Techniques to program cellular behaviors are central in synthetic biology. Scientists have actually established many artificial genetic circuits that allow crafted cells to produce restorative representatives on demand in response to specific stimuli, such as oncoproteins (e.g., mutant KRAS), chemicals (e.g., caffeine), and physical stimuli (e.g., light). However, the majority of circuits are based on gene changes that control transcription from a synthetic expression system. This delays downstream signal transduction and as an outcome, transcription-based circuits are too slow for some therapeutic applications.1-3 “Killing a virus-infected cell, needing insulin to get your blood sugar to the best level … that all requirements to happen incredibly fast and gene switches are just too slow … [They go] through transcription, translation, protein secretion, which takes some time,” described Martin Fussenegger, professor of biotechnology and bioengineering at the University of Basel. In a study released in NAR, Fusseneggers group developed a programmable synthetic circuit to manage secretion of a protein of interest after translation.1 Synthetic circuits that count on post-translational control, mediated by engineered proteins, use a more fast method than transcription-based gene switches. Fusseneggers group produced a protease-mediated circuit they named POSH (post-translational switch). This circuit includes the transmembrane domain of a cleavable endoplasmic reticulum (ER) retention signal fused to a protein of interest. The system produces the preferred protein, which remains in the ER under resting conditions due to the retention signal. The POSH platform depends upon a personalized inducer-sensitive protease to launch the protein of interest from the ER. The protease is revealed in two parts, which integrate in the presence of an inducer to cleave the ER retention signal of the chimeric protein. The protein of interest is then launched from the ER and undergoes trafficking to the Golgi for protein secretion. “Its an extremely smart style,” said Mads Kaern, associate teacher in the Department of Molecular and cellular Medicine at University of Ottawa, who was not associated with the study. “Youre using cells as pharmacies, essentially … you trick the cells into producing the medicine that the body needs, and after that youre managing when and where and how much is launched.”Killing a virus-infected cell, needing insulin to get your blood glucose to the best level … that all needs to happen incredibly fast and gene switches are just too sluggish. – Martin Fussenegger, University of BaselFusseneggers group revealed that they might manage POSH with a variety of stimuli, including chemical inducers, light, and electrostimulation. They also demonstrated the adaptability of this circuit to program protein secretion in numerous mammalian cell lines. The scientists initially defined POSH control in HEK293T cells, and then developed inducible protein secretion in cancer-derived cells lines (HeLa and HEPG2) and a non-human mammalian cell line (COS-7). This variety of efficient stimuli and applicable systems highlighted that POSH is a versatile, controllable platform for a variety of scientific opportunities.” [We were] hoping that others would find what they need to utilize this technology in their particular research,” Fussenegger explained.POSH is not the very first reported protease-mediated post-translational synthetic circuit. Fusseneggers research study follows publications on programmable systems by research study groups at Stanford University and the University of Ljubljana– all three groups developed robust synthetic circuits for managing cellular behavior in culture.2,3 Fusseneggers team took their system one step further, into a rodent design of type 1 diabetes. They verified their platform in vivo as a cell treatment by implanting POSH-engineered HEK293T cells into the mice, and as a gene treatment by injecting mice with a plasmid coding for POSH components. The researchers then injected mice with abscisic acid to induce insulin secretion. This caused a prolonged increase of insulin levels in the blood stream, showing that POSH can be utilized for on-demand secretion of insulin to normalize hyperglycemia in type 1 diabetic mice.This work positions post-translational circuits like POSH for potential restorative avenues in the future treatment of hormonal agent secretion-based conditions such as diabetes. Swank functions a quicker reaction than transcription-based gene switches, and is an important tool for post-translational control and programs of cell habits in culture and beyond.ReferencesM. Mansouri et al., “Design of programmable post-translational switch control platform for on-demand protein secretion in mammalian cells,” NAR, gkac916, 2022. A.E. Vlahos et al., “Protease-controlled secretion and display screen of intercellular signals,” Nat Commun, 13( 1 ):912, 2022. A. Praznik et al., “Regulation of protein secretion through chemical regulation of endoplasmic reticulum retention signal cleavage,” Nat Commun, 13( 1 ):1323, 2022.
In a study published in NAR, Fusseneggers group established a programmable synthetic circuit to control secretion of a protein of interest after translation.1 Synthetic circuits that rely on post-translational control, mediated by crafted proteins, provide a more quick method than transcription-based gene switches. The protein of interest is then released from the ER and goes through trafficking to the Golgi for protein secretion. They likewise showed the flexibility of this circuit to program protein secretion in a number of mammalian cell lines. The scientists first identified POSH control in HEK293T cells, and then developed inducible protein secretion in cancer-derived cells lines (HeLa and HEPG2) and a non-human mammalian cell line (COS-7). Mansouri et al., “Design of programmable post-translational switch control platform for on-demand protein secretion in mammalian cells,” NAR, gkac916, 2022.