Unusual pattern of ribonucleic acid components in the ribosome of Crithidia fasciculata, a trypanosomatid protozoan.

Abstract

In a previous study from this laboratory, presumptive ribosomal ribonucleic acid (RNA) species were identified in the total cellular RNA directly extracted from intact cells of the trypanosomatid protozoan Crithidia fasciculata (M. W. Gray, Can. J. Biochem. 57:914-926, 1979). The results suggested that the C. fasciculata ribosome might be unusual in containing three novel, low-molecularweight ribosomal RNA components, designated e, f, and g (apparent chain lengths 240, 195, and 135 nucleotides, respectively), in addition to analogs of eucaryotic 5S (species h) and 5.8S (species i) ribosomal RNAs. In the present study, all of the presumptive ribosomal RNAs were indeed found to be associated with purified C. fasciculata ribosomes, and their localization was investigated in subunits produced under different conditions of ribosome dissociation. When ribosomes were dissociated in a high-potium (880 mM K+, 12.5 mM Mg2") medium, species e to i were all found in the large ribosomal subunit, which also contained an additionaL transfer RNA-sized component (speciesj). However, when subunits were prepared in a low-magnesium (60 mM K+, 0.1 mM Mg2+) medium, two of the novel species (e and g) did not remain with the large subunit, but were released, apparently as free RNAs. Control experiments have eliminated the possibility that the small RNAs are generated by quantitative and highly specific (albeit artifactual) ribonuclease cleavage of large ribosomal RNAs during isolation. In terms of RNA composition and dissociation properties, therefore, the ribosome of C. fasciculata is the most "atypical" eucaryotic ribosome yet described. These observations raise interesting questions about the function and evolutionary origin of C. fasciculata ribosomes and about the organization and expression of ribosomal RNA genes in this organism.Most eucaryotic ribosomes conform to a pattern in which the large subunit contains three ribonucleic acid (RNA) components (25 to 28S, or "L-ribosomal RNA [rRNA]"; 5.8S, or "L3-rRNA"; and 5S), whereas the small subunit contains one (17 to 18S, or "S-rRNA") (3). I have recently shown (14) that the rRNA of Crithidia fasciculata, a trypanosomatid protozoan, differs from that of a typical eucaryote in several respects. (i) The L-rRNA (species a) is heat labile, dissociating into two high-molecular-weight components, species c and d (apparent molecular weights, 0.67 x 10' and 0.575 x 106, respectively). (ii) Heat denaturation of C. fasciculata rRNA also releases a small RNA (species i) analogous in this respect to eucaryotic 5.8SrRNA, but which migrates appreciably more slowly than wheat 5.8S rRNA in nondenaturing polyacrylamide gels. (iii) The S-rRNA of C. fasciculata also displays an anomalously slow migration during polyacrylamide gel electrophoresis, so that its apparent molecular weight (0.825 x 106) is substantially greater than that of the S-rRNA ofmost other eucaryotes (typically, this is -0.7 x 106). (iv) In addition to 5S rRNA (species h), three other small RNAs are found in nondenatured C. fascicu...

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