Identification of RNA aptamer which specifically interacts with PtdIns(3)P

The phosphinositide PtdIns(3)P plays an important role in autophagy; however, the detailed mechanism of its activity remains unclear. Here, we used a Systematic Evolution of Ligands by EXponential enrichment (SELEX) screening approach to identify an RNA aptamer of 40 nucleotides that specifically recognizes and binds to intracellular lysosomal PtdIns(3)P. Binding occurs in a magnesium concentration- and pH-dependent manner, and consequently inhibits autophagy as determined by LC3II/I conversion, p62 degradation, formation of LC3 puncta, and lysosomal accumulation of Phafin2. These effects in turn inhibited lysosomal acidification, and the subsequent hydrolytic activity of cathepsin D following induction of autophagy. Given the essential role of PtdIns(3)P as a key targeting molecule for autophagy induction, identification of this novel PtdIns(3)P RNA aptamer provides new opportunities for investigating the biological functions and mechanisms of phosphoinositides.