and an uncharacterized SANT domain protein are stable components of ATAC, a novel dAda2A/dGcn5-containing histone acetyltransferase complex in Drosophila. Mol. Cell. Biol. 26:871-882.
30. Guelman, S., T. Suganuma, L. Florens, V. Weake, S. K. Swanson, M. P. Washburn, S. M. Abmayr, and J. L. Workman. 2006. The essential gene wda encodes a WD40 repeat subunit of Drosophila SAGA required for histone H3 acetylation. Mol. Cell. Biol. 26:7178-7189.
31. Henikoff, S. 1997. Nuclear organization and gene expression: homologous pairing and long-range interactions. Curr. Opin. Cell Biol. 9:388-395.[CrossRef][Medline]
32. Horiuchi, J., N. Silverman, G. A. Marcus, and L. Guarente. 1995. ADA3, a putative transcriptional adaptor, consists of two separable domains and interacts with ADA2 and GCN5 in a trimeric complex. Mol. Cell. Biol. 15:1203-1209.[Abstract]
33. Ito, T. 2007. Role of histone modification in chromatin dynamics. J. Biochem. (Tokyo) 141:609-614.
34. Jin, Y., Y. Wang, J. Johansen, and K. Johansen. 2000. JIL-1, a chromosomal kinase implicated in regulation of chromatin structure, associates with the male specific lethal (MSL) dosage compensation complex. J. Cell Biol. 149:1005-1010.
35. Kulesza, C. A., H. A. Van Buskirk, M. D. Cole, J. C. Reese, M. M. Smith, and D. A. Engel. 2002. Adenovirus E1A requires the yeast SAGA histone acetyltransferase complex and associates with SAGA components Gcn5 and Tra1. Oncogene 21:1411-1422.[CrossRef][Medline]
36. Kumar, A., Y. Zhao, G. Meng, M. Zeng, S. Srinivasan, L. M. Delmolino, Q. Gao, G. Dimri, G. F. Weber, D. E. Wazer, H. Band, and V. Band. 2002. Human papillomavirus oncoprotein E6 inactivates the transcriptional coactivator human ADA3. Mol. Cell. Biol. 22:5801-5812.
37. Kusch, T., S. Guelman, S. M. Abmayr, and J. L. Workman. 2003. Two Drosophila Ada2 homologues function in different multiprotein complexes. Mol. Cell. Biol. 23:3305-3319.
38. Lee, K. K., and J. Workman. 2007. Histone acetyltransferase complexes: one size doesn't fit all. Nat. Rev. Mol. Cell Biol. 8:284-295.[CrossRef][Medline]
39. Lloyd, V. K., D. A. Sinclair, and T. A. Grigliatti. 1997. Competition between different variegating rearrangements for limited heterochromatic factors in Drosophila melanogaster. Genetics 145:945-959.[Abstract]
40. Lo, W., L. Duggan, N. Emre, R. Belotserkovskya, W. Lane, R. Shiekhattar, and S. Berger. 2001. Snf1—a histone kinase that works in concert with the histone acetyltransferase Gcn5 to regulate transcription. Science 293:1142-1146.
41. Lo, W. S., R. C. Trievel, J. R. Rojas, L. Duggan, J. Y. Hsu, C. D. Allis, R. Marmorstein, and S. L. Berger. 2000. Phosphorylation of serine 10 in histone H3 is functionally linked in vitro and in vivo to Gcn5-mediated acetylation at lysine 14. Mol. Cell 5:917-926.[CrossRef][Medline]
42. Martinez, C., and J. Modolell. 1991. Cross-regulatory interactions between the proneural achaete and scute genes of Drosophila. Science 251:1485-1487.
43. McGill, S., W. Chia, R. Karp, and M. Ashburner. 1988. The molecular analyses of an antimorphic mutation of Drosophila melanogaster, Scutoid. Genetics 119:647-661.
44. Morales, V., C. Giamarchi, C. Chailleux, F. Moro, V. Marsaud, S. Le Ricousse, and H. Richard-Foy. 2001. Chromatin structure and dynamics: functional implications. Biochimie 83:1029-1039.[Medline]
45. Nagy, Z., and L. Tora. 2007. Distinct GCN5/PCAF-containing complexes function as co-activators and are involved in transcription factor and global histone acetylation. Oncogene 26:5341-5357.[CrossRef][Medline]
46. Narlikar, G. J., H. Y. Fan, and R. E. Kingston. 2002. Cooperation between complexes that regulate chromatin structure and transcription. Cell 108:475-487.[CrossRef][Medline]
47. Nowak, S., and V. Corces. 2004. Phosphorylation of histone H3: a balancing act between chromosome condensation and transcriptional activation. Trends Genet. 20:214-220.[CrossRef][Medline]
48. Ogryzko, V. V., T. Kotani, X. Zhang, R. L. Schiltz, T. Howard, X. J. Yang, B. H. Howard, J. Qin, and Y. Nakatani. 1998. Histone-like TAFs within the PCAF histone acetylase complex. Cell 94:35-44.[CrossRef][Medline]
49. Pankotai, T., O. Komonyi, L. Bodai, Z. Ujfaludi, S. Muratoglu, A. Ciurciu, L. Tora, J. Szabad, and I. Boros. 2005. The homologous Drosophila transcriptional adaptors ADA2a and ADA2b are both required for normal development but have different functions. Mol. Cell. Biol. 25:8215-8227.
50. Perrimon, N. 1998. New advances in Drosophila provide opportunities to study gene functions. Proc. Natl. Acad. Sci. USA 95:9716-9717.
51. Phelps, C. B., and A. H. Brand. 1998. Ectopic gene expression in Drosophila using GAL4 system. Methods 14:367-379.[CrossRef][Medline]
52. Pile, L. A., and D. A. Wassarman. 2002. Localizing transcription factors on chromatin by immunofluorescence. Methods 26:3-9.[CrossRef][Medline]
53. Pina, B., S. Berger, G. A. Marcus, N. Silverman, J. Agapite, and L. Guarente. 1993. ADA3: a gene, identified by resistance to GAL4-VP16, with properties similar to and different from those of ADA2. Mol. Cell. Biol. 13:5981-5989.
54. Pirrotta, V. 1988. Vectors for P-mediated transformation in Drosophila. Biotechnology 10:437-456.[Medline]
55. Platero, J. S., A. K. Csink, A. Quintanilla, and S. Henikoff. 1998. Changes in chromosomal localization of heterochromatin-binding proteins during the cell cycle in Drosophila. J. Cell Biol. 140:1297-1306.
56. Prado, A., I. Canal, and A. Ferrus. 1999. The haplolethal region at the 16F gene cluster of Drosophila melanogaster: structure and function. Genetics 151:163-175.
57. Sass, G. L., and S. Henikoff. 1998. Comparative analysis of position-effect variegation mutations in Drosophila melanogaster delineates the targets of modifiers. Genetics 148:733-741.
58. Sass, G. L., and S. Henikoff. 1999. Pairing-dependent mislocalization of a Drosophila brown gene reporter to a heterochromatic environment. Genetics 152:595-604.
59. Smith, E. R., J. M. Belote, R. L. Schiltz, X. J. Yang, P. A. Moore, S. L. Berger, Y. Nakatani, and C. D. Allis. 1998. Cloning of Drosophila GCN5: conserved features among metazoan GCN5 family members. Nucleic Acids Res. 26:2948-2954.
60. Strahl, B. D., and C. D. Allis. 2000. The language of covalent histone modifications. Nature 403:41-45.[CrossRef][Medline]
61. Struhl, K. 1998. Histone acetylation and transcriptional regulatory mechanisms. Genes Dev. 12:599-606.
62. Struhl, K., and Z. Moqtaderi. 1998. The TAFs in the HAT. Cell 94:1-4.[CrossRef][Medline]
63. Sudarsanam, P., and F. Winston. 2000. The Swi/Snf family nucleosome-remodeling complexes and transcriptional control. Trends Genet. 16:345-351.[CrossRef][Medline]
64. Sun, Y. H., C. J. Tsai, M. M. Green, J. L. Chao, C. T. Yu, T. J. Jaw, J. Y. Yeh, and V. N. Bolshakov. 1995. White as a reporter gene to detect transcriptional silencers specifying position-specific gene expression during Drosophila melanogaster eye development. Genetics 141:1075-1086.[Abstract]
65. Workman, J. L., and R. E. Kingston. 1998. Alteration of nucleosome structure as a mechanism of transcriptional regulation. Annu. Rev. Biochem. 67:545-579.[CrossRef][Medline]
66. Zeng, M., A. Kumar, G. Meng, Q. Gao, G. Dimri, D. Wazer, H. Band, and V. Band. 2002. Human papilloma virus 16 E6 oncoprotein inhibits retinoic X receptor-mediated transactivation by targeting human ADA3 coactivator. J. Biol. Chem. 277:45611-45618.
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