Human NEIL1 localizes with the centrosomes and condensed chromosomes during mitosis.
The DNA glycosylase hNEIL1 initiates base excision repair (BER) of a number of oxidized purines and pyrimidines in cellular DNA and is one of three mammalian orthologs of the Escherichia coli Nei/Fpg enzymes. Human NEIL1 has been purified and extensively characterized biochemically, however, not much is known about its intracellular distribution. In the present work, we have studied the cellular localization of hNEIL1 using both antibodies raised against the full-length recombinant protein and a stable HeLa cell line expressing hNEIL1 fused N-terminal to EGFP. The results presented reveal an intricate mitotic distribution of hNEIL1. Centrosomal localization of hNEIL1 was observed when mitotic HeLa cells were immunostained with hNEIL1 antibodies. This localization was confirmed when Western blots of isolated centrosomes from stably expressing hNEIL1-EGFP HeLa cells were probed with GFP or hNEIL1 antibodies, even though a fluorescent signal could not be detected in the centrosomes of these cells. Human NEIL1 was also shown to be associated with mitotic condensed chromosomes. Notably, the interaction of hNEIL1 with condensed chromatin was disrupted when cells were fixed with chemical fixatives that are regularly used in immunodetection techniques.
Hildrestrand
Gunn A
GA
Centre for Molecular Biology and Neuroscience and Institute of Medical Microbiology, Rikshospitalet-Radiumhospitalet HF, 0027 Oslo, Norway.
Rolseth
Veslemøy
V
Bjørås
Magnar
M
Luna
Luisa
L
D002453:Cell Cycle; D018385:Centrosome / Q000378:metabolism*; D002877:Chromosomes, Human; D045647:DNA Glycosylases / Q000235:genetics / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis; D011994:Recombinant Proteins / Q000378:metabolism
D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D045647:DNA Glycosylases; C473634:NEIL1 protein, human