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Within a cell, ATP is both the energy currency inside and an extracellular messenger outside. Recent data show that adenosine concentration is much higher in the insterstitium (outside) of solid tumors compared to healthy tissues, and this information corroborates the growing awareness that tumor cells build up a 'self-advantage' micro-environment that limits the anti-tumor immune response. Thus, understanding ATP concentration dynamics outside the cells is a must for upcoming cancer research, particularly if this can be done in the context of the natural tissue.
The italian researcher Patrizia Pellegatti and colleagues from Ferrara University, have engineered a chimeric plasma membrane-targeted luciferase that allows in vivo real-time imaging of extracellular ATP. With this novel probe, they have measured the ATP concentration within the niche of several experimentally-induced tumors. How they did it?
They previously engineered a chimeric luciferase-folate receptor construct in which they appended to luciferase cDNA the targeting sequences (leader sequence and GPI anchor) derived from the folate receptor. This novel probe, named pmeLUC (plasma membrane luciferase) is in fact targeted to and retained at the plasma membrane thus detecting ATP in the aqueous layer close to the cell surface.
So-called "reporter genes" are in fact reporting more than transcriptional activity, and some companies are exploring this new market sector.
Monitoring gene expression it is just one piece of the big reporter gene cake. In a recent issue of ACS Chemical Biology, the group of Keith Wood from Promega introduces new covalent, non-covalent and allosteric design configurations to turn firefly luciferase in a biosensor to detect different intermolecular interactions trough modulation of its luminescence activity. Basically the group aims to branch new strategies for luciferases as intracellular probes: not only "genetic reporters", but even reporters (biosensors) of intracellular molecules and events.
Some years ago Promega was working on design of luciferin (the luciferase substrate) to make biosensors. Moving the design from the substrate to the enzyme it’s a big paradigm shift inside the company: it means that to date it’s easier to design (and then market at lower price) a genetic entity like luciferase instead of a chemical entity like the luciferin substrate. Genetic engineering is still in its early stages: how much “copy and paste” we have to do yet with reporter sequences!
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