Cation-chromatin binding as shown by ion microscopy is essential for the structural integrity of chromosomes

Abstract

Mammalian interphase and mitotic cells were analyzed for their cation composition using a three-dimensional high resolution scanning ion microprobe. This instrument maps the distribution of bound and unbound cations by secondary ion mass spectrometry (SIMS). SIMS analysis of cryofractured interphase and mitotic cells revealed a cell cycle dynamics of Ca 2+, Mg 2+, Na +, and K +. Direct analytical images showed that all four, but no other cations, were detected on mitotic chromosomes. SIMS measurements of the total cation content for diploid chromosomes imply that one Ca 2+ binds to every 12.5-20 nucleotides and one Mg 2+ to every 20-30 nucleotides. Only Ca 2+ was enriched at the chromosomal DNA axis and colocalized with topoisomerase IIα (Topo II) and scaffold protein II (ScII). Cells depleted of Ca 2+ and Mg 2+ showed partially decondensed chromosomes and a loss of Topo II and ScII, but not hCAP-C and histones. The Ca 2+ induced inhibition of Topo 11 catalytic activity and direct binding of Ca 2+ to Topo II by a fluorescent filter-binding assay supports a regulatory role of Ca 2+ during mitosis in promoting solely the structural function of Topo II. Our study directly implicates Ca 2+, Mg 2+, Na +, and K + in higher order chromosome structure through electrostatic neutralization and a functional interaction with nonhistone proteins.