It’s that time of year again: time to attempt the sisyphean task of maintaining peace between relatives who are constitutionally incompatible. Suppose you want both the uncanny gifting skills of Aunt Mildred and the mouth-watering cooking prowess of Uncle Frank. But if you allow them in the same room at the same time, both the gifts and the casseroles will end up smashed into indistinguishable, tear-soaked smears against the wall.
A similar problem often faces biochemical engineers working with cell-free transcription/translation (TXTL) mixes. These extracts – essentially cell juice – are powerful tools for testing genetic circuits, producing proteins, and making making other small molecules without the complication of living cells. However, sometimes people may want to combine chemical reactions or gene circuits that simply can’t work together. Without the membrane barriers of cells, combining those reactions would be as disastrous as sitting Uncle Frank and Aunt Mildred next to each other for a holiday dinner.
Gene Circuit Communication Across Vesicles
One workaround is to encapsulate TXTL mixes inside artificial lipid vesicles – making what amount to cell-free cells. These offer physical separation without the messiness of real cells. A recent paper from Katarzyna Adamala and Daniel Martin-Alarcon, in Edward Boyden’s lab at MIT, demonstrates some tips for separating incompatible reactions inside vesicles, just in time for the holidays.
In one demonstration, they made two different gene circuits inside separate vesicles communicate with each other. The first “sensor” vesicles contained IPTG and an arabinose-inducible gene for a membrane pore. When membrane-permeable arabinose diffused into the sensor vesicles, they expressed the pore and released IPTG. The second “reporter” vesicles had an IPTG-inducible luciferase gene and were primed with pre-expressed membrane pores. So when the sensor vesicles sensed arabinose and released IPTG, the reporter vesicles sensed that IPTG and expressed luciferase.
Encapsulation Within Vesicles Allows Incompatible Reactions To Play Nice
In another demonstration, the authors turned to reactions that are chemically incompatible, placing mammalian nuclear extract (for transcription) in one set of vesicles and cytoplasmic extract (for translation) in another set. The two extracts are incompatible – they don’t work if placed together in the same vesicle. However, if they incubated the transcription mix with DNA encoding luciferase in one set of vesicles, and then used membrane SNARE proteins to fuse those transcriptional vesicles with a second set containing translation extract, the stepwise combination effectively converted the luciferase gene from DNA to RNA to protein.
If you have chemically incompatible in-laws coming for the holidays, you may want to take a page from this paper and keep them physically constrained in different compartments – maybe encapsulate Aunt Mildred in the living room playing piano and Uncle Frank in the family room playing with the kids. Just be sure to get dinner out in time, or you may have to read up on vesicle cannibalism. For more, check out the paper in Nature Chemistry, 2016.