μDamID: A Microfluidic Approach for Joint Imaging and Sequencing of Protein-DNA Interactions in Single Cells

Overview

Journal Cell Syst

Publisher Cell Press

Specialties Biology
Cell Biology
Molecular Biology

Date 2020 Oct 25

PMID 33099405

Citations 9

Authors

Nicolas Altemose

Annie Maslan

Carolina Rios-Martinez

Andre Lai

Jonathan A White

Aaron Streets

Affiliations

Soon will be listed here.

Abstract

DNA adenine methyltransferase identification (DamID) measures a protein's DNA-binding history by methylating adenine bases near each protein-DNA interaction site and then selectively amplifying and sequencing these methylated regions. Additionally, these interactions can be visualized using A-Tracer, a fluorescent protein that binds to methyladenines. Here, we combine these imaging and sequencing technologies in an integrated microfluidic platform (μDamID) that enables single-cell isolation, imaging, and sorting, followed by DamID. We use μDamID and an improved A-Tracer protein to generate paired imaging and sequencing data from individual human cells. We validate interactions between Lamin-B1 protein and lamina-associated domains (LADs), observe variable 3D chromatin organization and broad gene regulation patterns, and jointly measure single-cell heterogeneity in Dam expression and background methylation. μDamID provides the unique ability to compare paired imaging and sequencing data for each cell and between cells, enabling the joint analysis of the nuclear localization, sequence identity, and variability of protein-DNA interactions. A record of this paper's transparent peer review process is included in the Supplemental Information.

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References

Park M, Patel N, Keung A, Khalil A . Engineering Epigenetic Regulation Using Synthetic Read-Write Modules. Cell. 2018; 176(1-2):227-238.e20. PMC: 6329643. DOI: 10.1016/j.cell.2018.11.002. View

Buchwalter A, Kaneshiro J, Hetzer M . Coaching from the sidelines: the nuclear periphery in genome regulation. Nat Rev Genet. 2018; 20(1):39-50. PMC: 6355253. DOI: 10.1038/s41576-018-0063-5. View

Southall T, Gold K, Egger B, Davidson C, Caygill E, Marshall O . Cell-type-specific profiling of gene expression and chromatin binding without cell isolation: assaying RNA Pol II occupancy in neural stem cells. Dev Cell. 2013; 26(1):101-12. PMC: 3714590. DOI: 10.1016/j.devcel.2013.05.020. View

Essletzbichler P, Konopka T, Santoro F, Chen D, Gapp B, Kralovics R . Megabase-scale deletion using CRISPR/Cas9 to generate a fully haploid human cell line. Genome Res. 2014; 24(12):2059-65. PMC: 4248322. DOI: 10.1101/gr.177220.114. View

Guelen L, Pagie L, Brasset E, Meuleman W, Faza M, Talhout W . Domain organization of human chromosomes revealed by mapping of nuclear lamina interactions. Nature. 2008; 453(7197):948-51. DOI: 10.1038/nature06947. View

Aughey G, Gomez A, Thomson J, Yin H, Southall T . CATaDa reveals global remodelling of chromatin accessibility during stem cell differentiation in vivo. Elife. 2018; 7. PMC: 5826290. DOI: 10.7554/eLife.32341. View

Yuan J, Sheng J, Sims P . SCOPE-Seq: a scalable technology for linking live cell imaging and single-cell RNA sequencing. Genome Biol. 2018; 19(1):227. PMC: 6305572. DOI: 10.1186/s13059-018-1607-x. View

Johnson D, Mortazavi A, Myers R, Wold B . Genome-wide mapping of in vivo protein-DNA interactions. Science. 2007; 316(5830):1497-502. DOI: 10.1126/science.1141319. View

Rodriguez-Mateos P, Azevedo N, Almeida C, Pamme N . FISH and chips: a review of microfluidic platforms for FISH analysis. Med Microbiol Immunol. 2020; 209(3):373-391. PMC: 7248050. DOI: 10.1007/s00430-019-00654-1. View

10.

Grosselin K, Durand A, Marsolier J, Poitou A, Marangoni E, Nemati F . High-throughput single-cell ChIP-seq identifies heterogeneity of chromatin states in breast cancer. Nat Genet. 2019; 51(6):1060-1066. DOI: 10.1038/s41588-019-0424-9. View

11.

Skene P, Henikoff J, Henikoff S . Targeted in situ genome-wide profiling with high efficiency for low cell numbers. Nat Protoc. 2018; 13(5):1006-1019. DOI: 10.1038/nprot.2018.015. View

12.

Kim S, De Jonghe J, Kulesa A, Feldman D, Vatanen T, Bhattacharyya R . High-throughput automated microfluidic sample preparation for accurate microbial genomics. Nat Commun. 2017; 8:13919. PMC: 5290157. DOI: 10.1038/ncomms13919. View

13.

Huang B, Babcock H, Zhuang X . Breaking the diffraction barrier: super-resolution imaging of cells. Cell. 2010; 143(7):1047-58. PMC: 3272504. DOI: 10.1016/j.cell.2010.12.002. View

14.

Singh J, Klar A . Active genes in budding yeast display enhanced in vivo accessibility to foreign DNA methylases: a novel in vivo probe for chromatin structure of yeast. Genes Dev. 1992; 6(2):186-96. DOI: 10.1101/gad.6.2.186. View

15.

Shen J, Jiang D, Fu Y, Wu X, Guo H, Feng B . H3K4me3 epigenomic landscape derived from ChIP-Seq of 1,000 mouse early embryonic cells. Cell Res. 2014; 25(1):143-7. PMC: 4650579. DOI: 10.1038/cr.2014.119. View

16.

Jakobsen J, Bagger F, Hasemann M, Schuster M, Frank A, Waage J . Amplification of pico-scale DNA mediated by bacterial carrier DNA for small-cell-number transcription factor ChIP-seq. BMC Genomics. 2015; 16:46. PMC: 4328043. DOI: 10.1186/s12864-014-1195-4. View

17.

Lenain C, de Graaf C, Pagie L, Visser N, De Haas M, de Vries S . Massive reshaping of genome-nuclear lamina interactions during oncogene-induced senescence. Genome Res. 2017; 27(10):1634-1644. PMC: 5630027. DOI: 10.1101/gr.225763.117. View

18.

Li D, Hsu S, Purushotham D, Sears R, Wang T . WashU Epigenome Browser update 2019. Nucleic Acids Res. 2019; 47(W1):W158-W165. PMC: 6602459. DOI: 10.1093/nar/gkz348. View

19.

Rooijers K, Markodimitraki C, Rang F, de Vries S, Chialastri A, de Luca K . Simultaneous quantification of protein-DNA contacts and transcriptomes in single cells. Nat Biotechnol. 2019; 37(7):766-772. PMC: 6609448. DOI: 10.1038/s41587-019-0150-y. View

20.

Bolger A, Lohse M, Usadel B . Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014; 30(15):2114-20. PMC: 4103590. DOI: 10.1093/bioinformatics/btu170. View