Highly expressed loci are vulnerable to misleading ChIP localization of multiple unrelated proteins
Open accessAbstract
Chromatin immunoprecipitation (ChIP) is the gold-standard technique for localizing nuclear proteins in the genome. We used ChIP, in combination with deep sequencing (Seq), to study the genome-wide distribution of the Silent information regulator (Sir) complex in Saccharomyces cerevisiae. We analyzed ChIP-Seq peaks of the Sir2, Sir3, and Sir4 silencing proteins and discovered 238 unexpected euchromatic loci that exhibited enrichment of all three. Surprisingly, published ChIP-Seq datasets for the Ste12 transcription factor and the centromeric Cse4 protein indicated that these proteins were also enriched in the same euchromatic regions with the high Sir protein levels. The 238 loci, termed "hyper-ChIPable", were in highly expressed regions with strong polymerase II and polymerase III enrichment signals, and the correlation between transcription level and ChIP enrichment was not limited to these 238 loci but extended genome-wide. The apparent enrichment of various proteins at hyper-ChIPable loci was not a consequence of artifacts associated with deep sequencing methods, as confirmed by ChIP-quantitative PCR. The localization of unrelated proteins, including the entire silencing complex, to the most highly transcribed genes was highly suggestive of a technical issue with the immunoprecipitations. ChIP-Seq on chromatin immunoprecipitated with a nuclear-localized GFP reproduced the above enrichment in an expression-dependent manner: induction of the GAL genes resulted in an increased ChIP signal of the GFP protein at these loci, with presumably no biological relevance. Whereas ChIP is a broadly valuable technique, some published conclusions based upon ChIP procedures may merit reevaluation in light of these findings.
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It remains unknown what proportion of candidate enhancers identified solely by biochemical marks are technical false positives or have enhancer-like features but no meaningful impact on expression of cis-encoded genes.
“Although unquestionably useful, it remains unknown what proportion of candidate enhancers identified solely by biochemical marks are technical false positives130-132 or a product of having enhancer-like biochemical features but no meaningful impact on the expression of cis encoded genes133.”
From Towards a comprehensive catalogue of validated and target-linked human enhancers (2020) · cites this paper as [130]
ChIP is not equally adept at measuring all interactions.
“While ChIP is not equally adept at measuring all interactions (Baranello et al., 2016; Teytelman et al., 2013), recent methods have overcome some of these limitations (Janssens et al., 2018; Skene and Henikoff, 2017; Skene et al., 2018; Zhu et al., 2019).”
From Unraveling Hematopoiesis through the Lens of Genomics (2020) · cites this paper as Teytelman et al., 2013
Cites
The Sequence Alignment/Map format and SAMtools
Li, H., Handsaker, B., Wysoker, A., Fennell, T., Ruan, J., Homer, N., Marth, G., Abecasis, G., Durbin, R., & 1000 Genome Project Data Processing Subgroup (2009) · Bioinformatics · doi:10.1093/bioinformatics/btp352 · Cited by 62,041 · see citing works
Fast and accurate short read alignment with Burrows-Wheeler transform
Li, H., & Durbin, R. (2009) · Bioinformatics · doi:10.1093/bioinformatics/btp324 · Cited by 58,607 · see citing works
Model-based Analysis of ChIP-Seq (MACS)
Zhang, Y., Liu, T., Meyer, C. A., Eeckhoute, J., Johnson, D. S., Bernstein, B. E., Nusbaum, C., Myers, R. M., Brown, M., Li, W., et al. (2008) · Genome Biology · doi:10.1186/gb-2008-9-9-r137 · Cited by 18,386 · see citing works
Fast and accurate long-read alignment with Burrows-Wheeler transform.
Li, H., & Durbin, R. (2010) · Bioinformatics · doi:10.1093/bioinformatics/btp698 · Cited by 12,063 · see citing works
Longtine, M. S., Mckenzie, A., DeMarini, D. J., Shah, N. G., Wach, A., Brachat, A., Philippsen, P., & Pringle, J. R. (1998) · Yeast · doi:10.1002/(sici)1097-0061(199807)14:10 · Cited by 5,367 · see citing works
Mapping short DNA sequencing reads and calling variants using mapping quality scores
Li, H., Ruan, J., & Durbin, R. (2008) · Genome Research · doi:10.1101/gr.078212.108 · Cited by 2,638 · see citing works
ChIP–seq: advantages and challenges of a maturing technology
Park, P. J. (2009) · Nature Reviews Genetics · doi:10.1038/nrg2641 · Cited by 2,217 · see citing works
Position effect at S. cerevisiae telomeres: Reversible repression of Pol II transcription
Gottschling, D. E., Aparicio, O. M., Billington, B. L., & Zakian, V. A. (1990) · Cell · doi:10.1016/0092-8674(90)90141-z · Cited by 1,325 · see citing works
Furey, T. S. (2012) · Nature Reviews Genetics · doi:10.1038/nrg3306 · Cited by 825 · see citing works
The Establishment, Inheritance, and Function of Silenced Chromatin in Saccharomyces cerevisiae
Rusché, L. N., Kirchmaier, A. L., & Rine, J. (2003) · Annual Review of Biochemistry · doi:10.1146/annurev.biochem.72.121801.161547 · Cited by 726 · see citing works
Self-citationPromoter-specific binding of Rap1 revealed by genome-wide maps of protein–DNA association
Lieb, J. D., Liu, X., Botstein, D., & Brown, P. O. (2001) · Nature Genetics · doi:10.1038/ng569 · Cited by 707 · see citing works
Chromosomal landscape of nucleosome-dependent gene expression and silencing in yeast
Wyrick, J. J., Holstege, F. C. P., Jennings, E. G., Causton, H. C., Shore, D., Grunstein, M. R., Lander, E. S., & Young, R. A. (1999) · Nature · doi:10.1038/46567 · Cited by 394 · see citing works
Aparicio, O. M., Geisberg, J. V., Sekinger, E. A., Yang, A., Moqtaderi, Z., & Struhl, K. (2005) · Current Protocols in Molecular Biology · doi:10.1002/0471142727.mb2103s69 · Cited by 277 · see citing works
ChIP-Seq: technical considerations for obtaining high-quality data
Kidder, B. L., Hu, G., & Zhao, K. (2011) · Nature Immunology · doi:10.1038/ni.2117 · Cited by 245 · see citing works
Facilitated Recycling Pathway for RNA Polymerase III
Dieci, G., & Sentenac, A. (1996) · Cell · doi:10.1016/s0092-8674(00)80979-4 · Cited by 193 · see citing works
Roberts, D. N., Stewart, A. J., Huff, J. T., & Cairns, B. R. (2003) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.2435566100 · Cited by 174 · see citing works
Moqtaderi, Z., & Struhl, K. (2004) · Molecular and Cellular Biology · doi:10.1128/mcb.24.10.4118-4127.2004 · Cited by 168 · see citing works
Systematic evaluation of factors influencing ChIP-seq fidelity
Chen, Y., Negre, N. N., Li, Q., Mieczkowska, J. O., Slattery, M. G., Liu, T., Zhang, Y., Kim, T.-K., He, H. H., Zieba, J. T., et al. (2012) · Nature Methods · doi:10.1038/nmeth.1985 · Cited by 164 · see citing works
ChIP-chip versus ChIP-seq: Lessons for experimental design and data analysis
Ho, J. W. K., Bishop, E. P., Karchenko, P. V., Negre, N. N., White, K. P., & Park, P. J. (2011) · BMC Genomics · doi:10.1186/1471-2164-12-134 · Cited by 156 · see citing works
Efficient yeast ChIP-Seq using multiplex short-read DNA sequencing
Lefrançois, P., Euskirchen, G. M., Auerbach, R. K., Rozowsky, J. S., Gibson, T., Yellman, C. M., Gerstein, M., & Snyder, M. P. (2009) · BMC Genomics · doi:10.1186/1471-2164-10-37 · Cited by 152 · see citing works
Hanlon, S. E., & Lieb, J. D. (2004) · Current Opinion in Genetics & Development · doi:10.1016/j.gde.2004.09.008 · Cited by 101 · see citing works
Radman‐Livaja, M., Ruben, G. J., Weiner, A., et al. (2011) · The EMBO Journal · doi:10.1038/emboj.2011.30 · Cited by 70 · see citing works
Where Does Mediator Bind In Vivo?
Fan, X., & Struhl, K. (2009) · PLoS ONE · doi:10.1371/journal.pone.0005029 · Cited by 69 · see citing works
Genome-wide analysis of functional sirtuin chromatin targets in yeast
Li, M., Valsakumar, V., Poorey, K., Bekiranov, S., & Smith, J. S. (2013) · Genome biology · doi:10.1186/gb-2013-14-5-r48 · Cited by 52 · see citing works
Telomeric silencing of a natural subtelomeric gene
Vega-Palas, M. A., Martín-Figueroa, E., & Florencio, F. J. (2000) · Molecular and General Genetics MGG · doi:10.1007/s004380051170 · Cited by 44 · see citing works
Co-Evolution of Transcriptional Silencing Proteins and the DNA Elements Specifying Their Assembly
Zill, O. A., Scannell, D. R., Teytelman, L., & Rine, J. (2010) · PLoS Biology · doi:10.1371/journal.pbio.1000550 · Cited by 40 · see citing works
Self-citationTelomeric Silencing of an Open Reading Frame in Saccharomyces cerevisiae
Barton, A. B., & Kaback, D. B. (2006) · Genetics · doi:10.1534/genetics.106.058420 · Cited by 15 · see citing works
SIR‐dependent repression of non‐telomeric genes in Saccharomyces cerevisiae?
Marchfelder, U., Rateitschak, K., & Ehrenhofer‐Murray, A. E. (2003) · Yeast · doi:10.1002/yea.1001 · Cited by 10 · see citing works
The Enigmatic Conservation of a Rap1 Binding Site in the Saccharomyces cerevisiae HMR-E Silencer
Teytelman, L., Nishimura, E. O., Özaydın, B., Eisen, M. B., & Rine, J. (2012) · G3 Genes Genomes Genetics · doi:10.1534/g3.112.004077 · Cited by 5 · see citing works
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CUT&Tag for efficient epigenomic profiling of small samples and single cells
Kaya-Okur, H. S., Wu, S. J., Codomo, C. A., Pledger, E. S., Bryson, T. D., Henikoff, J. G., Ahmad, K., & Henikoff, S. (2019) · Nature Communications · doi:10.1038/s41467-019-09982-5 · Cited by 2,187 · see citing works
Transcriptional enhancers: from properties to genome-wide predictions
Shlyueva, D., Stampfel, G., & Stark, A. (2014) · Nature Reviews Genetics · doi:10.1038/nrg3682 · Cited by 1,356 · see citing works
ChEA3: transcription factor enrichment analysis by orthogonal omics integration
Keenan, A. B., Torre, D., Lachmann, A., Leong, A. K., Wojciechowicz, M. L., Utti, V., Jagodnik, K. M., Kropiwnicki, E., Wang, Z., & Ma’ayan, A. (2019) · Nucleic Acids Research · doi:10.1093/nar/gkz446 · Cited by 1,136 · see citing works
Genome-wide binding of the CRISPR endonuclease Cas9 in mammalian cells
Wu, X., Scott, D. A., Kriz, A. J., Chiu, A. C., Hsu, P. D., Dadon, D. B., Cheng, A. W., Trevino, A. E., Konermann, S., Chen, S., et al. (2014) · Nature Biotechnology · doi:10.1038/nbt.2889 · Cited by 944 · see citing works
Targeted in situ genome-wide profiling with high efficiency for low cell numbers
Skene, P. J., Henikoff, J. G., & Henikoff, S. (2018) · Nature Protocols · doi:10.1038/nprot.2018.015 · Cited by 803 · see citing works
Peak calling by Sparse Enrichment Analysis for CUT&RUN chromatin profiling
Meers, M. P., Tenenbaum, D., & Henikoff, S. (2019) · Epigenetics & Chromatin · doi:10.1186/s13072-019-0287-4 · Cited by 556 · see citing works
Absence of a simple code: how transcription factors read the genome
Slattery, M., Zhou, T., Yang, L., Dantas Machado, A. C., Gordân, R., & Rohs, R. (2014) · Trends in Biochemical Sciences · doi:10.1016/j.tibs.2014.07.002 · Cited by 537 · see citing works
Latest Developed Strategies to Minimize the Off-Target Effects in CRISPR-Cas-Mediated Genome Editing
Naeem, M., Majeed, S., Hoque, M. Z., & Ahmad, I. (2020) · Cells · doi:10.3390/cells9071608 · Cited by 457 · see citing works
Towards a comprehensive catalogue of validated and target-linked human enhancers
Gasperini, M., Tome, J. M., & Shendure, J. (2020) · Nature Reviews Genetics · doi:10.1038/s41576-019-0209-0 · Cited by 396 · see citing works
Wang, X., Brandão, H. B., Le, T. B. K., Laub, M. T., & Rudner, D. Z. (2017) · Science · doi:10.1126/science.aai8982 · Cited by 293 · see citing works
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Jain, D., Baldi, S., Zabel, A., Straub, T., & Becker, P. B. (2015) · Nucleic Acids Research · doi:10.1093/nar/gkv637 · Cited by 156 · see citing works
Andersson, R. (2015) · BioEssays · doi:10.1002/bies.201400162 · Cited by 127 · see citing works
ChIP bias as a function of cross-linking time
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Merhej, J., Thiebaut, A., Blugeon, C., Pouch, J., Ali Chaouche, M. E. A., Camadro, J.-M., Le Crom, S., Lelandais, G., & Devaux, F. (2016) · Frontiers in Microbiology · doi:10.3389/fmicb.2016.00645 · Cited by 36 · see citing works
Maintenance of Nucleosomal Balance in<i>cis</i>by Conserved AAA-ATPase Yta7
Lombardi, L. M., Davis, M. D., & Rine, J. (2015) · Genetics · doi:10.1534/genetics.114.168039 · Cited by 16 · see citing works
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- Authors
- Teytelman, L., Thurtle, D. M., Rine, J., & van Oudenaarden, A.
- Affiliations
- Departments of aPhysics and
- Department of Molecular and Cell Biology and California Institute for Quantitative Biosciences, University of California, Berkeley, CA 94720; and
- QB3 (US)
- ORCID
- Rine, Jasper
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 2013-10-30
- Volume
- 110
- Issue
- 46
- Pages
- 18602-18607
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC3831989
- OpenAlex
- W2080466240
- Cited by
- 389
- References
- 29
- Citations by year
- 2026: 6 · 2025: 19 · 2024: 21 · 2023: 21 · 2022: 29 · 2021: 18 · 2020: 47 · 2019: 42 · 2018: 35 · 2017: 29 · 2016: 36 · 2015: 42 · 2014: 40 · 2013: 4
- FWCI
- 18.20
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Licence
- other-oa
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion, other-oa)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Data Archiving and Networked Services (DANS) (submittedVersion)
- DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc)
- KNAW research portal (Royal Academy of Art and Sciences (KNAW)) (submittedVersion)
- Funders
- National Cancer Institute (U54CA143874, DP1CA174420)
- National Institute of General Medical Sciences (GM31105, R01GM068957, R01GM031105, R37GM031105, T32GM007232)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Epigenetics and DNA Methylation
- Keywords
- SIR complex, Sir2, Sir3, Sir4, heterochromatin silencing, transcriptional regulation, Ste12, Cse4, nuclear protein localization, Saccharomyces cerevisiae
- MeSH terms
- Polymerase Chain Reaction, Artifacts, DNA Primers, Silent Information Regulator Proteins, Saccharomyces cerevisiae, Chromatin Immunoprecipitation, Genetic Loci, High-Throughput Nucleotide Sequencing
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