C-BERST: defining subnuclear proteomic landscapes at genomic elements with dCas9-APEX2

Xin D Gao; Li-Chun Tu; Aamir Mir; Tomás Rodriguez; Yuehe Ding; John Leszyk; Job Dekker; Scott A Shaffer; Lihua Julie Zhu; Scot A Wolfe; Erik J Sontheimer

doi:10.1038/s41592-018-0006-2

C-BERST: defining subnuclear proteomic landscapes at genomic elements with dCas9-APEX2

Nat Methods. 2018 Jun;15(6):433-436. doi: 10.1038/s41592-018-0006-2. Epub 2018 May 7.

Authors

Xin D Gao¹, Li-Chun Tu¹, Aamir Mir¹, Tomás Rodriguez¹, Yuehe Ding¹, John Leszyk^{2

3}, Job Dekker^{3

4

5}, Scott A Shaffer^{2

3}, Lihua Julie Zhu^{6

7

8}, Scot A Wolfe^{3

6}, Erik J Sontheimer^{9

10}

Affiliations

¹ RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, MA, USA.
² Proteomics and Mass Spectrometry Facility, University of Massachusetts Medical School, Worcester, MA, USA.
³ Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, MA, USA.
⁴ Program in Systems Biology, University of Massachusetts Medical School, Worcester, MA, USA.
⁵ Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, MA, USA.
⁶ Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, USA.
⁷ Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School, Worcester, MA, USA.
⁸ Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA, USA.
⁹ RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, MA, USA. erik.sontheimer@umassmed.edu.
¹⁰ Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA, USA. erik.sontheimer@umassmed.edu.

Abstract

Mapping proteomic composition at distinct genomic loci in living cells has been a long-standing challenge. Here we report that dCas9-APEX2 biotinylation at genomic elements by restricted spatial tagging (C-BERST) allows the rapid, unbiased mapping of proteomes near defined genomic loci, as demonstrated for telomeres and centromeres. C-BERST enables the high-throughput identification of proteins associated with specific sequences, thereby facilitating annotation of these factors and their roles.

Publication types

Research Support, N.I.H., Extramural

MeSH terms

CRISPR-Associated Protein 9 / metabolism*
Cell Line, Tumor
Chromosome Mapping
DNA-(Apurinic or Apyrimidinic Site) Lyase / metabolism*
Endonucleases
Gene Expression Regulation
Genome
Genomics
Humans
Multifunctional Enzymes
Protein Engineering
Proteome
Proteomics / methods*

Substances

Multifunctional Enzymes
Proteome
CRISPR-Associated Protein 9
Endonucleases
APEX2 protein, human
DNA-(Apurinic or Apyrimidinic Site) Lyase

Abstract

Publication types

MeSH terms

Substances

Grants and funding