Open AccessCharacteristics of UV-induced repair patches relative to the nuclear skeleton in human fibroblasts
Author(s) -
Parimal Karmakar
Publication year - 2000
Publication title -
mutagenesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.723
H-Index - 91
eISSN - 1464-3804
pISSN - 0267-8357
DOI - 10.1093/mutage/15.2.115
Subject(s) - xeroderma pigmentosum , cockayne syndrome , nucleotide excision repair , dna repair , biology , microbiology and biotechnology , t cell receptor , chemistry , biochemistry , genetics , dna , t cell , immune system
We have tried to characterize the nucleotide excision repair (NER) events associated with the nuclear skeleton in both repair-proficient and repair-deficient human cell lines following UV irradiation. The repair patches were labelled with biotin-16-dUTP and the repair sites were visualized by fluorescence microscopy using fluorescence-conjugated antibodies to biotin. The intensities of repair labelling measured for the three human cell lines of normal, xeroderma pigmentosum group C (XP-C) and Cockayne syndrome group B (CS-B) are in good agreement with their known repair capabilities. Digestion of nuclei with DNase I markedly solubilized the repair patches in normal (3-fold reduction after 1 h post-UV incubation) and transcription-coupled repair (TCR)-defective Cockayne syndrome cells (6-fold reduction after 1 h post-UV incubation). The intensity of repair labelling remained the same in TCR-proficient XP-C cells after DNase I digestion, indicating that the repair events mediated by the TCR pathway are tightly associated with the nuclear skeleton. Treatment with ammonium sulphate after DNase I digestion further reduced the intensity of repair patches in both normal and Cockayne syndrome cells, but not in XP-C cells. The tight association of repair patches generated by the TCR pathway with the nucleoskeleton in XP-C cells reinforces the concept of functional compartmentalization of the nucleus, where NER is highly heterogeneous.