Identification of a Novel “Chromosome Scaffold” Protein That Associates with Tec Elements Undergoing En Masse Elimination inEuplotes crassus

Abstract

During macronuclear development in the ciliate Euplotes crassus, the highly repetitive, transposon-like Tec elements possess an unusual chromatin structure. We observed that the Tec element chromatin is highly resistant to salt extraction and behaves like a nuclear matrix/chromosome scaffold-associated structure. Standard matrix/scaffold extraction procedures identified two major proteins: 1) an ∼140-kDa protein that seems to be topoisomerase II based on its reactivity with anti-topoisomerase II antibodies, and 2) an 85-kDa protein that we further purified by acid extraction and have shown to be a novel protein by sequence analysis of its gene. The 85-kDa protein (p85) is a developmental stage-specific protein and is located exclusively in the developing macronucleus. Immunolocalization studies of p85 show that it colocalizes with topoisomerase II in chromatin. In addition, in situ hybridization combined with immunofluorescence localization of the proteins indicates that 100% of the Tec elements colocalize with 70% of the p85, whereas no significant colocalization with a total macronuclear sequence-specific probe is observed. p85 is the first developmental stage-specific protein identified as being specifically associated with sequences undergoing elimination in E. crassus.