Exposure of histone antigenic determinants in chromatin

Abstract

The exposure of antigenic determinants of histones present in "native" chromatin was studied by: (1) testing their ability to elicit anti-histone antibodies and (2) measuring their ability to interact with anti-histone sera. To this end, antisera specific to purified histone fractions and to purified rat liver chromatin were elicited in rabbits. The anti-chromatin sera did not react with pure histone fractions and pure histone fractions F2b, F3, F2a1, and F2a2 failed to inhibit the complement fixation resulting from the binding of anti-chromatin to chromatin. These results suggest that in native chromatin, determinants in these histones are not immunogenic. Histone F1, however, inhibited the reaction between chromatin and anti-chromatin. Antisera elicited by histone fractions reacted weakly with "native" chromatin. The maximal complement fixations (obtained with 5-10 mug of chromatin DNA) were as follows: 60% with anti-F2b, 20% with anti-F1 and anti-F3, and less than 5% with either anti-F2a1 or anti-F2a2. Studies of the interaction between anti-histone antibodies and chromatin in which chromatin was used as an immunoadsorbent indicated that antibodies against different histones were adsorbed to a different degree by the same amount of chromatin. Differences in the immunoadsorbing capacity between sonicated and nonsonicated chromatin were found. Quantitative adsorbtion studies revealed that in the "native" chromatin structure, antigenic determinants of F1 and F2b were more available to interact with homologous antibody than those of F3 and F2a1 and that determinants in F2a2 were the least available. It could be calculated that the "equivalent antigenicity" of the histones in chromatin was 9.6% for F1, 3.2% for F2b, and 0.90% for F3 and F2a1. Upon sonication these values did not change for F1 but increased two-, three-, and fourfold for F2b, F3, and F2a1, respectively. Digestion of chromatin with trypsin totally abolished the ability of chromatin to adsorb anti-histone antibodies.