Yeast Two‐hybrid Reveals a Potato Polypyrimidine Tract Binding (PTB) Protein Interacts with a Nova‐like Protein

Abstract

PTB proteins are RNA‐binding proteins that contain four RNA recognition motifs. In potato, four PTB‐like genes, designated as PTB1, PTB6, TC201749 and PTB7, have been identified. In plants, these proteins play an important role in alternative splicing. In order to understand the multifunctional role of PTB proteins of plants we performed yeast two‐hybrid screening using PTB1 protein as bait. PTB1 protein was found to interact with a Nova‐like protein, termed StNova‐like protein. Nova (neuron‐oncological ventral antigen) protein is a RNA‐binding protein that contains three K‐homology domains. In humans, these proteins play an important role in regulation of neuronal RNA metabolism but the role of Nova‐like proteins in plants has not been explored. In this study, the StNova‐like gene has been cloned into the pMAL‐c2E vector, expressed in T7 express cells and purified over Ni‐NTA resin. Further, putative interactions between StPTB proteins and StNova were confirmed by pull‐down experiments and by blue‐native PAGE. Using a random 21‐mer phage‐peptide library, we also identified a consensus peptide sequence motif [S/G][I/L][L/I][S] that binds to PTB1, suggesting that the PTB1 protein might be interacting with the [S][I][I][S] sequence present in StNova‐like protein. Our studies suggest that in plants, Nova‐like proteins may be involved in processes of RNA metabolism mediated by PTB proteins.