Nat. Biotechnol. published online 16 September 2012; doi:10.1038/nbt.2355

Synthetic biology efforts have generated modular gene circuits that enable the engineering of genetic and metabolic pathways that are integrated with or are orthogonal to normal cellular pathways. However, experience has shown that the complex regulatory elements that govern transcription and translation can confound synthetic biology efforts that require precise, switchable and predictable changes in gene expression within cells. Qi et al. report a way to mitigate these challenges by fragmenting the mRNA of multigene constructs into modular functional expression units. The clustered regularly interspaced short palindromic repeat (CRISPR) pathway from bacteria—in which a CRISPR-associated endoribonuclease (Cys4) binds and cleaves at specific repetitive RNA sequences—was used as a targeted mRNA cleavage platform. By inserting CRISPR cleavage sites between promoters, 5′ UTRs possibly containing regulatory sequences and open reading frames, the authors showed that CRISPR-mediated RNA processing could normalize previously variable protein expression across multiple genes of a construct, thereby rendering their composite behavior more predictable. Physical separation of RNA expression elements was also shown to enhance the responsiveness of individual modules to engineered regulatory motifs, such as antisense RNA. The demonstration that the system can be applied in several bacteria and in yeast suggests that CRISPR-mediated RNA processing may offer a useful adjunct tool to afford synthetic biologists greater control over their engineered circuits within cells.