Poly(ADP-ribose) degradation by post-nuclear extracts from human cells.
The nuclear metabolism of poly(ADP-ribose) is mainly regulated by poly(ADP-ribose) polymerase-1 (PARP-1) and by poly(ADP-ribose) glycohydrolase (PARG). A PARP-like enzyme, V-PARP, and a PARG isoform are present in the extra-nuclear compartment of mammalian cells, even if poly(ADP-ribose) has never been detected therein. In this work, we demonstrate the ability of post-nuclear extracts from HeLa and HL60 cells to degrade synthetic 32P-polymers of ADP-ribose to ADP-ribose and, further, to AMP. This reaction implies the combined action of PARG and of an ADP-ribose-degrading activity, possibly corresponding to a phosphodiesterase and/or to an ADP-ribose pyrophosphatase. The inhibition of PARG or ADP-ribose-degrading enzymes allowed the demonstration that in vitro synthesized 32P-poly(ADP-ribose) is first digested to ADP-ribose monomers by a typical PARG reaction, and that ADP-ribose is further rapidly converted into AMP by an Mg(2+)-dependent activity. Collectively, our results demonstrate the ability of the human cell post-nuclear fraction to convert synthetic poly(ADP-ribose) into utilizable AMP units by the concerted action of PARG and ADP-ribose-degrading activities.
Rossi
Laura
L
Istituto di Genetica Molecolare CNR, Pavia, Italy.
Denegri
Marco
M
Torti
Mauro
M
Poirier
Guy G
GG
Ivana Scovassi
A
A
D002467:Cell Nucleus / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000032:analysis; D004492:Edetic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism; D018922:HL-60 Cells; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009687:Nuclear Proteins / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000302:isolation & purification / Q000378:metabolism; D011064:Poly Adenosine Diphosphate Ribose / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011506:Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D013997:Time Factors
D009687:Nuclear Proteins; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011506:Proteins; C024857:poly(ADP)-ribosylated proteins; D011064:Poly Adenosine Diphosphate Ribose; D004247:DNA; D004492:Edetic Acid; D011065:Poly(ADP-ribose) Polymerases; D010727:Phosphoric Diester Hydrolases; D006026:Glycoside Hydrolases; C022407:poly ADP-ribose glycohydrolase