Predicting in vivo binding sites of RNA-binding proteins using mRNA secondary structure

Abstract

While many RNA-binding proteins (RBPs) bind RNA in a sequence-specific manner, their sequence preferences alone do not distinguish known target RNAs from other potential targets that are coexpressed and contain the same sequence motifs. Recently, the mRNA targets of dozens of RNA-binding proteins have been identified, facilitating a systematic study of the features of target transcripts. Using these data, we demonstrate that calculating the predicted structural accessibility of a putative RBP binding site allows one to significantly improve the accuracy of predicting in vivo binding for the majority of sequence-specific RBPs. In our new in silico approach, accessibility is predicted based solely on the mRNA sequence without consideration of the locations of bound trans-factors; as such, our results suggest a greater than previously anticipated role for intrinsic mRNA secondary structure in determining RBP binding target preference. Target site accessibility aids in predicting target transcripts and the binding sites for RBPs with a range of RNA-binding domains and subcellular functions. Based on this work, we introduce a new motif-finding algorithm that identifies accessible sequence-specific RBP motifs from in vivo binding data.

Quantifying RNA binding sites transcriptome-wide using DO-RIP-seq

“…As shown in Figure 2G, site accessibility was a stronger predictor of HuR binding when using DO-RIP-seq than for our previous HuR PAR-CLIP data (Friedersdorf and Keene 2014). This finding corroborates previous studies in concluding that HuR binds to specific sequences in unstructured regions of RNA (Meisner et al 2004;Li et al 2010). Taken together, these data show that increased sequence coverage allows for the unique ability of DO-RIP-seq to identify most of the strong binding sites, and thus also identify meaningful nonbound sites as demonstrated by the presence of local secondary structure surrounding nonbound sites.…”

Identifying mRNA sequence elements for target recognition by human Argonaute proteins

“…Nucleotide accessibility in the RNA secondary structure was calculated using RNAplfold (Bernhart et al 2006), which averages over all the sliding windows of size W to compute the probability of a nucleotide being unpaired with a maximal span of L nucleotides. In these experiments, we used W = 80 and L = 40, which were previously adopted for predicting siRNA and RBP binding (Tafer et al 2008;Li et al 2010). We used RNAplfold over other RNA folding programs because of its capability of robustly computing the local base-pairing probabilities without predefining the exact position of sequence window.…”

An Untranslated cis-Element Regulates the Accumulation of Multiple C4 Enzymes in Gynandropsis gynandra Mesophyll Cells

“…It is also possible that an RNA binding protein may recognize MEM2 in G. gynandra bundle sheath cells. PUF proteins recognize cis-elements present in mRNA that are typically within 39 UTRs, of a similar size to MEM2, and are dependent on secondary structure (Tadauchi et al, 2001;Francischini and Quaggio, 2009;Li et al, 2010b;Filipovska et al, 2011). However, PUF proteins typically recognize a sequence beginning with 59 UGUR (whereby R represents A or G) (Lu et al, 2009), which is not part of the conserved MEM2 sequence or directly upstream in any of the MEM2-containing UTRs.…”