Unraveling the mysteries of gene silencing in nucleolar dominance and RNA-directed DNA methylation

Invited and Hosted by Plant Biology Graduate Students

Abstract:
My lab is interested in how genes can be singled out for silencing. A longstanding interest is the selective silencing of some, but not all, of the hundreds of nearly identical 45S ribosomal RNA (rRNA) genes present in a eukaryotic genome, an epigenetic phenomenon known as nucleolar dominance. Decades ago, we found that the inactive set of rRNA genes is selectively silenced, involving DNA methylation and histone modifications. However, it remained a mystery how different subsets of rRNA genes can be chosen for activation or silencing. A breakthrough came when we found that the active and inactive rRNA genes of Arabidopsis thaliana were present on different chromosomes, indicating that silencing is not decided one rRNA gene at a time but instead occurs on a sub-chromosomal scale, involving rRNA gene clusters, known as nucleolus organizer regions (NORs), that span 4-5 million base pairs of DNA. We recently sequenced the two NORs of A. thaliana, closing the final two gaps in the genome and allowing us to determine, for the first time in any eukaryote, the positions of active and inactive rRNA genes and their associated chromatin modifications. These studies have revealed surprising new insights into how NORs are regulated.

A second interest of the lab is RNA-directed DNA methylation, a process that is critical for silencing selfish genetic elements, such as transposons, throughout the genome. The process involves two plant-specific multi-subunit RNA polymerases, abbreviated as Pol IV and Pol V, that evolved as specialized forms of DNA-dependent RNA polymerase II, the enzyme that synthesizes mRNAs. Pol IV collaborates with an RNA-dependent RNA polymerase, RDR2 to convert double-stranded DNA sequences into double-stranded RNA. These double-stranded RNAs are then diced by DCL3 into short interfering RNAs (siRNAs) that associate with the ARGONAUTE 4 protein to direct the DNA methylation and silencing of matching DNA sequences transcribed by Pol V. By recapitulating the enzymatic reactions of the pathway in the test tube, we have uncovered how the reaction mechanisms of Pol IV, RDR2, DCL3, AGO4 and Pol V generate an RNA code that provides the instructions for what will happen in each successive step of the pathway.

About the Speaker:
Craig Pikaard earned a B.S. degree in Horticulture at The Pennsylvania State University in 1980, earned a Ph.D. in Plant Physiology at Purdue University in 1985, conducted postdoctoral research at the Fred Hutchinson Cancer Research Center in Seattle from 1996-1990, and began his faculty career at Washington University in St. Louis in 1990 before moving to Indiana University in 2009. At Indiana University, Craig holds the Carlos O. Miller Professorship and is a Distinguished Professor in the Department of Biology and the Department of Molecular and Cellular Biochemistry. He is a fellow of the American Association for the Advancement of Science, a member of the National Academy of Sciences and an Investigator of the Howard Hughes Medical Institute. He is the recipient of the American Society of Plant Biologists’ Martin Gibbs Medal and Indiana University’s Bicentennial Medal. Pikaard’s lab is known for their studies gene activation and silencing that combine the disciplines of genetics, genomics, biochemistry and cell biology.

Contact

  Alisa Chernikova
  azc6415@psu.edu