An examination of chromatin states in Oxytricha trifallax nanochromosomes by psoralen crosslinking

Abstract

The macronuclear genome of the ciliate Oxytricha trifallax consists of thousands of small nanochromosomes, which are each amplified to an average of 1900 copies per cell. The copy number of individual nanochromosomes is likely not stable within asexually dividing cultures and is also not well correlated with cellular levels of RNA transcripts. These observations suggest that Oxytricha may possibly regulate transcriptional activity through novel mechanisms unique to ciliates.An assay combining psoralen crosslinking with Southern hybridization has previously determined that, in a number of eukaryotes, the highly amplified ribosomal DNA copies exist in alternative chromatin states. Psoralen distinguishes between gene copies with different chromatin states based on different nucleosomal occupancies. We have optimized a protocol for psoralen crosslinking of Oxytricha macronuclei. Using this protocol, we have determined that alternative chromatin states likely exist in Oxytricha rDNA, but not in any other nanochromosomes we tested. Additional studies will be necessary to elucidate potential mechanisms of transcriptional regulation in Oxytricha.