The peculiar features of human telomeric nucleosomes: sequence-dependent intrinsic mobility and specific interactions with telomeric proteins

The structural organization of human telomeres has not yet completely characterized, mainly because of the complexity of telomere structure and functions. To date, only three proteins have been identified that bind to telomeric sequences, TRF1 and TRF2 that bind to TTAGGG duplex telomeric repeats and POT1 which recognizes 3’ single stranded overhangs. Several other proteins are involved in the formation of the telomeric complex, binding directly or indirectly to these three proteins. Besides TRF1 and TRF2 there is another protein complex that binds to duplex telomeric DNA, the histone octamer. Most of vertebrate telomeres are organized in arrays of tightly spaced nucleosomes, having a shorter repeat size than bulk nucleosomes. The role of nucleosomes in the establishment of telomeric structures is still largely unknown. We set up model systems to study sequence-dependent features of telomeric nucleosomes, and to analyze the relationship between telomeric proteins and nucleosomes. We find that telomeric nucleosomes are the least stable nucleosomes so far studied. By means of a restriction enzyme assay, and by AFM visualization, we find that telomeric nucleosomes are intrinsically mobile, i.e. they are able to move along DNA without the action of ATP-dependent remodeling enzymes. By electrophoretic mobility shift assay, we demonstrate that TRF1, and to a lesser extent TRF2, are able to recognize their binding sites in a nucleosomal context. DNase I footprinting of the complex TRF1-nucleosome shows that the formation of a ternary complex is strongly dependent on the orientation of binding sites on the nucleosome surface more than on the distance from the nucleosome dyad axis. Moreover, TRF1 binding causes alterations in nucleosome structure without dissociation of histone subunits. These results indicate that nucleosomes contribute to the establishment of a telomeric capping structure, and suggest that they may play a role in telomere dynamics.