The COOH-terminal Domain of the JIL-1 Histone H3S10 Kinase Interacts with Histone H3 and Is Required for Correct Targeting to Chromatin

Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa 50011, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 10/2008; 283(47):32741-50. DOI: 10.1074/jbc.M806227200
Source: PubMed


The JIL-1 histone H3S10 kinase in Drosophila localizes specifically to euchromatic interband regions of polytene chromosomes and is enriched 2-fold on the male X chromosome.
JIL-1 can be divided into four main domains including an NH2-terminal domain, two separate kinase domains, and a COOH-terminal domain. Our results demonstrate that the COOH-terminal
domain of JIL-1 is necessary and sufficient for correct chromosome targeting to autosomes but that both COOH- and NH2-terminal sequences are necessary for enrichment on the male X chromosome. We furthermore show that a small 53-amino acid
region within the COOH-terminal domain can interact with the tail region of histone H3, suggesting that this interaction is
necessary for the correct chromatin targeting of the JIL-1 kinase. Interestingly, our data indicate that the COOH-terminal
domain alone is sufficient to rescue JIL-1 null mutant polytene chromosome defects including those of the male X chromosome. Nonetheless, we also found that a truncated
JIL-1 protein which was without the COOH-terminal domain but retained histone H3S10 kinase activity was able to rescue autosome
as well as partially rescue male X polytene chromosome morphology. Taken together these findings indicate that JIL-1 may participate
in regulating chromatin structure by multiple and partially redundant mechanisms.

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Available from: Weiguo Zhang, Apr 17, 2015
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    • "To explore this possibility, we modeled the 3D structure of JIL-1 using the I-TASSER structure prediction program (37) and compared it to the nucleosome crystal structure (41). As illustrated in Supplementary Figure S4, considering that JIL-1 is known to bind the tail of H3 by the end of its carboxy-terminal domain (42) and that its folded structure is larger than a nucleosome, it is likely to have the capacity to phosphorylate histone H3 of one or more nucleosomes some distance away from its actual binding site depending on the state of higher order nucleosome packaging. Thus, as indicated by the present data, the distribution of JIL-1 and H3S10 phosphorylation may not necessarily be coincident. "
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    ABSTRACT: In this study we have determined the genome-wide relationship of JIL-1 kinase mediated H3S10 phosphorylation with gene expression and the distribution of the epigenetic H3K9me2 mark. We show in wild-type salivary gland cells that the H3S10ph mark is predominantly enriched at active genes whereas the H3K9me2 mark is largely associated with inactive genes. Comparison of global transcription profiles in salivary glands from wild-type and JIL-1 null mutant larvae revealed that the expression levels of 1539 genes changed at least 2-fold in the mutant and that a substantial number (49%) of these genes were upregulated whereas 51% were downregulated. Furthermore, the results showed that downregulation of genes in the mutant was correlated with higher levels or acquisition of the H3K9me2 mark whereas upregulation of a gene was correlated with loss of or diminished H3K9 dimethylation. These results are compatible with a model where gene expression levels are modulated by the levels of the H3K9me2 mark independent of the state of the H3S10ph mark, which is not required for either transcription or gene activation to occur. Rather, H3S10 phosphorylation functions to indirectly maintain active transcription by counteracting H3K9 dimethylation and gene silencing.
    Full-text · Article · Mar 2014 · Nucleic Acids Research
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    • "While difficult to discern because of the grossly perturbed chromosome morphology [15] such discrete labeling was also present in the JIL-1 null background (Fig. 1B). Thus, in order to verify this we expressed a CFP-tagged JIL-1 carboxy-terminal construct (JIL-1-CTD-CFP) in the JIL-1 null background that rescues chromosome morphology to near wild-type without JIL-1 kinase activity [27]. As illustrated in Fig. 1C discrete bands similar to those of wild-type preparations were clearly present under these conditions indicating that they were not caused by JIL-1 kinase activity. "
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    ABSTRACT: JIL-1 is the major kinase controlling phosphorylation of histone H3S10 and has been demonstrated to function to counteract heterochromatization and gene silencing. However, an alternative model has been proposed in which JIL-1 is required for transcription to occur, additionally phosphorylates H3S28, and recruits 14-3-3 to active genes. Since these findings are incompatible with our previous demonstration that there are robust levels of transcription in the complete absence of JIL-1 and that JIL-1 is not present at developmental or heat shock-induced polytene chromosome puffs, we have reexamined JIL-1's possible role in H3S28 phosphorylation and 14-3-3 recruitment. Using two different H3S28ph antibodies we show by immunocytochemistry and immunoblotting that in Drosophila the H3S28ph mark is not present at detectable levels above background on polytene chromosomes at interphase but only on chromosomes at pro-, meta-, and anaphase during cell division in S2 cells and third instar larval neuroblasts. Moreover, this mitotic H3S28ph signal is also present in a JIL-1 null mutant background at undiminished levels suggesting that JIL-1 is not the mitotic H3S28ph kinase. We also demonstrate that H3S28ph is not enriched at heat shock puffs. Using two different pan-specific 14-3-3 antibodies as well as an enhancer trap 14-3-3ε-GFP line we show that 14-3-3, while present in salivary gland nuclei, does not localize to chromosomes but only to the nuclear matrix surrounding the chromosomes. In our hands 14-3-3 is not recruited to developmental or heat shock puffs. Furthermore, using a lacO repeat tethering system to target LacI-JIL-1 to ectopic sites on polytene chromosomes we show that only H3S10ph is present and upregulated at such sites, not H3S28ph or 14-3-3. Thus, our results argue strongly against a model where JIL-1 is required for H3S28 phosphorylation and 14-3-3 recruitment at active genes.
    Full-text · Article · Apr 2013 · PLoS ONE
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    • "JIL-1 z2 /JIL-1 z2 flies, viability was restored to 65% (Table S4), strongly indicating that the complete lethality of JIL-1 z2 /JIL-1 z2 flies (Table 1) is not due to a second site lethal. These results were confirmed in crosses with the hsp70 promoter-driven full-length JIL- 1-V5 transgene, JIL-1-FL (Bao et al. 2008). In these crosses, viability was restored to 54.8% at 25° (Table S5) and to 94.2% at 21° (Table S6). "
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    ABSTRACT: The essential JIL-1 histone H3S10 kinase is a key regulator of chromatin structure that functions to maintain euchromatic domains while counteracting heterochromatization and gene silencing. In the absence of the JIL-1 kinase, two of the major heterochromatin markers H3K9me2 and HP1a spread in tandem to ectopic locations on the chromosome arms. Here we address the role of the third major heterochromatin component, the zinc-finger protein Su(var)3-7. We show that the lethality but not the chromosome morphology defects associated with the null JIL-1 phenotype to a large degree can be rescued by reducing the dose of the Su(var)3-7 gene and that Su(var)3-7 and JIL-1 loss-of-function mutations have an antagonistic and counterbalancing effect on position-effect variegation (PEV). Furthermore, we show that in the absence of JIL-1 kinase activity, Su(var)3-7 gets redistributed and upregulated on the chromosome arms. Reducing the dose of the Su(var)3-7 gene dramatically decreases this redistribution; however, the spreading of H3K9me2 to the chromosome arms was unaffected, strongly indicating that ectopic Su(var)3-9 activity is not a direct cause of lethality. These observations suggest a model where Su(var)3-7 functions as an effector downstream of Su(var)3-9 and H3K9 dimethylation in heterochromatic spreading and gene silencing that is normally counteracted by JIL-1 kinase activity.
    Full-text · Article · May 2010 · Genetics
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