ON THE BIOLOGICAL ROLE OF THE NUCLEAR POLYMERIZING NAD(-PROTEIN(ADP-RIBOSYL) TRANSFERASE (ADPRT) - ADPRT FROM DICTYOSTELIUM-DISCOIDEUM AND INACTIVATION OF THE ADPRT GENE IN THE MOUSE())

Two approaches have been used to elucidate the role of the nuclear pol ymerizing NAD(+):protein(ADP-ribosyl)transferase (ADPRT): i) compariso n of the primary structure of Dictyostelium discoideum ADPRT derived f rom a 2 lib, partial cDNA sequence with the mammalian, fish, amphibian and insect counterparts revealed an overall homology of 25%. Whereas the automodification domain was not conserved at all, the NAD(+) bindi ng domain (aa 859-908) showed more than 70% identical amino acids in a ll species. Together with the similar enzymatic properties of the ADPR Ts the genetic conservation underlined the notion that ADPRT plays a m ajor role in various cellular processes; and ii) inactivation of the A DPRT gene in murine embryonic stem cells by homologous recombination l ed to mouse strains with a complete lack of nuclear poly(ADP-ribosyl)a tion. These ADPRT mutant mice were viable and fertile indicating that ADPRT is dispensable in mouse development. Moreover, repair of UV and MNNG induced DNA damage was not affected in ADPRT/3T3 Like fibroblasts , as measured by reactivation of in vitro damaged reporter plasmids an d unscheduled DNA synthesis. However, about 30% of the ADPRT mutant mi ce developed pathological skin aberrations on a mixed 129/Sv x C57B1/6 genetic background. These mice will be extremely useful to define the precise biological role of poly(ADP-ribosyl)ation.