Bridging top-down and bottom-up approaches to synthetic biology
- Gene regulation and imaging
- Top-down
Regulation and readout of gene expression via general mRNA binding protein. - Communication
- Bottom-up
Synthetic minimal cells - artificial bioreactors interfacing with natural cells.
My research focuses on developing and applying tools for readout of mammalian cell states and for control of cellular processes, achieved via combining top-down and bottom-up approaches to synthetic biology.
The top-down approach involves building on the biological ensemble, modifying existing cellular pathways to explore and control biology.
My works focuses on building gene expression quantification and control tools via engineering single strand RNA binding proteins (more).
The bottom-up approach involves building chemical systems capable of mimicking cellular processes, such as protocells with replicating RNA and minimal peptide enzymes that couple catalysis to protocell fitness.
We aim at using those systems to develop synthetic minimal cell technology, to process chemical signals between mammalian cells and the environment (more).
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bottom-up synthetic biology
Building artificial chemical systems mimicking cells
I have developed the first known system capable of non-enzymatic RNA synthesis inside model protocells.
In this work, I have shown how citric acid can stabilize fatty acid liposomes in presence of divalent cations, allowing for non-enzymatic template director RNA primer extension inside the liposomes. This discovery bridges the RNA-world hypothesis or earliest life having RNA-based metabolism with the origin of compartmentalization based on fatty acid liposomes. (Adamala and Szostak, Science 2013)
I was also involved in discovering a chemically-driven replication mechanism for model protocells (Zhu, Adamala, Zhang and Szostak, PNAS 2012).
Together, those two projects allow for drawing a plausible, complete protocell cycle: inside the protocells, RNA is replicated by template-directed non-enzymatic synthesis, and the protocells grow into filamentous threads by absorbing fatty acid molecules from micelles and divide in response to shear stress.
in protocells
via semi-permeable bilayer
As part of the same overarching goal of building a chemical system capable of Darwinian evolution, I have shown that an encapsulated small peptide catalyst can impact the fitness of model protocells.
I have studies catalysis of Ser-His and other di- and tri- amino acid catalytic peptides. I have shown that the minimal serine protease analogue, Ser-His, can catalyze formation of peptide nucleic acids (Gorlero et.al FEBS Letters 2009). I have also demonstrated how the same dipeptide catalyst's activity can be enhanced by the presence of fatty acid protocell vesicles, and in turn the product of the reaction catalyzed by said dipeptide allows the protocell to enhance uptake of membrane building blocks, resulting in protocell growth.
This couples the activity of encapsulated catalyst with the fitness of the protocell, in a model system exploring the origin of Darwinian selection. (Adamala and Szostak, Nature Chemistry 2013).
Work done with Pier Luigi Luisi from University Roma Tre and Jack Szostak from Harvard University.
Peer reviewed publications list: protobiology.org/publications
Broader impact
The effort towards elucidating the origin and earliest evolution of life has always been receiving interest from broad audience.
As an example: an external review detailing my work on RNA replication in protocells: Angewandte Chemie International Edtion Citric Acid and the RNA World 2014, 53, 5245 - 5247, external link
My work on protocells have been featured in science news outlets and editorials, including:
Science Focus: Robert Service, The Life Force 29 Nov 2013 external link
BioTechniques: How Cellular Life Evolved 06 Jan 2014 external link
Science News: To cook up life, just add citrate 185(1):15 external link
Science Daily: Researchers find missing component in effort to create primitive, synthetic cells, 28 Nov 2013 external link
Chemical & Engineering News: Lab-Made Protocells Show Hints Of Evolution, 91(21), 27 May 2013
The Panda's Thumb: New Szostak protocell is closest approximation to origin of life and Darwinian evolution so far, 13 Dec 2013 external link
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top-down synthetic biology
Engineering generalized RNA-protein interactions: a toolbox for regulation and readout of gene expression
We are developing and validating a protein architecture which binds to single stranded RNA. Using this protein technology, we are developing a tool for visualization and quantification of levels of expression of genes of interest.
This tool will work by following the reconstitution of a protein probe upon interaction of sequence-specific RNA binding proteins with the mRNA of the gene of interest.
We are also aiming to edit the transcriptome of the gene of interest, selectively decreasing the level of expression of one splice variant (as opposed to cutting the DNA of the gene, which targets all splice variants indiscriminately).
This technology could potentially help in studying non-ER translation events, elucidating mechanisms of synaptic plasticity, as well as studying healthy and diseased translational profiles of genes, e.g., those involved in oncogenesis and other disease processes.
The ability to monitor and perturb RNA in living neurons - which would open up the investigation of many processes that contribute to development, plasticity, and disease progression - would benefit greatly from a method of systematically targeting unmodified RNA sequences for observation and control.
The currrent ssRNA targeting methods are based on the RNA binding protein aptamers, like MS2 or PP7; this require the introduction of aptamer binding sites into the RNA. My work shows that it is possible to develop a ssRNA binding protein that can be engineered to target arbitrary sequences of variable length, thus eliminating the need to engineer the target sites into the RNA of interest.
Work done in Synthetic Neurobiology lab with Ed Boyden.