New molecules required for silencing and the important role of siRNAs are detailed in Molecular Cell paper.
A team of investigators report a breakthrough in understanding nucleolar dominance, the silencing of an entire parental set of ribosomal RNA (rRNA) genes in a hybrid plant or animal. Their study identified several new players necessary for the silencing machinery in nucleolar dominance to function and also highlighted the key role of siRNA.
“siRNAs are not just regulating the selfish DNA or the junk DNA, but they’re regulating the really essential genes too,” points out Craig Pikaard, Ph.D., WUSTL professor of biology in Arts & Sciences, who led the research. Dr. Pikaard believes that siRNAs might be the key to understanding the choice mechanism underlying which parental genes get switched off and which get left on.
Nucleolar dominance occurs when nucleoli, protein-rich, dense regions of RNA within the nucleus, form on the chromosomes inherited from one parent, but not on the chromosomes inherited from the other parent. Expression of ribosomal RNA genes drives the formation of these nucleoli.
First in the pathway is RNA-dependent RNA Polymerase 2 (RDR2), which prepares a stretch of RNA for DICER-LIKE 3 (DCL3), an enzyme that chops up RNA transcripts into smaller segments. These smaller fragments of RNA become siRNAs, which then guide the de novo cytosine methyltransferase , DRM2, to the targeted genes. DRM2 is required to put a methyl group on ribosomal genes that had been active in the parental genome. MBD6 and MBD10, methylcytosine binding proteins, then adhere to the segments of DNA that have been methylated by DRM2. At the same time, HDA6, a histone deacetylase, modifies the proteins that act as spools for the DNA.
The end result of this convergent, siRNA-mediated pathway is the large-scale silencing of hundreds of clustered rRNA genes that span millions of basepairs of DNA, the scientists note.
If researchers could harness the silencing machinery involved in nucleolar dominance to limit the expression of rRNA genes, they could potentially slow the growth rate of tumor cells and thereby slow the progression of diseases like cancer, according to Dr. Pikaard.
Completely silencing all ribosomal genes would not be a viable therapeutic approach for cancer patients because ribosomes are necessary for survival. But Dr. Pikaard and his collaborators’ research suggests that siRNAs can direct silencing agendas that are much more sophisticated than an all or nothing approach.
To determine the pathway regulating nucleolar dominance, Dr. Pikaard’s team exploited RNAi to knockdown expression of target genes in a hybrid of two species of Arabidopsis. The genes knocked down were those coding for products that prior research had suggested might be involved in silencing. By knocking these suspects down one by one and assessing whether nucleolar dominance had been disrupted after each knockdown, Dr. Pikaard and his collaborators were able to determine which proteins and RNAs were necessary to keep the silenced parental genes off.
The study was published December 4 in Molecular Cell.