Studies on the methylation of cytoplasmic ribosomal RNA from cultured higher plant cells.

The methylation of cytoplasmic ribosomal RNA of cultured sycamore cells (Acer pseudoplatanus L.) was investigated. Labelled 17-S and 26-S rRNA were prepared from cells that had been incubated with either [32P]phosphate, [Me-3H]methionine or [Me-14C]methionine. Ion-exchange resin chromatography of 0.3 M KOH or 1 M HCl hydrolysates and two-dimensional chromatographic analyses of phosphodiesterase plus phosphatase digests of 17-S and 26-S rRNA were performed. 17-S and 26-S rRNA contain 49 and 91 methyl groups per molecule, respectively. These values were verified in sevemral ways. The high degree of methylation of sycamore rRNA, particularly for the 26-S rRNA, contrasts with the situation in all other investigated organisms. Several methylated bases were identified. 7-Methylguanine and 5-methylcytosine both occur in 17-S and 26-S rRNA. N6-Methyladenine and N6,N6-dimethyladenine are restricted to the 17-S rRNA while 3-methyluracil and 1-methyladenine occur in the 26-S rRNA. One hypermodified uridine was also tentatively identified in the small rRNA. In 17-S rRNA, there is one copy of 7-methylguanine, N6-methyladenine and hypermodified uridine and two copies of N6,N6-dimethyladenine. 3-Methyluracil, 1-methyladenine and 5-methylcytosine occur twice, twice and three times, respectively, in 26-S rRNA. 7-Methylguanine and 5-methylcytosine are only in submolar amounts in the 26-S and 17-S rRNA, respectively. There are 40 +/- 2 and 83 +/- 3 2'-O-methylriboses per 17-S and 26-S rRNA molecule, respectively. In addition to the four 2'-O-methylnucleosides, one 2'-O-methylpseudouridine is present in the 17-S rRNA. Several lines of evidence argues for a non-random distribution of the methylriboses. In particular, one and seven Nm-Nm-Np structures occur in the 17-S and 26-S rRNA, respectively. The data are discussed comparatively with the methylation pattern of Escherichia coli, yeast and HeLa cell rRNA.