| In Dictyostelium discoideum most of the rRNA genes are known to be present in linear, extrachromosomal palindromic molecules. Here I report the identification, molecular cloning and partial DNA sequence analysis of the terminal Eco RI restriction fragment (Eco RI 1) of the rDNA. I identify a length heterogeneity confined to the most distal region of uncloned Eco RI 1, and demonstrate, by DNA sequence analysis of five independently isolated recombinant plasmids containing the Eco RI 1 fragment, that the heterogeneity can be accounted for by variable tracts of an unprecedented irregular satellite sequence with the general formula (C(,n)T)(,m); n varies from 1 to 8 and accounts for the irregularity, while m varies from 18 to 34 and accounts for the length heterogeneity. The proximal C(,n)T pattern is identical in all five clones of Eco RI 1, while the distal C(,n)T sequence appears to vary between clones. Immediately proximal to the C(,n)T satellite are found four nearly perfect tandem repeats of a 29 bp sequence. I discuss the evolution of these simple sequences at the rDNA termini and speculate about possible functions for the C(,n)T satellite. Using the cloned terminal restriction fragment as probe, I provide evidence that the C(,n)T satellite sequence does not occur at multiple sites elsewhere in the genome and that virtually all the rDNA in Dictyostelium exists in the linear palindromic form.;I also present tangential findings obtained in the course of the hybridization experiments. Certain sites of hybridization apparently resulted from aberrant restriction digestion, while others were shown to represent pBR322-like sequences present in at least one genomic DNA preparation. Finally, I identified in undigested DNA discrete clusters of bands able to hybridize with rDNA terminal sequences.;In the ciliate Tetrahymena, a single chromosomal copy of the rDNA appears to generate the extrachromosomal rDNA through a gene amplification process. To investigate the possible existence in Dictyostelium of chromosomal rDNA, I cloned sequences from both the rDNA center and termini and used these to probe restriction digests of nuclear DNA. Each probe was found to hybridize to one or a few sites in the genome, and hybridization intensities were consistent with those sites corresponding to single copy chromosomal sequences. Inconsistencies between the numbers of sites which hybridized with the two probes suggests an intriguing model for Dictyostelium rDNA integration. |
