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Fly 4:4, 299-301; October/November/December 2010; © 2010 Landes Bioscience
Positive regulation of euchromatic gene expression by HP1 Lucia Piacentini and Sergio Pimpinelli* Istituto Pasteur; Fondazione Cenci Bolognetti and Dipartimento di Genetica e Biologia molecolare; Università “La Sapienza”; Roma, Italy
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Key words: HP1, gene expression, hnRNPs, heterochromatin, PEV, Drosophila Submitted: 05/06/10 Revised: 08/03/10 Accepted: 08/03/10 Previously published online: www.landesbioscience.com/journals/fly/ article/13261 DOI: 10.4161/fly.4.4.13261 *Correspondence to: Sergio Pimpinelli; Email:
[email protected] Addendum to: Piacentini L, Fanti L, Negri R, Del Vescovo V, Fatica A, Altieri F, Pimpinelli S. Heterochromatin protein 1 (HP1a) positively regulates euchromatic gene expression through RNA transcript association and interaction with hnRNPs in Drosophila. PLoS Genet. 2009 Oct;5(10):e1000670; PMCID: PMC2782133; DOI: 10.1371/journal.pgen.1000769.
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P1 is a conserved prototype p rotein that plays an essential role in heterochromatin formation and epigenetic gene silencing through its interaction with histone methyltransferase enzymes (HMTases) and the histone H3 at lysine 9 (H3-MeK9). HP1 is also involved in telomere capping and, more surprisingly, in positive regulation of gene expression. Recently, a wide expression analysis, using a RIP-chip assays (RNAimmunoprecipitation on microarrays), has shown that HP1 associates with the transcripts of more than one hundred euchromatic genes and interacts with the heterogeneous nuclear ribonucleoproteins (hnRNPs) that are known to be involved in RNA processing. By these results, HP1 seems to be part of a complex that stabilizes RNA transcripts. Though previously unsuspected, it was also found that HP1-interacting hnRNPs have a functional role in heterochromatin formation. These proteins bind heterochromatin and are dominant suppressors of position effect variegation. Taken together, the results in the paper by Piacentini et al. open a window on a possible new conceptual landscape in which similar epigenetic mechanisms could have a significant role, both in the metabolism of RNA transcripts and in heterochromatin formation, producing opposite functional effects. These data seem to establish a functional link between euchromatin and heterochromatin. Besides the most commonly cited role of HP1 in heterochromatin formation, gene silencing1-5 and telomere capping,6 recent studies have shown that HP1 is present at multiple active euchromatic loci along
the polytene chromosomes of D. melanogaster, including developmental and heat shock-induced puffs7,8 (Fig. 1). The removal of HP1 from euchromatic sites by in vivo RNase treatment also indicates its association with nascent RNA transcripts.7 Most recently, it has been shown that HP1 associates with the active form of Pol II Phospho Ser2 and binds nascent transcripts by its chromodomain.9 A wide genome screening using a RIPchip assay (RNA-immunoprecipitation on microarrays) in S2 cells identified more than 100 genuine HP1 transcript targets; although these transcripts were identified in cultured somatic cells, their genes overlap, for the most part, with HP1 binding sites observed along the polytene chromosomes of larval salivary glands.9 An expression analysis, by real time RT-PCR, of a sample of genes corresponding to the HP1 target transcripts, showed a significant reduction in transcript amounts in both S2 cells lacking HP1 and mutant larvae expressing an inactive form of HP1 chromodomain.9 This approach permitted, for the first time, the systematic identification of the direct targets of HP1 in the euchromatin and HP1’s positive regulatory role on the corresponding genes.9 A positive role of HP1 in gene expression was also suggested by further analyses showing that HP1 colocalizes on polytene chromosomes and coprecipitates with different classes of hnRNPs.9 These include the multi-KH-domain vigilin DDP1,10 the Drosophila homolog to mammalian A/B type hnRNP HRB87F,11 and the zinc finger protein PEP (Peptide on Ecdysone Puffs),12 all capable of binding DNA and, with higher affinity, RNA. In addition, genetic and biochemical experiments have demonstrated that all
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these proteins form an HP1-containing hnRNP nuclear sub-complex.9 Since these hnRNPs seem to play a central role in RNA packaging and stability, it is probable that the positive role of HP1 in gene
expression depends on its involvement in these two functions. In support of this idea, it has been found that, after blocking transcription with actinomycin D, the HP1a target transcripts are less stable
Figure 1. Immunopattern of HP1 localization (green signals) along polytene chromosomes (red) of Drosophila melanogaster. The arrowheads indicated some of the many euchromatic HP1 immunosignals, including developmental puffs (yellow arrowheads) and the arrows indicated the telomeric HP1 immunosignals. Chr = chromocenter.
in S2 cells lacking HP1a compared to control cells.9 Complementary to these conclusions, the study has intriguingly shown that HRB87F and PEP proteins, like DDP1 and HP1, are also located on the heterochromatic chromocenter of polytene chromosomes, where they probably bind both RNA and single-stranded DNA, since they are partially removed from the chromocenter of polytene chromosomes after in vivo treatments with RNase.9 Importantly, these proteins, like DDP1, are dominant suppressors of heterochromatin-induced gene silencing.9 This strongly suggests that the HP1-containing hnRNP sub-complex also plays a novel and unexpected role in heterochromatin formation. In conclusion, these results reveal, for the first time, new and unexpected functional properties of HP1 and hnRNPs: together these proteins seem to be involved in both heterochromatin formation and positive regulation of gene expression. What molecular mechanisms are responsible for the functional versatility of these proteins? It seems reasonable to propose that their main function is nucleic acid compaction (Fig. 2). As shown in Figure 3, DNA
Figure 2. Diagramatic representation of a tentative model suggesting that a HP1 and hnRNP proteins complex performs DNA packaging in the chromocenter and RNA packaging during transcription. 300
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Figure 3. The cartoon represents the functional consequences of nucleic acid packaging. The DNA packaging in the chromocenter should cause the formation of heterochromatic domains while the packaging and stabilization of RNA transcripts, should reinforce gene expression.
compaction results in heterochromatin formation and probably gene silencing, while RNA compaction and then RNA stabilization, reinforces gene expression. The sharing of similar protein complexes also suggests a possible functional interdependency between heterochromatin and euchromatic gene activity. Their quantitative balance between heterochromatin and RNA transcripts could be one of the mechanisms underlying a fine regulation of gene expression by modulating RNA transcripts stability. On the other hand, although by different mechanistic point of view, it has been already suggested that gene expression could depend on the dynamic interplay between heterochromatin and euchromatin domains.13
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8. Fanti L, Berloco M, Piacentini L, Pimpinelli S. Chromosomal distribution of heterochromatin protein 1 (HP1) in Drosophila: a cytological map of euchromatic HP1 binding sites. Genetica 2003; 117:135-47. 9. Piacentini L, Fanti L, Negri R, Del Vescovo V, Fatica A, Altieri F, et al. Heterochromatin protein 1 (HP1a) positively regulates euchromatic gene expression through RNA transcript association and interaction with hnRNPs in Drosophila. PLoS Genet 2009; 5:1000670. 10. Cortés A, Huertas D, Fanti L, Pimpinelli S, Marsellach FX, Piña B, et al. DDP1, a single-stranded nucleic acid-binding protein of Drosophila, associates with pericentric heterochromatin and is functionally homologous to the yeast Scp160p, which is involved in the control of cell ploidy. EMBO J 1999; 18:3820-33. 11. Haynes SR, Johnson D, Raychaudhuri G, Beyer AL. The Drosophila Hrb87F gene encodes a new member of the A and B hnRNP protein group. Nucleic Acids Res 1991; 19:25-31. 12. Amero SA, Elgin SC, Beyer AL. A unique zinc finger protein is associated preferentially with active ecdysone-responsive loci in Drosophila. Genes Dev 1991; 5:188-200. 13. Girton JR, Johansen KM. Chromatin structure and the regulation of gene expression: the lessons of PEV in Drosophila. Adv Genet 2008; 61:1-43.
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