MOLECULAR ANALYSIS OF HISTONE GENE CLUSTER OF PSAMMECHINUS MILIARIS .1. FRACTIONATION AND IDENTIFICATION OF 5 INDIVIDUAL HISTONE MESSENGER-RNAS

The electrophoretic separation of labeled 9S histone mRNAs obtained from cleaving sea urchin polysomes was at first highly unreproducible. The secondary structure of the individual mRNAs had a greater effect on their relative electrophoretic mobilities than did their molecular weight differentials. The parameters affecting electrophoretic mobility by the novel method of running the labeled polysomal RNA in slab gels across polyacrylamide and urea gradients were determined. The initially complex and species-specific electrophoretic pattern could then, by a judicious choice of denaturing conditions, be simplified to yield 5 well defined classes of labeled mRNAs. Using optimal conditions for the separation of the RNA components, 5 messengers were isolated from Psammechinus embryos by preparative disc electrophoresis, 4 of which, after 2 electrophoretic separations, exhibited a unimodal distribution. Each of the mRNAs was translated in vitro, 4 of the 5 fractions promoting the synthesis of one major protein. The in vitro products were characterized by comparison of their electrophoretic mobilities with those of known sea urchin histones. It was thus possible to correlate individual mRNAs with specific histones. We propose that the 5 mRNAs designated a-e in order of decreasing electrophoretic mobility code for the histones H4, H2A, H2B, H3 and H1.