Characterization of the constriction with neocentric activity of 5RL chromosome in wheat

In wheat-5RL monotelosomic and ditelosomic addition lines, a proximal constriction located on the long arm of rye chromosome 5R shows neocentric activity at metaphase I of meiosis. In some pollen mother cells this region is unusually stretched, acquires kinetic activity and co-orients with the true centromeres. In the work described here we characterized the putative neocentric constriction of 5RL using various approaches. Fluorescence in situ hybridization (FISH) revealed that the rye subtelomeric repetitive DNA sequence pSc119.2 is a constituent of the 5RL constriction. This FISH site corresponds with a heterochromatic C-band in normal rye. Other subtelomeric (pSc34, pSc74, pSc200), centromeric (CCS1, Bilby) and Arabidopsis-type telomeric sequences produce no detectable hybridization signal on the constriction. Immunolocalization with anti-alpha-tubulin antibodies showed that microtubules are bound to the constriction in a similar way to their binding to true centromeres. Silver staining demonstrated that proteins are accumulated at the constriction, the signal being more prominent than that observed at the centromere and telomeres of 5RL. The frequency of neocentric activity in different plants varied dramatically in different generations and in siblings grown in different years, suggesting that activation of the neocentric site is dependent on internal features and environmental conditions.