Studies of human histone messenger RNA. II. The resolution of fractions containing individual human histone messenger RNA species

Abstract

Polyribosomal 4 to 18 S RNA from S phase HeLa S-3 cells has been fractionated by chromatography on oligo(dT)-cellulose and resolved into multiple discrete components by continuous elution preparation electrophoresis. The human histone messenger RNA (mRNA) species associated with various polyadenylated [poly(A(+))] and nonpolyadenylated [poly(A(-))] components of 4 to 18 S RNA were determined by translation of these RNA fractions in vitro using a Krebs II ascites cell-free system followed by resolution of histones synthesized in vitro on polyacrylamide gels containing Triton X-100. The results of these studies indicate that poly(A(-)) 4 to 18 S RNA from S phase HeLa polyribosomes contains: (a) large quantities of discrete 7.4 and 8 S RNA species which are not functional histone mRNA; (b) a discrete 8.6 S RNA fraction which contains the templates of human histone H4; (c) 9.2 to 10.7 S RNA which contains mixtures of incompletely resolved histone H2B, H2A, and H3 mRNA (These mRNA species do not closely correspond to discrete RNA subfractions resolvable by our techniques.); (d) discrete 12 and 13 S RNA fractions which contain templates of human histone H1 polypeptides. The present studies also indicate that the mRNA templates of histone variants H3.2 and H3.3 have a slightly lower electrophoretic mobility than H3.1 mRNA and that H2A.2 mRNA has a slightly lower electrophoretic mobility than H2A.1 mRNA. In addition, appreciable quantities of H3.2, H3.3, and H2A.2 mRNA are bound to oligo(dT)-cellulose in 0.5 M KCl. These results indicate that mRNA species of the same histone class differ slightly in primary structure and are consistent with the hypothesis that some histone mRNA species contain short tracts of poly(A).