Minisymposium 16: Gene Regulation

Abs # 28001: Epigenetic mechanisms and chromatin modifications controlling nucleolar dominance

Presenter:	Pikaard, Craig S., pikaard@biology.wustl.edu
Authors	Pikaard, Craig S. (A)   Lawrence, Richard  (A)   Lewis, Michelle  (A)   Pikaard, Diane J (A)   Preuss, Sasha  (A)   Brower-Toland, Brent  (A)
Affiliations:	(A): Biology Department, Washington University, St. Louis, MO USA
Web Site:	http://www.biology.wustl.edu/pikaard/

A common occurrence in inter-species hybrids and allopolyploids is for the chromosomes of only one parent to organize nucleoli, the sites of ribosome biogenesis. This epigenetic phenomenon, known as nucleolar dominance, was first discovered 70 years ago as a change in chromosome morphology at the loci where nucleoli form. The molecular explanation is the transcription of only one parental set of rRNA genes. In Arabidopsis suecica, the allotetraploid hybrid of A. thaliana and A. arenosa, and in diploid hybrids of A. thaliana x A. lyrata, the rRNA genes derived from A. thaliana are silenced. Chemicals that interfere with DNA methylation (i.e. 5 aza-2’ deoxycytosine) or histone deacetylation (e.g. trichostatin A), derepress the A. thaliana rRNA genes, inspiring a search for the responsible chromatin modifying activities. As part of an NSF funded consortium(see www.chromdb.org) we are using RNA interference (RNAi) to knock down chromatin modifying protein expression in Arabidopsis and then assaying a variety of epigenetic phenomena including nucleolar dominance. Our lab has developed a method for genetic transformation of A. suecica using the floral dip method commonly used for A. thaliana. In A. suecica, we have targeted by RNAi 13 genes representing the four families of histone deacetylases (HDACs). An HDAC within the family related to yeast Rpd3 is required for rRNA gene silencing in nucleolar dominance. In an effort to determine which histones and which amino acids the HDAC affecting nucleolar dominance might modify, we are turning to mass spectroscopy. Thus far, we have adapted methods for obtaining pure nucleosomal histones and have identified all the core histones, including some histone variants. We have also identified the amino-terminal lysines of histones H3 and H4 that are acetylated and are working to determine if any of these acetylated amino acids are deacetylated by the HDAC. In the course of making hybrids or allopolyploids containing chromatin RNAi transgenes we have found that knocking down a variety of chromatin modifying activities, including HDACs, has little phenotypic consequence in A. thaliana but causes dramatic developmental abnormalities in hybrids. These results suggest that chromatin modifications are important for something we might call “epigenetic homeostasis” - mechanisms that help suppress conflicting instructions encoded by two competing genomes in a hybrid nucleus.