TY - CHAP
T1 - Analysis of nuclear uracil-DNA glycosylase (nUDG) turnover during the cell cycle
AU - Fischer, Jennifer A.
AU - Caradonna, Salvatore
PY - 2011/8/4
Y1 - 2011/8/4
N2 - Uracil-DNA glycosylases (UDG/UNG) are enzymes that remove uracil from DNA and initiate base-excision repair. These enzymes play a key role in maintaining genomic integrity by reducing the mutagenic events caused by G:C to A:T transition mutations. The recent finding that a family of RNA editing enzymes (AID/APOBECs) can deaminate cytosine in DNA has raised the interest in these base-excision repair enzymes. The methodology presented here focuses on determining the regulation of the nuclear isoform of uracil-DNA glycosylase (nUDG), a 36,000 Da protein. In synchronized HeLa cells, nUDG protein levels decrease to barely detectable levels during the S phase of the cell cycle. Immunoblot analysis of immunoprecipitated or affinity-isolated nUDG reveals ubiquitin-conjugated nUDG when proteolysis is inhibited by agents that block proteasomal-dependent protein degradation.
AB - Uracil-DNA glycosylases (UDG/UNG) are enzymes that remove uracil from DNA and initiate base-excision repair. These enzymes play a key role in maintaining genomic integrity by reducing the mutagenic events caused by G:C to A:T transition mutations. The recent finding that a family of RNA editing enzymes (AID/APOBECs) can deaminate cytosine in DNA has raised the interest in these base-excision repair enzymes. The methodology presented here focuses on determining the regulation of the nuclear isoform of uracil-DNA glycosylase (nUDG), a 36,000 Da protein. In synchronized HeLa cells, nUDG protein levels decrease to barely detectable levels during the S phase of the cell cycle. Immunoblot analysis of immunoprecipitated or affinity-isolated nUDG reveals ubiquitin-conjugated nUDG when proteolysis is inhibited by agents that block proteasomal-dependent protein degradation.
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U2 - 10.1007/978-1-61779-182-6_9
DO - 10.1007/978-1-61779-182-6_9
M3 - Chapter
C2 - 21755446
AN - SCOPUS:79960946697
SN - 9781617791819
T3 - Methods in Molecular Biology
SP - 137
EP - 149
BT - Cell Cycle Synchronization
A2 - Banfalvi, Gaspar
ER -