John Danzer
Mentor: Lori Wallrath, Ph.D.
Lab Phone: 335-7916
Tethered HP1 generates silent chromatin in transcriptionally permissive regions of the genome
Gene silencing by heterochromatin may involve the spread of non-histone chromatin proteins along the chromosome that correlates with alterations in chromatin structure that antagonize transcription. Heterochromatin Protein 1 (HP1) is a key player in this process. HP1 localizes to centric regions, telomeric regions, along the fourth chromosome, and at specific sites within euchromatin. The current model for HP1 association at centric regions involves the methylation of lysine nine of histone H3 by SU(VAR)3-9. This methylation serves as a docking site for the chromo domain of HP1. To determine whether HP1 is sufficient to nucleate the formation of silent chromatin in the absence of the methyl mark, we utilized a LacI tethering system. HP1 was fused to the DNA binding domain of the LacI repressor and expressed in flies carrying reporter genes wherein the lac operator repeats were located two and five kilobases upstream of two heat shock reporter genes, hsp26 and hsp70, respectively. Tethering HP1 at five independent insertion sites within transcriptionally permissive euchromatic regions caused a 20- to 30- fold decrease in heat shock activated gene expression. Tethering a control LacI-GFP fusion protein had no effect on heat shock activation. The decreased level of gene expression correlates with alterations in chromatin structure. Heat shock elements within the hsp26 promoter region are less accessible to digestion in isolated nuclei when HP1 is tethered, compared to non-tethered conditions, or when LacI-GFP is bound. In conclusion, long range gene silencing (5 kb) is observed in transcriptionally permissive regions of the genome upon tethering HP1. Thus, HP1 is sufficient to form silent chromatin in the absence of the initial epigenetic methylation mark and repetitive DNA sequences typically found in centric heterochromatin.
Abstracts:
1999 - DyrkH, a new heterochromatic gene? J.R. Danzer. J.C. Limas, D.E. Cryderman. L.L. Wallrath. Midwest Drosophila Research Conference. Allerton, IL.
2001 - DyrkH, a novel gene located near the 4th chromosome telomere, exhibits characteristics of heterchromatic genes. J.R. Danzer, D.E.Cryderman, J.C. Limas, J. Holmes, L.L. Wallrath. 42th Annual Drosophila Research Conference. Washington D.C. Poster #272B.
2003 - Tethered HP1 generates silent chromatin in transcriptionally permissive regions. J.R. Danzer, Y. Li, L.L. Wallrath. 44th Annual Drosophila Research Conference. Chicago, Poster # 274A.
2000 - Gene regulation from a heterochromatic point of view. J.R. Danzer, D.E.Cryderman, J.C. Limas, J. Holmes, L.L. Wallrath. Midwest Drosophila Research Conference. Allerton, IL.
2002 - Laying a foundation for heterochromatic gene silencing. J.R. Danzer, Y. Li, L.L. Wallrath. Midwest Drosophila Research Conference. Allerton, IL.
Publications:
Danzer JR, Wallrath LL. Mechanisms of HP1-mediated gene silencing in Drosophila. Development. 2004 Aug;131(15):3571-80. Epub 2004 Jun 23. PubMed PMID:15215206.
Li Y, Danzer JR, Alvarez P, Belmont AS, Wallrath LL. Effects of tethering HP1 to euchromatic regions of the Drosophila genome. Development. 2003 May;130(9):1817-24. PubMed PMID: 12642487.
Lab Phone: 335-7916
Tethered HP1 generates silent chromatin in transcriptionally permissive regions of the genome
Gene silencing by heterochromatin may involve the spread of non-histone chromatin proteins along the chromosome that correlates with alterations in chromatin structure that antagonize transcription. Heterochromatin Protein 1 (HP1) is a key player in this process. HP1 localizes to centric regions, telomeric regions, along the fourth chromosome, and at specific sites within euchromatin. The current model for HP1 association at centric regions involves the methylation of lysine nine of histone H3 by SU(VAR)3-9. This methylation serves as a docking site for the chromo domain of HP1. To determine whether HP1 is sufficient to nucleate the formation of silent chromatin in the absence of the methyl mark, we utilized a LacI tethering system. HP1 was fused to the DNA binding domain of the LacI repressor and expressed in flies carrying reporter genes wherein the lac operator repeats were located two and five kilobases upstream of two heat shock reporter genes, hsp26 and hsp70, respectively. Tethering HP1 at five independent insertion sites within transcriptionally permissive euchromatic regions caused a 20- to 30- fold decrease in heat shock activated gene expression. Tethering a control LacI-GFP fusion protein had no effect on heat shock activation. The decreased level of gene expression correlates with alterations in chromatin structure. Heat shock elements within the hsp26 promoter region are less accessible to digestion in isolated nuclei when HP1 is tethered, compared to non-tethered conditions, or when LacI-GFP is bound. In conclusion, long range gene silencing (5 kb) is observed in transcriptionally permissive regions of the genome upon tethering HP1. Thus, HP1 is sufficient to form silent chromatin in the absence of the initial epigenetic methylation mark and repetitive DNA sequences typically found in centric heterochromatin.
Abstracts:
1999 - DyrkH, a new heterochromatic gene? J.R. Danzer. J.C. Limas, D.E. Cryderman. L.L. Wallrath. Midwest Drosophila Research Conference. Allerton, IL.
2001 - DyrkH, a novel gene located near the 4th chromosome telomere, exhibits characteristics of heterchromatic genes. J.R. Danzer, D.E.Cryderman, J.C. Limas, J. Holmes, L.L. Wallrath. 42th Annual Drosophila Research Conference. Washington D.C. Poster #272B.
2003 - Tethered HP1 generates silent chromatin in transcriptionally permissive regions. J.R. Danzer, Y. Li, L.L. Wallrath. 44th Annual Drosophila Research Conference. Chicago, Poster # 274A.
2000 - Gene regulation from a heterochromatic point of view. J.R. Danzer, D.E.Cryderman, J.C. Limas, J. Holmes, L.L. Wallrath. Midwest Drosophila Research Conference. Allerton, IL.
2002 - Laying a foundation for heterochromatic gene silencing. J.R. Danzer, Y. Li, L.L. Wallrath. Midwest Drosophila Research Conference. Allerton, IL.
Publications:
Danzer JR, Wallrath LL. Mechanisms of HP1-mediated gene silencing in Drosophila. Development. 2004 Aug;131(15):3571-80. Epub 2004 Jun 23. PubMed PMID:15215206.
Li Y, Danzer JR, Alvarez P, Belmont AS, Wallrath LL. Effects of tethering HP1 to euchromatic regions of the Drosophila genome. Development. 2003 May;130(9):1817-24. PubMed PMID: 12642487.
Honors and Awards
- 1999 - DyrkH, a new heterochromatic gene? J.R. Danzer. J.C. Limas, D.E. Cryderman. L.L. Wallrath. Mi