MODEL SYSTEMS FOR THE STUDY OF THE ROLE OF PADPRP IN ESSENTIAL BIOLOGICAL PROCESSES

Citation
Ds. Rosenthal et al., MODEL SYSTEMS FOR THE STUDY OF THE ROLE OF PADPRP IN ESSENTIAL BIOLOGICAL PROCESSES, Biochimie, 77(6), 1995, pp. 439-443
Citations number
28
Categorie Soggetti
Biology
Journal title
ISSN journal
03009084
Volume
77
Issue
6
Year of publication
1995
Pages
439 - 443
Database
ISI
SICI code
0300-9084(1995)77:6<439:MSFTSO>2.0.ZU;2-T
Abstract
The nuclear enzyme poly(ADP-ribose) polymerase (PADPRP) is implicated in a number of cellular processes, including DNA repair, replication, and differentiation. We have been using several model systems to exami ne these potential roles of PADPRP. A human keratinocyte model system has been developed in which stable lines of epidermal cells contain an inducible construct harboring the antisense cDNA to PADPRP. When PADP RP antisense RNA is induced in culture, endogenous PADPRP mRNA, protei n, and enzymatic activity is lowered, and the pattern of poly(ADP) rib osylation in response to alkylating agents is altered. When keratinocy te clones containing the antisense construct or empty vector alone wer e grafted onto nude mice, they formed histologically normal human skin . The PADPRP antisense construct was also inducible in vivo by the top ical application of dexamethasone to the reconstituted epidermis, as d etermined by in situ hybridization. In addition, poly(ADP-ribose) poly mer could be induced and detected in vivo following the topical applic ation of a DNA alkylating agent to the grafted transfected skin layers . Thus, a model system has been developed in which the levels of PADPR P can be selectively manipulated in human keratinocytes in cell cultur e, and potentially in reconstituted epidermis as these keratinocyte li nes can be grafted to nude mice, whereupon they form a histologically and immunocytochemically normal human epidermis. Another system that w e have been utilizing to examine the role of PADPRP in proliferation a nd differentiation is stable lines of mouse preadipocytes that contain an inducible antisense PADPRP RNA. Similar to the keratinocyte system , these cells can inducibly express antisense PADPRP RNA, and subseque ntly lower levels of endogenous PADPRP. In this system, the normal dif ferentiation to adipocytes is blocked by the lowering of endogenous PA DPRP, apparently resulting from inhibition of replication immediately preceding terminal differentiation. This inhibition in turn may stem f rom the requirement for the physical association between PADPRP and po lymerase alpha during the S phase of the cell cycle. These systems wil l be useful tools to study the role of PADPRP in essential biological processes.