2D gel electrophoresis reveals dynamics of t-loop formation during the cell cycle and t-loop in maintenance regulated by heterochromatin state

Zepeng Zhang, Tianpeng Zhang, Yuanlong Ge, Mengfan Tang, Wenbin Ma, Qinfen Zhang, Shengzhao Gong, Woodring E Wright, Jerry W Shay, Haiying Liu, Yong Zhao

Research output: Contribution to journalArticle

Abstract

Linear chromosome ends are capped by telomeres that have been previously reported to adopt a t-loop structure. The lack of simple methods for detecting t-loops has hindered progress in understanding the dynamics of t-loop formation and its function in protecting chromosome ends. Here, we employed a classical two-dimensional agarose gel method (2D gel method) to innovatively apply to t-loop detection. Briefly, restriction fragments of genomic DNA were separated in a 2D gel, and the telomere sequence was detected by in-gel hybridization with telomeric probe. Using this method, we found that t-loops are present throughout the cell cycle, and t-loop formation tightly couples to telomere replication. We also observed that t-loop abundance positively correlates with chromatin condensation, i.e. cells with less compact telomeric chromatin (ALT cells and trichostatin A (TSA)-treated HeLa cells) exhibited fewer t-loops. Moreover, we observed that telomere dysfunction-induced foci, ALT-associated promyelocytic leukemia bodies, and telomere sister chromatid exchanges are activated upon TSAinduced loss of t-loops. These findings confirm the importance of the t-loop in protecting linear chromosomes from damage or illegitimate recombination.

Original languageEnglish (US)
Pages (from-to)6645-6656
Number of pages12
JournalJournal of Biological Chemistry
Volume294
Issue number16
DOIs
StatePublished - Jan 1 2019

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Heterochromatin
Telomere
Electrophoresis, Gel, Two-Dimensional
Electrophoresis
Cell Cycle
Chromosomes
Gels
Cells
Maintenance
Chromatin
trichostatin A
Sister Chromatid Exchange
Sepharose
Condensation
HeLa Cells
Genetic Recombination
Leukemia
DNA

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

2D gel electrophoresis reveals dynamics of t-loop formation during the cell cycle and t-loop in maintenance regulated by heterochromatin state. / Zhang, Zepeng; Zhang, Tianpeng; Ge, Yuanlong; Tang, Mengfan; Ma, Wenbin; Zhang, Qinfen; Gong, Shengzhao; Wright, Woodring E; Shay, Jerry W; Liu, Haiying; Zhao, Yong.

In: Journal of Biological Chemistry, Vol. 294, No. 16, 01.01.2019, p. 6645-6656.

Research output: Contribution to journalArticle

Zhang, Zepeng ; Zhang, Tianpeng ; Ge, Yuanlong ; Tang, Mengfan ; Ma, Wenbin ; Zhang, Qinfen ; Gong, Shengzhao ; Wright, Woodring E ; Shay, Jerry W ; Liu, Haiying ; Zhao, Yong. / 2D gel electrophoresis reveals dynamics of t-loop formation during the cell cycle and t-loop in maintenance regulated by heterochromatin state. In: Journal of Biological Chemistry. 2019 ; Vol. 294, No. 16. pp. 6645-6656.
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AU - Gong, Shengzhao

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AU - Zhao, Yong

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