Inhibition of cell proliferation in Saccharomyces cerevisiae by expression of human NAD+ ADP-ribosyltransferase requires the DNA binding domain ("zinc fingers").
Constitutive expression of human nuclear NAD+: protein ADP-ribosyltransferase (polymerizing) [pADPRT; poly(ADP-ribose)polymerase; EC 2.4.2.30] as an active enzyme in Saccharomyces cerevisiae, under the control of the alcohol dehydrogenase promoter, was only possible with simultaneous inhibition of ADP-ribosylation by 3-methoxybenzamide. Induction of fully active pADPRT from the inducible galactose epimerase promoter resulted in inhibition of cell division and morphological changes reminiscent of cell cycle mutants. Expression of a pADPRT cDNA truncated at its 5' end had no influence on cell proliferation at all. Obviously the amino-terminal part of the DNA binding domain containing the first "zinc finger", which is essential for inducibility of pADPRT activity by DNA breaks, is also required for inhibition of cell growth on expression in yeast. Full-length as well as truncated pADPRT molecules were directed to the cell nucleus where the fully active enzyme produced large amounts of poly(ADP-ribose) by automodification. Since pADPRT turned out to be the only target for ADP-ribosylation in these cells, elevated levels of poly(ADP-ribose) were the most likely cause of inhibition of cell division, presumably resulting from interaction with chromosomal proteins.
Kaiser
P
P
Institut für Biochemie, Universität Innsbruck, Austria.
Auer
B
B
Schweiger
M
M
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D015153:Blotting, Western; D002455:Cell Division; D004247:DNA / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D006801:Humans; D008856:Microscopy, Fluorescence; D010957:Plasmids; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D014170:Transformation, Genetic
D004274:DNA, Recombinant; D000246:Adenosine Diphosphate Ribose; D004247:DNA; D011065:Poly(ADP-ribose) Polymerases