DNAzyme-Mediated Genetically Encoded Sensors for Ratiometric Imaging of Metal Ions in Living Cells.

ABSTRACT

Genetically encoded fluorescent proteins (FPs) have been used for metal ion detection. However, their applications are restricted to a limited number of metal ions owing to the lack of available metal-binding proteins or peptides that can be fused to FPs and the difficulty in transforming the binding of metal ions into a change of fluorescent signal. We report herein the use of Mg2+ -specific 10-23 or Zn2+ -specific 8-17 RNA-cleaving DNAzymes to regulate the expression of FPs as a new class of ratiometric fluorescent sensors for metal ions. Specifically, we demonstrate the use of DNAzymes to suppress the expression of Clover2, a variant of the green FP (GFP), by cleaving the mRNA of Clover2, while the expression of Ruby2, a mutant of the red FP (RFP), is not affected. The Mg2+ or Zn2+ in HeLa cells can be detected using both confocal imaging and flow cytometry. Since a wide variety of metal-specific DNAzymes can be obtained, this method can likely be applied to imaging many other metal ions, expanding the range of the current genetically encoded fluorescent protein-based sensors.