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 transcriptional competence and protein patterns, the latter indicating according to results previously obtained, that the testis-specific H1t is preferentially associated to the soluble fraction, whereas the other H1 variants are localized in the pellet. S and P chromatins also differed in the distribution of the poly(ADP-ribosyl)ating system, (poly(ADP-ribose)polymerase, reaction product and acceptor proteins), detected by incubating nuclei with 32P-NAD. The 32P-modified H1s and core histones of both fractions, known as specific ADPribose target proteins, were separated by high performance liquid chromatography and it was demonstrated that the H1 variants from S and P are differently ADPribosylated, being H1t always the best acceptor, and that most of the ADPribosylated variants were solubilized after DNase I treatment. The further digestion of P chromatin with the nuclease produced a fraction (pP) devoid of most DNA, but particularly enriched in transcriptionally competent tracts. The low DNA content of pP chromatin, which reflects the typical feature of a nuclear matrix, corresponded to a relevant poly(ADPribosyl)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 chromatin.
Dipartimento di Chimica Organica e Biologica, Facoltà di Scienze, Università Federico II, Napoli, Italy.
F
F
De Lucia
M R
MR
Mennella
P
P
Quesada
B
B
Farina
D000818:Animals; D001345:Autoradiography; D002843:Chromatin / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000235:genetics; D008297:Male; D011065:Poly(ADP-ribose) Polymerases / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D051381:Rats; D013737:Testis / Q000378:metabolism*; D014158:Transcription, Genetic
D002843:Chromatin; D004247:DNA; D011065:Poly(ADP-ribose) Polymerases