An APEX2 proximity ligation method for mapping interactions with the nuclear lamina

The nuclear lamina (NL) is a meshwork found beneath the inner nuclear membrane. The study of the NL is hindered by the insolubility of the meshwork and has driven the development of proximity ligation methods to identify the NL-associated/proximal proteins, RNA, and DNA. To simplify and improve temp...

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Published in:The Journal of cell biology Vol. 220; no. 1
Main Authors: Tran, Joseph R, Paulson, Danielle I, Moresco, James J, Adam, Stephen A, Yates, John R, Goldman, Robert D, Zheng, Yixian
Format: Journal Article
Language:English
Published: United States 04.01.2021
Subjects:
3' Untranslated Regions - genetics
Base Sequence
DNA-(Apurinic or Apyrimidinic Site) Lyase - chemistry
DNA-(Apurinic or Apyrimidinic Site) Lyase - metabolism
Endonucleases - chemistry
Endonucleases - metabolism
HCT116 Cells
HEK293 Cells
Humans
K562 Cells
Lamin Type B - metabolism
Multifunctional Enzymes - chemistry
Multifunctional Enzymes - metabolism
Nuclear Lamina - metabolism
Protein Domains
Protein Interaction Mapping
Proteome - metabolism
RNA - metabolism
RNA Splicing - genetics
ISSN:1540-8140, 1540-8140
Online Access:Get more information
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Summary:The nuclear lamina (NL) is a meshwork found beneath the inner nuclear membrane. The study of the NL is hindered by the insolubility of the meshwork and has driven the development of proximity ligation methods to identify the NL-associated/proximal proteins, RNA, and DNA. To simplify and improve temporal labeling, we fused APEX2 to the NL protein lamin-B1 to map proteins, RNA, and DNA. The identified NL-interacting/proximal RNAs show a long 3' UTR bias, a finding consistent with an observed bias toward longer 3' UTRs in genes deregulated in lamin-null cells. A C-rich motif was identified in these 3' UTR. Our APEX2-based proteomics identifies a C-rich motif binding regulatory protein that exhibits altered localization in lamin-null cells. Finally, we use APEX2 to map lamina-associated domains (LADs) during the cell cycle and uncover short, H3K27me3-rich variable LADs. Thus, the APEX2-based tools presented here permit identification of proteomes, transcriptomes, and genome elements associated with or proximal to the NL.
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ISSN:1540-8140
1540-8140
DOI:10.1083/jcb.202002129