DNA and Chromatin Structure of Telomere-Subtelomere Junctions inSilene latifolia
Genomic variants of telomere-subtelomere junctions in Silene latifolia chromosomes were cloned and sequenced. In addition to the previously isolated subtelomeric sequence X43.1 (Buzek et al., 1997), several novel telomere-associated sequences have been characterised, including high-copy tandem repeat 15Ssp and low-copy sequence BstB19. Their subtelomeric location has been confirmed by pulsed-field gel electrophoresis and fluorescent in situ hybridisation. Analysis of chromatin structures (nucleosome phasing, nucleosome positioning) at these sequences revealed their specific features: 1) In case of X43.1, two distinct nucleosome periodicities have been observed and preferred translational positions of its nucleosomes have been mapped by both low and high resolution techniques. These positions correspond to computer-predicted nucleosome positions. Sites of X43.1 attachment to telomere are nonrandom; all in all, just three variants have been found, all of them corresponding either to a start or an end of a nucleosome-wrapped DNA. 2) A newly described tandemly repeated 159 bp telomere associated sequence, 15Ssp, shows a presence of MNase hypersensitive sites when either naked DNA or chromatin is digested. An alternative use of chemical nuclease MPE-Fe(II) digestion results in regular nucleosome ladder. Positions of preferential MNase cleavage sites correspond to phased oligo-A/T tracts causing local deformation of DNA axis. Further, the 15SspI nucleosomes show extremely short spacing (157*6 bp), instability of mononucleosomes in both MNase and MPE-Fe(II) digestion and absence of translation positioning ? the features typical for telomeric chromatin. In our results we suggest that two subsets of telomere-subtelomere junctions may be distinguished: the first, represented by X43.1 in S. latifolia, or by HRS60 (Fajkus et al., 1995) or TAS4.9 (Horáková and Fajkus, 2000) in tobacco, is characterised by change in telomere-specific chromatin structure into subtelomeric heterochromatin immediately in telomere-subtelomere boundary. In the second subset, first reported here in case of 15Ssp in S. latifolia, telomere-specific chromatin pervades to the telomere-associated region and the transition to heterochromatin occurs in centromere-proximal part of subtelomere.
This work was supported by the Czech Ministry of Education, project VS97032.
E. Sykorova1,2, M. Horakova1, I. Mikako3, K. Fukui3, and J. Fajkus1,2
1Institute of Biophysics, Czech Academy of Sciences,