Poster abstracts

Poster number 78 submitted by Christopher Strulson

RNA Function Through Compartmentalization

Christopher A. Strulson (Chemistry, Pennsylvania State University), Rosalynn C. Molden (Chemistry, Pennsylvania State University; current institution Princeton University), Christine D. Keating (Chemistry, Pennsylvania State University), Philip C. Bevilacqua (Chemistry, Pennsylvania State University)

Abstract:
Compartmentalization of RNA is essential for many cellular functions such as RNP processing and assembly, gene silencing, and transcription control, and may have played a pivotal role in the emergence of life. Variation in local RNA concentration impacts numerous biomolecular functions, from binding to catalysis. However, effects of cellular compartmentalization on RNA function, including catalysis, are largely unknown. We use RNA partitioning in an aqueous two-phase system (ATPS) to mimic intracellular compartmentalization and crowding, and test the effect on the kinetics of phosphodiester bond cleavage by a series of two-piece hammerhead ribozymes. The ATPS consists of poly(ethyleneglycol) (PEG) 8 kDa, dextran 10 kDa, and reaction buffer, in which the PEG-rich and dextran-rich phases of the ATPS serve as chemically distinct compartments. Partitioning induces a 1,000-fold enrichment of long RNA into the dextran-rich phase. Local concentration of hammerhead ribozyme is controlled by varying the volume ratio of the PEG-rich and dextran-rich phases. For ribozymes with an accessible k_cat/K_m region, increasing the local concentration of RNA via compartmentation into the dextran-rich phase of the ATPS enhances the observed rate of cleavage. In particular, a 20-fold enhancement in cleavage rate is observed in an ATPS consisting of a 1:100 ratio of dextran to PEG-rich phase. This study demonstrates that compartmentalization can enhance RNA function, which may be important in RNA therapeutics, in vivo RNA folding, and the evolution of functional protocells.

Keywords: Compartmentalization, Catalysis