The role of RNA binding proteins in post-transcriptional gene regulation and plant biology.

From our interactome capture project we envision that we will isolate many proteins that we previously not known to bind RNA. For such RBPs, we intend to carry out in-depth functional analysis to gain greater insights into post-transcriptional gene regulatory mechanisms from which many interesting avenues of research may arise. Firstly, mutational analysis will be carried out to not only reveal the roles of the genes in the plant are, but what roles the RNA binding function confers. Mutagenesis may include targeted mutagenesis using CRISPR, to specifically edit a potential RNA domain, or inhibition of the RNA binding domain via competition with a dominant negative transgene in which only the RNA binding domain is expressed. Additionally, do the RNA-binding domains of these proteins correspond to important mutational hotspots, as is the case for many animal RBPs? Such RBPs can then be specifically analysed via CLIP-seq to investigate which mRNAs they are binding; in the case of miRNA associated RBPs, are they selectively binding to strongly-regulated miRNA targets? Can overexpression of these RBPs promote miRNA-mediated silencing? Are there housekeeping enzymes that have a bi-functional role, where they can bind RNA? Will this reveal novel mechanisms of post-transcriptional regulation in plants?