Programmed RNA self-assembly

Chenhui Hao, Purdue University

Abstract

RNA nanotechnology has attracted tremendous attention recently and is rapidly developing. The versatile functionalities of RNA molecules are the hotspots of RNA nanotechnology. And one major direction for realizing functionality is that well defined and assembled RNA nanostructures acting as carriers decorated by diversely functional biological molecules. Upon cell delivery, the multi-functional RNA nanostructures release the large copies of functional molecules in cells and effect specifically. However, the well defined RNA nanostructures are limitedly available, yet the fragility of RNA against the RNase and the complexity of RNA self-folding and tertiary structures shy away scientists. My major research topics focus on developing novel RNA motifs and study their programmed self-assembly. Both canonical basepaired and non-canonical natural RNA motifs are utilized to program the assembly of RNA nanostructures, specifically RNA nanorings and RNA prisms. RNA structures resemble DNA structures in terms of canonical Watson-Crick basepairs: A (Adenine) complementary to U (Uracil) and C (Cytosine) complementary to G (Guanine). Since DNA has been explored to build diverse DNA motif such as double crossover, three-point-star motif, we expect that this nanotechnology can be adapted to RNA self-assembly. T-linkage has been developed as a connection between two DNA motifs for the DNA assembly. In my Ph.D research, we grafted the "T-linkage" onto RNA self-assembly to connect two RNA motifs. RNA nanorings were assembled with high yield and uniform size by the designed curvature. Replacement part of RNA molecule with DNA molecule can increase the stability of RNA nanostructures. Thus, we replace one strand of the RNA duplex with DNA strand or all the RNA duplex with DNA duplex for the assembly. We found RNA/DNA hybrid nanorings were formed because it retains A-form of duplex, while DNA/DNA formed random linear structures due to its B-form duplex. Non-canonical basepairing makes varieties of natural RNA motifs, which provide a large scope platform to assemble RNA nanostructures. One of those natural RNA motifs is packaging RNA (pRNA) of Φ29 bacteriophage, which has been firstly assembled into RNA oligomers in 1998. However, the well controlled and addressable pRNA nanostructure has not yet been reported. In my Ph.D research, we designed the two pRNA trigonal prisms and tetragonal prism referring to the pRNA crystal structure. The prisms were assembled with high yield and characterized by Cryo-EM and single particle reconstruction. Since pRNA oligomers were proved to efficiently carry functional RNA moleculs, our prisms will be competitive candidates for RNA therapeutic applications.

Degree

Ph.D.

Advisors

Mao, Purdue University.

Subject Area

Analytical chemistry

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