Chromatin structure of the 5S ribosomal RNA genes and satellites in Xenopus laevis

The chromatin structure of the 5S rRNA genes and two satellite DNAs in Xenopus laevis were investigated. The average nucleosome repeat lengths of these three repeated sequences were found to be the same as the bulk nucleosome repeat length of 192±4 bp in erythrocyte chromatin, although widths of the nucleosome bands were noticeably different for the three DNAs. The 5S DNA nucleosome bands were much broader than those from bulk chromatin while the 388 bp satellite bands were noticeably sharper. This may reflect the difference in the variations of nucleosome repeat lengths of the three sequences. The positions of micrococcal nuclease sensitive regions in the 5S DNA repeats of purified DNA and chromatin from erythrocytes were determined using an indirect end-labeling hybridization technique. While most of the sites found in purified DNA are present in chromatin, the patters of intensities are strikingly different in the two cases. In 5S chromatin, three nuclease-sensitive regions are spaced approximately a nucleosome length apart, suggesting that nucleosomes occupy a single regular arrangement on most of the 5S DNA repeats. Because the preferred locations appear to be reestablished in each repeating unit, despite spacer length heterogeneity, the regular chromatin structure may reflect the presence of a sequence-specific DNA-binding component on inactive 5S rRNA genes. The arrangements of nucleosomes on 5S DNA and the satellite DNAs were also investigated by studying the availability of restriction enzymes Hind III, Rsa I, Mbo I, and Hae III cleave a fraction of their sites in bulk chromatin producing DNA distributions which indicate that they probably cleave chromatin only in the linker DNA between nucleosome cores. The 5S DNA fragments produced from these digests, as analyzed by Southern blot hybridization, indicate that only a fraction of the restriction sites in the 5S DNA repeats are available for cleavage in chromatin. These results are analyzed with respect to the single nucleosome arrangement of 5S DNA repeats deduced from the micrococcal nuclease cleavage map. Preliminary analyses of the chromatin structure of the 741 bp satellite I do not indicate a phased nucleosome arrangement on this DNA sequence, while the 388 bp repeating sequence may be organized in a regular chromatin structure. The close relation between the 388 bp sequence repeat length and the nucleosome dimer length suggests that nucleosomes may be phased. Also, analysis of micrococcal nuclease cleavage of the 388 base pair satellite indicates that the positions of sensitive sties in erythrocyte chromatin and purified DNA are different.