Related Experiment Video
Updated: Apr 21, 2026
Quantification of γH2AX Foci in Response to Ionising Radiation
Published on: April 6, 2010
TPX2 impacts acetylation of histone H4 at lysine 16: implications for DNA damage response
Gernot Neumayer1, Minh Dang Nguyen1
1Departments of Clinical Neurosciences, Cell Biology & Anatomy, Biochemistry & Molecular Biology, and Hotchkiss Brain Institute, University of Calgary, Cumming School of Medicine, Calgary, Alberta, Canada.
Abstract:
During interphase, the spindle assembly factor TPX2 is compartmentalized in the nucleus where its roles remain largely uncharacterized. Recently, we found that TPX2 regulates the levels of serine 139-phosphoryated H2AX (γ-H2AX) at chromosomal breaks induced by ionizing radiation. Here, we report that TPX2 readily associates with the chromatin in the absence of ionizing radiation. Overexpression of TPX2 alters the DAPI staining pattern of interphase cells and depletion of TPX2 constitutively decreases the levels of histone H4 acetylated at lysine16 (H4K16ac) during G1-phase. Upon ionizing irradiation, this constitutive TPX2 depletion-dependent decrease in H4K16ac levels correlates with increased levels of γ-H2AX. The inversely correlated levels of H4K16ac and γ-H2AX can also be modified by altering the levels of SIRT1, herein identified as a novel protein complex partner of TPX2. Furthermore, we find that TPX2 depletion also interferes with formation of 53BP1 ionizing radiation-induced foci, known to depend on γ-H2AX and the acetylation status of H4K16. In brief, our study is the first indication of a constitutive control of TPX2 on H4K16ac levels, with potential implications for DNA damage response.
Insights
TPX2 protein controls histone acetylation levels in the G1 phase, impacting DNA damage repair. Its depletion reduces H4K16ac, increasing DNA damage marker γ-H2AX.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- TPX2 is a spindle assembly factor primarily studied in mitosis.
- Nuclear roles of TPX2 during interphase are largely unknown.
- Previous work linked TPX2 to regulating γ-H2AX at DNA breaks.
Purpose of the Study:
- Investigate the constitutive role of TPX2 in interphase chromatin.
- Determine TPX2's impact on histone modifications and DNA damage response.
- Identify novel TPX2 interacting partners.
Main Methods:
- Chromatin association assays
- Overexpression and depletion studies of TPX2
- Analysis of histone modifications (H4K16ac) and DNA damage markers (γ-H2AX, 53BP1 foci)
- Co-immunoprecipitation to identify protein partners
Main Results:
- TPX2 associates with chromatin independently of DNA damage.
- TPX2 depletion constitutively reduces H4K16ac in G1 phase.
- Reduced H4K16ac correlates with increased γ-H2AX upon irradiation.
- SIRT1 identified as a novel TPX2 binding partner.
- TPX2 depletion impairs 53BP1 foci formation.
Conclusions:
- TPX2 plays a novel, constitutive role in regulating H4K16ac during interphase.
- TPX2's function in H4K16ac regulation has implications for DNA damage response pathways.
- TPX2 and SIRT1 interaction may be critical for maintaining chromatin integrity and DNA repair.
Related Concept Videos
02:25Spreading of Chromatin Modifications
Writers
The writer...
02:32Histone Modification
Acetylation
The enzyme histone acetyltransferase adds acetyl group to the histones. Another enzyme, histone...
02:36DNA Damage can Stall the Cell Cycle
02:36DNA Damage Can Stall the Cell Cycle
01:38Nucleotide Excision Repair
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
01:08Nucleotide Excision Repair