In hybrids, genes inherited from both parents are typically expressed, producing intermediate phenotypes for characters such as flower or leaf morphology. This is true for Arabidopsis thaliana (A.t.), Arabidopsis arenosa (A.a) and their hybrid, A. suecica (A.s.), whose flowers are shown at the top left corner of this page. However, some genes are expressed from the chromosomes inherited from only one parent. An example is nucleolar dominance, an epigenetic phenomenon in hybrids which describes the formation of a nucleolus (or nucleoli) on the chromosomes inherited from only one of the progenitors, regardless of whether that progenitor was the maternal or the paternal parent. Nucleolar dominance occurs in both the plant and animal kingdoms and is due to the expression of only one parental set of rRNA genes.

As shown below, rRNA genes are inherited from both progenitors in Arabidopsis suecica and are readily detected by PCR (left panel) but transcriptional analyses show that only the A. arenosa-derived rRNA genes in A. suecica are actually expressed into RNA (right panel; lane 5).The A. thaliana rRNA genes are not expressed in the hybrid (lane 8).

In 1997 we found that nucleolar dominance in Brassica hybrids or A. suecica can be reversed by chemical inhibitors of DNA methylation or histone deacetylation (see Figure below), which led to the realization that nucleolar dominance is due to the selective silencing of one set of rRNA genes rather than the selective activation of the other. Because rRNA genes are clustered by the hundreds, spanning millions of basepairs of chromosomal DNA, nucleolar dominance is one of the most extensive gene silencing phenomena known.

Further study has shown that rRNA gene silencing involves concerted changes in DNA methylation and histone modification and we have proposed a model whereby DNA and histone modifications are each upstream of one another in a self-reinforcing, circular pathway (see Figure below). Changes in DNA methylation, histone acetylation and histone methylation are critical to the on-off switch mechanism that controls the number of active rRNA genes, both in hybrids displaying nucleolar dominance and in non-hybrids that regulate the effective dosage of their rRNA genes in response to the physiological needs of the cell.

Members of the lab are identifying the chromatin modifying activities involved in the epigenetic on-off switch, using RNA interference (RNAi) to knock down the expression of targeted genes in A. suecica. Histone deacetylases HDA6 and HDT1, the de novo DNA methyltransferase DRM2, and methylcytosine binding proteins MBD6 and MBD10 have been identified in the screens thus far. Both HDA6 (see photo below) and HDT1 localize, at least in part, within the nucleolus, where rRNA gene transcription takes place, suggesting that they may act directly on the rRNA genes. MBD6 is interesting because it colocalizes with condensed, inactive rRNA genes. We are making use of a variety of genetic, cytogenetic and biochemical approaches in order to understand the mechanism(s) of action of these chromatin modifying activities and to understand how their actions are intertwined to comprise the on-off switch.

In 2008, we discovered evidence that biogenesis of 24 nt siRNAs is required for nucleolar dominance. Knocking down RNA-DEPENDENT RNA POLYMERASE 2 (RDR2) or DICER-LIKE 3 (DCL3), which are required for 24 nt siRNA production, is sufficient to turn on the normally silenced A. thaliana rRNA genes in A. suecica.24 nt siRNAs direct the de novo methylation of corresponding DNA sequences, consistent with our finding that DRM2 is also required for nucleolar dominance. Because differential basepairing of siRNAs with A.thaliana versus A. arenosa rRNA gene sequences has the potential to explain the choice mechanism in nucleolar dominance, we are eager to understand the role of siRNAs in rRNA gene silencing.

For more details, see the following papers:

Preuss, Sasha, Pedro Costa-Nunes, Sarah Tucker, Olga Pontes, Richard J. Lawrence, Rebecca Mosher, Kristin D. Kasschau, James C. Carrington, David C. Baulcombe, Wanda Viegas and Craig S. Pikaard (2008). Multi-megabase silencing in nucleolar dominance involves siRNA-directed DNA methylation and specific methylcytosine binding proteins. Molecular Cell (Dec. 5, 2008 issue).

Preuss Sasha and Craig S. Pikaard (2007). rRNA gene silencing and nucleolar dominance: Insights into a chromosome-scale epigenetic on/off switch.Biochim Biophys Acta. 1769: 383-392 Pubmed; PDF

Earley, Keith, Richard J. Lawrence, Olga Pontes, Rachel Reuther, Angel J. Enciso, Manuela Silva, Nuno Neves, Michael Gross, Wanda Viegas, and Craig S. Pikaard (2006). Erasure of histone acetylation by Arabidopsis HDA6 mediates large-scale gene silencing in nucleolar dominance. Genes & Development 20:1283-1293. Pubmed PDF

Lawrence, Richard J., Keith Earley, Olga Pontes, Manuela Silva, Z. Jeffrey Chen, Nuno Neves, Wanda Viegas and Craig S. Pikaard (2004). A concerted DNA methylation/histone methylation switch regulates rRNA gene dosage control and nucleolar dominance. Molecular Cell 13:599-609. Pubmed PDF

Probst, Aline V., Mathilde Fagard, Florence Proux, Philippe Mourrain, Stéphanie Boutet, Keith Earley, Richard J. Lawrence, Craig S. Pikaard, Jane Murfett, Ian Furner, Hervé Vaucheret & Ortrun Mittelsten Scheid (2004). Arabidopsis histone deacetylase HDA6 is required for maintenance of transcriptional gene silencing and determines nuclear organization of rDNA repeats. Plant Cell 16:1021-1034. Pubmed PDF

Lewis, Michelle S., James M. Cheverud, and Craig S. Pikaard. (2004). Evidence for NORs as the units of regulation in nucleolar dominance in Arabidopsis thaliana inter-ecotype hybrids. Genetics 167:931-939. Pubmed PDF

Chen, Z. Jeffrey, Luca Comai and Craig S. Pikaard (1998). Gene dosage and stochastic effects determine the severity and direction of uniparental rRNA gene silencing (nucleolar dominance) in Arabidopsis allopolyploids. Proc. Natl. Acad. Sci. USA 95: 14891-14896. Pubmed PDF

Chen, Z. Jeffrey and Craig S. Pikaard (1997). Transcriptional analysis of nucleolar dominance in polyploid plants: biased expression/silencing of progenitor rRNA genes is developmentally regulated in Brassica. Proc. Natl. Acad. Sci. USA 94: 3442-3447. Pubmed PDF

Chen, Z. J. and Craig S. Pikaard (1997). Epigenetic silencing of RNA polymerase I transcription: a role for DNA methylation and histone modification in nucleolar dominance. Genes & Development 11: 2124-2136. Pubmed PDF