110th Anniversary: Engineered Ribonucleic Acid Control Elements as Biosensors for in Vitro Diagnostics

Abstract

Regulatory mechanisms in biological systems are analogous to process control exerted in many chemical and biomolecular engineering processes. Ribonucleic acid (RNA) has been identified as a ubiquitous element for tight regulation of cellular activities, such as gene expression, given its high tunability of structures and functions. Enhanced understanding of interactions of RNA with nucleic acids, proteins, and small molecules has enabled programming of robust gene circuits for detection of specific target analytes and signal transduction. Bottom up approaches in synthetic biology have developed in silico engineered RNA modules to work as biosensors for nucleic acids in vitro detection. In this review, we describe light-up aptamers and toehold switches as exciting examples of RNA engineered modules that were inspired by evolution and process control in biological organisms. We emphasize how the programmability of these elements has empowered the assembly circuitry to sense and compute information relevant for biomedical applications.

Developments of Riboswitches and Toehold Switches for Molecular Detection—Biosensing and Molecular Diagnostics

“…In addition to the regulatory applications, the RNA-based regulatory system, specifically riboswitches and toehold switches, has attracted scientific interest to develop systems for molecular detection applications as biosensing and molecular diagnostics [1,2]. A riboswitch is a cis-regulation element of gene expression at the transcription or translation level [3][4][5].…”

Circular single-stranded DNA as a programmable vector for gene regulation in cell-free protein expression systems

“…Cell-free protein expression (CFE) systems have become an attractive alternative platform for engineering and designing biological systems in vitro [1][2][3] , and have thus been developed for various applications, including high-throughput drug screening 4 , point-of-care diagnostics [5][6][7][8][9][10] , biomanufacturing, and artificial cellular communication [11][12][13][14][15][16] . Vectors for protein expression in CFE systems are commonly circular double-stranded DNA molecules such as plasmids, or linear double-stranded DNA, which is usually obtained as a polymerase chain reaction (PCR) product 17 .…”

“…On the other hand, RNA-based CFE systems have increasingly received attention due to the abundant availability of RNA secondary structures and the possibility to manipulate these structures via hybridization and strand displacement reactions 6,29,30 . Riboswitches and toehold switches are the most representative types of RNA-based CFE systems at the post-transcriptional level that allow to activate or repress gene expression through structural transformations triggered by DNA or RNA sequences or other target molecules [31][32][33] .…”