Methylation of ribosomal proteins in HeLa cells

Abstract

The in vivo post-translational methylation of individual ribosomal proteins has been studied. Methylation was detected by: (1) 3H labeling of cells incubated with a mixture of [ Me- 3H]methionine plus [ 35S]methionine, the incorporation of the 3H-labeled amino acid methionine being corrected by the 35S uptake, (2) 3H incorporation from [ Me- 3H]methionine during inhibition of protein synthesis with cycloheximide, and (3) amino acid analysis. A minimum of seven methylated ribosomal proteins were detected. The levels of methylation differed among several proteins (and possibly between two sites of the same protein) in relationship to suppression of protein synthesis or ribosome formation. The methylation of some ribosomal proteins (most of Nos. 30, 62, 63 and part of 18 and 42) occurred on ribosomes that were being processed, i.e., it was inhibited when ribosome formation was suppressed with actinomycin D. The methylation of other ribosomal proteins occurred in mature ribosomes (protein No. 38 and apparently another site of 42), as it was not suppressed by actinomycin D. In another experiment cells were both pulsed with [ Me- 3H]methionine and chased with an excess of non-radioactive methionine during suppression of protein synthesis (in the presence of cycloheximide); when protein synthesis was allowed to recover protein 18 was preferentially labeled (methylated), in the continued presence of an excess of non-radioactive methionine. The level of methylation of protein 30 as well as one site of protein 42, both apparently methylated during ribosome processing, seemed to increase in the absence of protein synthesis. The methylation of protein 42 in mature ribosomes did not appear to be affected by suppression of protein synthesis. In general, the ribosomal proteins which appeared to be methylated in nascent ribosomes contained methyllysine, while methylarginine was found in those methylated in mature ribosomes.