Related Experiment Videos
A damage-recognition protein which binds to DNA containing interstrand cross-links is absent or defective in Fanconi
B Hang1, A T Yeung, M W Lambert
1Department of Laboratory Medicine and Pathology, UMDNJ, New Jersey Medical School, Newark 07103.
Abstract:
A DNA binding protein with specificity for DNA containing interstrand cross-links induced by 4,5',8-trimethylpsoralen (TMP) plus long wavelength ultraviolet (UVA) light has been identified in normal human chromatin. Protein binding to DNA was determined using a gel mobility shift assay and an oligonucleotide containing a hot spot for formation of psoralen interstrand cross-links. Specificity of the damage-recognition protein for cross-links was demonstrated both by a positive correlation between level of cross-link formation in DNA and extent of protein binding and by effective competition by treated but not undamaged DNA for the binding protein. Chromatin protein extracts from cells from individuals with the genetic disorder, Fanconi anemia, complementation group A (FA-A), which have decreased ability to repair damage produced by TMP plus UVA light, failed to show any protein binding to TMP plus UVA treated DNA. We have previously shown that these chromatin protein extracts contain a DNA endonuclease complex, pI 4.6, which specifically recognizes and incises DNA containing interstrand cross-links and which in FA-A cells is defective in its ability to incise this damaged DNA (Lambert et al. (1992) Mutation Res., 273, 57-71). Together, these findings suggest that the DNA binding protein identified is involved in recognition and repair of DNA interstrand cross-links.
Insights
Researchers identified a DNA binding protein that recognizes DNA interstrand cross-links caused by 4,5
Area of Science:
- Molecular Biology
- DNA Repair Mechanisms
- Genetics
Background:
- DNA interstrand cross-links (ICLs) are severe DNA lesions.
- 4,5',8-trimethylpsoralen (TMP) plus UVA light induces ICLs.
- Fanconi anemia (FA) is a genetic disorder characterized by impaired ICL repair.
Purpose of the Study:
- To identify and characterize DNA binding proteins involved in ICL recognition.
- To investigate the role of these proteins in DNA repair pathways.
Main Methods:
- Gel mobility shift assay using oligonucleotides with TMP-induced ICLs.
- Analysis of chromatin protein extracts from normal and FA-A cells.
Main Results:
- A novel DNA binding protein specific for TMP-induced ICLs was identified in normal human chromatin.
- Protein binding correlated with the level of DNA cross-linking.
- FA-A cell extracts, deficient in ICL repair, lacked this DNA binding activity.
Conclusions:
- The identified DNA binding protein is likely involved in the recognition of DNA interstrand cross-links.
- This protein may play a crucial role in the DNA repair pathway for ICLs.
- Defects in this recognition protein could contribute to the FA-A phenotype.