Detecting Telomeric DNA Damage by Immuno-Telo FISH.
Marianna Paulis1,2, Ombretta Garbarino1, Francesca Faggioli3,4
1Institute of Genetics and Biomedical Research, UoS of Milan, National Research Council, Milan, Italy.
Methods in Molecular Biology (Clifton, N.J.)
|March 14, 2025
Summary
This study introduces a combined immuno-fluorescence in situ hybridization (FISH) method to detect DNA damage at telomeres, crucial for understanding oncogene-induced senescence (OIS). This technique helps identify dysfunctional telomeres and monitor OIS in cell cultures and senescent cells.
Area of Science:
- Cytogenetics
- Molecular Biology
- Cellular senescence
Background:
- Telomere erosion and DNA damage are hallmarks of oncogene-induced senescence (OIS).
- Detecting DNA lesions that activate DNA repair is key to understanding OIS.
- Existing methods may not fully capture telomere dysfunction in senescence.
Purpose of the Study:
- To describe a novel protocol combining immuno-FISH to detect dysfunctional telomeres.
- To provide a method for monitoring OIS in experimental settings.
- To explore applications for analyzing ex vivo senescent cells.
Main Methods:
- Utilizing fluorescence in situ hybridization (FISH) for DNA integrity assessment.
- Employing immunofluorescence with γH2AX (DNA double-strand break marker) and PML (senescence marker).
- Combining immuno-FISH to identify DNA lesions at telomeres.
Main Results:
- The combined immuno-FISH protocol effectively detects dysfunctional telomeres.
- The method identifies DNA damage associated with OIS activation.
- The protocol is suitable for cell cultures and potentially ex vivo senescent cells.
Conclusions:
- The developed immuno-FISH protocol is a reliable tool for studying telomere dysfunction in OIS.
- This technique facilitates monitoring of OIS under experimental conditions.
- The method has potential applications in analyzing senescent cells isolated from tissues.
