Ultraviolet damage and nucleosome folding of the 5S ribosomal RNA gene

The Xenopus borealis somatic 5S ribosomal RNA gene was used as a model syst em to determine the mutual effects of nucleosome folding and formation of u ltraviolet (UV) photoproducts (primarily cis-syn cyclobutane pyrimidine dim ers, or CPDs) in chromatin. We analyzed the preferred rotational and transl ational settings of 5S rDNA on the histone octamer surface after induction of up to 0.8 CPD/nucleosome core (2.5 kJ/m(2) UV dose). DNase I and hydroxy l radical footprints indicate that UV damage at these levels does not affec t the average rotational setting of the 5S rDNA molecules. Moreover, a comb ination of nuclease trimming and restriction enzyme digestion indicates the preferred translational positions of the histone octamer are not affected by this level of UV damage, We also did not observe differences in the UV d amage patterns of irradiated 5S rDNA before or after nucleosome formation, indicating there is little difference in the inhibition of nucleosome foldi ng by specific CPD sites in the 5S rRNA gene. Conversely, nucleosome foldin g significantly restricts CPD formation at all sites in the three helical t urns of the nontranscribed strand located in the dyad axis region of the nu cleosome, where DNA is bound exclusively by the histone H3-H4 tetramer. Fin ally, modulation of the CPD distribution in a 14 nt long pyrimidine tract c orrelates with its rotational setting on the histone surface, when the stro ng sequence bias for CPD formation in this tract is minimized by normalizat ion. These results help establish the mutual roles of histone binding and U V photoproducts on their formation in chromatin.