POLY(ADPRIBOSYL)ATION SYSTEM IN TRANSCRIPTIONALLY ACTIVE-RAT TESTIS CHROMATIN FRACTIONS

The rat testis chromatin fractions (soluble, S, and insoluble, P) were prepared by mild digestion of nuclei with DNAase I. They appeared to be different in specific biochemical features such as their transcript ional competence and protein patterns, the latter indicating, accordin g to results previously obtained, that the testis-specific Hit is pref erentially associated to the soluble fraction, whereas the other H1 va riants are localized in the pellet. S and P chromatins also differed i n the distribution of the poly(ADP-ribosyl)ating system, (poly(ADP-rib ose)polymerase, reaction product and acceptor proteins), detected by i ncubating nuclei with P-32-NAD. The P-32-modified His and core histone s of both fractions, known as specific ADPribose target proteins, were separated by high performance liquid chromatography and it was demons trated that the H1 variants from S and P are differently ADPribosylate d, being Hit always the best acceptor, and that most of the ADPribosyl ated variants were solubilized after DNase I treatment. The further di gestion of P chromatin with the nuclease produced a fraction (pP) devo id of most DNA, but particularly enriched in transcriptionally compete nt tracts. The low DNA content of pP chromatin, which reflects the typ ical feature of a nuclear matrix, corresponded to a relevant poly(ADPr ibosyl)ation, the highest as compared to S and P fractions. Moreover, long and branched chains of poly(ADP-ribose) were found associated to pP sample which resemble the products determined in the soluble chroma tin. (C) 1996 Wiley-Liss, Inc.