sxRNA was originally described by the Tenenbaum Lab at SUNY Polytechnic Institute. Broadly speaking, sxRNA provides a method to make a message “switchable”, where we can control the functionality of the RNA-RBP complex as a result of a trans-interaction with a miRNA. We are using this mechanism both to control translation of an mRNA in order to selectively express protein in specific cell types, and to create a new class of miRNA inhibitors which can potentially be used as therapeutics.
In the cell, RNA messages are able to perform a wide array of vital functions because of their ability to dynamically fold into many different conformations. These structures are frequently regulated by RNA-Binding Proteins (RBPs). One common structure found in ribosomal RNA is the 3-way junction, and results from the RNA message folding on itself in a manner known as cis, producing 3 helices that meet around an unpaired region.
The sxRNA technology takes advantage of this mechanism by demonstrating a trans interaction, wherein a separate miRNA binds this unpaired region, thereby stabilizing the 3-way junction and potentially reinforcing the binding of an RBP. These trans interactions have allowed us to design RNA messages (sxRNAs) which are targeted to miRNAs naturally occurring in the cell. Once the two meet, the stem loop can either be stabilized and potentially increase translation, or destabilized in order to turn “off” the message.
The image on the left shows a variety of situations where an mRNA is shown (i) without associated miRNAs and (ii) with these miRNAs. A number of different functional mechanisms for trans-acting modulation are depicted:
(A) An RBP whose binding site is hidden by an alternate structure. In the presence of the associated trans-acting ncRNA, the site is revealed.
(B) An RBP that recognizes a double stranded binding site that only forms via interaction with a separate ncRNA. (C) An RBP whose binding site must be presented in a particular orientation, with respect to its flanking sequence, that is facilitated by a 3WJ. (D) An RBP whose binding site is hidden by its sequence hybridizing with an ncRNA. (E) An RBP whose binding site structure is weak and may sample alternate structures over time. A trans-acting 3WJ stabilizes the active conformation, allowing the protein to bind. This combined model allows a single mRNA transcript variant to exhibit multiple behaviors amongst cell types via the combinatorial absence and presence (in varying concentrations) of both the relevant RBPs and, more specifically, ncRNAs. It is these latter that may facilitate a distinct regulation for sub-populations of expressed mRNAs, where a much broader group may hold the potential for control by the same RBP.