Degradation of human exonuclease 1b upon DNA synthesis inhibition.
In response to DNA damage, signaling pathways are triggered that either block the cell division cycle at defined transitions (G1-S and G2-M) or slow down progression through the S phase. Nucleases play important roles in DNA synthesis, recombination, repair, and apoptosis. In this study, we have examined the regulation of human exonuclease 1 (hEXO1b). The endogenous hEXO1b protein was only detected upon enrichment by immunoprecipitation. We found that hEXO1b was constantly expressed throughout the cell cycle. However, treatment of cells with agents that cause arrest of DNA replication led to rapid degradation of hEXO1b. This effect was fully reversed upon removal of the block. Analysis of synchronized cells showed that degradation of hEXO1b during the S phase was strictly dependent on DNA synthesis inhibition. DNA damage caused by UV-C radiation, ionizing radiation, cisplatin, or the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine did not affect hEXO1b stability. We show that hEXO1b was phosphorylated in response to inhibition of DNA synthesis and that phosphorylation coincided with rapid protein degradation through ubiquitin-proteasome pathways. Our data support the evidence that control of exonuclease 1 activity may be critical for the maintenance of stalled replication forks.
El-Shemerly
Mahmoud
M
Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland.
Janscak
Pavel
P
Hess
Daniel
D
Jiricny
Josef
J
Ferrari
Stefano
S
D000906:Antibodies / Q000737:chemistry / Q000276:immunology; D002460:Cell Line; D004249:DNA Damage; D045643:DNA Repair Enzymes / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects / Q000502:physiology*; D004273:DNA, Neoplasm / Q000037:antagonists & inhibitors* / Q000096:biosynthesis*; D004795:Enzyme Stability; D005090:Exodeoxyribonucleases / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D016196:S Phase / Q000502:physiology; D025801:Ubiquitin / Q000378:metabolism
D000906:Antibodies; D004273:DNA, Neoplasm; D025801:Ubiquitin; C114331:EXO1 protein, human; D005090:Exodeoxyribonucleases; D045643:DNA Repair Enzymes; D006918:Hydroxyurea