Elucidation of the RNA Recognition Code for Pentatricopeptide Repeat Proteins Involved in Organelle RNA Editing in Plants

Abstract

Pentatricopeptide repeat (PPR) proteins are eukaryotic RNA-binding proteins that are commonly found in plants. Organelle transcript processing and stability are mediated by PPR proteins in a gene-specific manner through recognition by tandem arrays of degenerate 35-amino-acid repeating units, the PPR motifs. However, the sequence-specific RNA recognition mechanism of the PPR protein remains largely unknown. Here, we show the principle underlying RNA recognition for PPR proteins involved in RNA editing. The distance between the PPR-RNA alignment and the editable C was shown to be conserved. Amino acid variation at 3 particular positions within the motif determined recognition of a specific RNA in a programmable manner, with a 1-motif to 1-nucleotide correspondence, with no gap sequence. Data from the decoded nucleotide frequencies for these 3 amino acids were used to assign accurate interacting sites to several PPR proteins for RNA editing and to predict the target site for an uncharacterized PPR protein.

An artificial PPR scaffold for programmable RNA recognition

“…The particular amino acids found at positions 4 and 34 of our cPPR are asparagine and glutamate, respectively ('ND'). Bioinformatic and recent structural analysis predicted that this combination would bind uracil [8][9][10] . We performed RNA electrophoretic shift assays with RNA homopolymers and observed specific binding of our designed cPPR to poly(U) RNA but no other RNA homopolymer (Fig.…”

“…A significant correlation was found between the identities of amino acids at positions 4 and 34 and particular bases within the RNA footprint [8][9][10] . Furthermore, one study indicated that the identity of the amino acid at position 1 might fine-tune base recognition 9 . Recently the structures of two distinct PPR-RNA complexes have been described at atomic resolution 18,19 .…”

“…Computational studies of a large family of RNA-binding proteins, known as pentatricopeptide repeat (PPR) proteins, have predicted that they bind their targets in a modular and sequencespecific manner [8][9][10] . PPR proteins contain a repeated motif that is typically 35 amino acids in length and folds into two anti-parallel alpha helices [11][12][13][14] .…”

“…Some PPR proteins appear to consist almost entirely of tandem PPRs, while some contain other domains, such as endonuclease or protein interaction domains [15][16][17] . Statistical correlations between specific PPR residues and RNA bases within their binding sites have elucidated a potential code for RNA recognition by PPR proteins [8][9][10] . A significant correlation was found between the identities of amino acids at positions 4 and 34 and particular bases within the RNA footprint [8][9][10] .…”

“…Statistical correlations between specific PPR residues and RNA bases within their binding sites have elucidated a potential code for RNA recognition by PPR proteins [8][9][10] . A significant correlation was found between the identities of amino acids at positions 4 and 34 and particular bases within the RNA footprint [8][9][10] . Furthermore, one study indicated that the identity of the amino acid at position 1 might fine-tune base recognition 9 .…”