Poster abstracts
Poster number 105 submitted by Jian Wu
Genome-wide functional analysis of PSTVd reveals wide-spread GU wobble base pairs critical for survival of an infectious noncoding viroid RNA
Jian Wu, Cuiji Zhou, Ying Wang, James Li, Dana Driver (Dept. of Molecular Genetics, Graduate Program in Molecular, Cellular, and Developmental Biology, Center for Applied Plant Sciences, and Center for RNA Biology, The Ohio State University, Columbus, OH ), Neocles Leontis (Dept. of Chemistry, Bowling Green State University, Bowling Green, OH 43403), Craig Zirbel (Dept. of Mathematics and Statistics, Bowling Green State University, Bowling Green, OH 43403), Xiaorong Tao (Dept. of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, PR China)
Abstract:
Potato spindle tuber viroid (PSTVd) is a noncoding circular RNA of 359 nt that replicates and systematically spreads in host plants, and thus all biological functions required to establish infection are accomplished through regulatory structural elements within the RNA genome itself. GU wobble base pairs are predicted to be present in most, if not all, structured RNAs. A few studies have demonstrated critical roles for certain specific wobble pairs in mediating RNA-protein interactions underpinning the functions of those RNAs. However, the lack of extensive molecular studies to identify and characterize the roles of multiple GU wobble pairs within a single RNA limits our understanding of their biological significance. To this end, we systematically analyzed the genome-wide presence and function of GU wobble pairs in PSTVd. Selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) confirmed the presence of 17 GU/UG wobble pairs in the PSTVd genome. Deep sequencing of natural full-length PSTVd variants derived from a cloned master sequence, performed using a novel approach, showed that GU/UG wobble pairs were conserved to the same extent as canonical Watson-Crick pairs. Comprehensive mutational analysis identified GU/UG pairs critical for PSTVd infectivity, including RNA replication, proper processing of replication intermediates, and/or systemic trafficking. In addition, we found that two specific wobble pairs are required for PSTVd trafficking into the host vascular system. This study represents the first genome-wide functional analysis of GU wobble pairs, and confirms their critical roles in the survival of an infectious RNA. Our findings have significant implications for understanding RNA structure-based regulation of replication and intercellular movement of both pathogen and cellular RNAs.
This work was supported by NSF grant IOS-1354636 to B. D. and D. M. B.
This abstract is dedicated to the memory of Dr. Biao Ding.
Keywords: Potato spindle tuber viroid , GU wobble, Replication