SynBioFromLeukipposInstitute

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You’ve probably heard the cliché “life is complicated.” Maybe you have even picked
up Marie Kondo’s best-selling book The Life-Changing Magic of Tidying Up or clicked
on a link to a self-help article promising to guide you toward the recreation of a
simpler you by cutting out excess baggage—the things you probably don’t really need
to maintain your desired lifestyle. Perhaps some of these things are unnecessary redundancies
within your home, which would only get in the way if you wanted to reconfigure the
functionality of your living space.

Author(s): Choe, Joseph Harim | Advisor(s): Lim, Wendell | Abstract: Cell-based therapeutics have become a powerful method to combat cancer. Unlike small-molecule drugs or biologics, cells are able to integrate multiple inputs to execute complex response behaviors. Although cell-based therapies are poised to revolutionize cancer treatment, the field is still in its infancy in that it is still risky and often ineffective at treating most cancers. This dissertation focuses on applying the principles and tools of synthetic biology in T cells to develop the next generation of engineered T cells that will lead to more reliable, safe, and effective therapeutics. In order for engineered T cells to safely and reliably eradicate solid tumors, we need to generate T cells that are targeted specifically enough to prevent on-target, off-tumor toxicities but flexible enough to overcome the heterogeneous antigen expression patterns. To accomplish this, we developed a two-receptor system (a “prime-and-kill” circuit) that is capable of integrating information across multiple cells such that the T cells can target and kill tumor cells without damaging normal cells. We have demonstrated in vitro and in vivo that the prime-and-kill circuit is capable of eradicating tumors while sparing the normal tissues for which they may have had on-target, off-tumor toxicity. Another limitation for CAR T cell therapy is dependence on endogenous T cell circuitry to activate T cells to kill tumor cells. Due to such dependence, it has been difficult to control the level of T cell cytotoxic response. One approach to overcome this limitation is to enable T cells to have customizable input/output functions independent of endogenous T cell signaling. We developed a new chimeric force sensing receptor (A2-Notch receptor) to allow better tuning and control of the input/output function. By engineering the T cells to tune the levels of immune response through the A2-Notch receptor, T cells may be able to overcome the immunosuppressive blockade without causing unwanted side effects.The next wave of T cell therapies will need to be programmed to autonomously regulate sensing and response behaviors. These studies demonstrate the utility of using synthetic biology principles to facilitate T cell activity in order to generate safer and effective cell-based therapies.

Structural DNA nanotechnology is beginning to emerge as a widely accessible research tool to mechanistically study diverse biophysical processes. Enabled by scaffolded DNA origami in which a long single strand of DNA is weaved throughout an entire target nucleic acid assembly to ensure its proper folding, assemblies of nearly any geometric shape can now be programmed in a fully automatic manner to interface with biology on the 1–100-nm scale. Here, we review the major design and synthesis principles that have enabled the fabrication of a specific subclass of scaffolded DNA origami objects called wireframe assemblies. These objects offer unprecedented control over the nanoscale organization of biomolecules, including biomolecular copy numbers, presentation on convex or concave geometries, and internal versus external functionalization, in addition to stability in physiological buffer. To highlight the power and versatility of this synthetic structural biology approach to probing molecular and cellular biophysics, we feature its application to three leading areas of investigation: light harvesting and nanoscale energy transport, RNA structural biology, and immune receptor signaling, with an outlook toward unique mechanistic insight that may be gained in these areas in the coming decade.

The availability of renewable energy technologies is increasing dramatically across the globe thanks to their growing maturity. However, large scale electrical energy storage and retrieval will almost certainly be a required in order to raise the penetration of renewable sources into the grid. No present energy storage technology has the perfect combination of high power and energy density, low financial and environmental cost, lack of site restrictions, long cycle and calendar lifespan, easy materials availability, and fast response time. Engineered electroactive microbes could address many of the limitations of current energy storage technologies by enabling rewired carbon fixation, a process that spatially separates reactions that are normally carried out together in a photosynthetic cell and replaces the least efficient with non-biological equivalents. If successful, this could allow storage of renewable electricity through electrochemical or enzymatic fixation of carbon dioxide and subsequent storage as carbon-based energy storage molecules including hydrocarbons and non-volatile polymers at high efficiency. In this article we compile performance data on biological and non-biological component choices for rewired carbon fixation systems and identify pressing research and engineering challenges.

Technologies harnessing CRISPR systems have been rapidly evolving and expanding the capacity of researchers for understanding of mammalian cell behavi…

Molecular Biology; Neuroscience; Bioengineering; Biotechnology; Cell Biology; Biological
Sciences Research Methodologies

High-fidelity convergent total synthesis is used to produce Escherichia coli with a 61-codon synthetic genome that uses 59 codons to encode all of the canonical amino acids.

Synthetic biologists would like to be able to make gene regulatory circuits that mimic key properties of eukaryotic gene regulation. Taking a cue from multimeric transcription factor complexes, Bashor et al. developed synthetic transcriptional circuits that produce nonlinear behavior from cooperativity (see the Perspective by Ng and El-Samad). Their system uses clamp proteins with multiple protein-interaction domains. Circuit behavior can be tuned by altering the number or affinities of the interactions according to a mathematical model. The authors created synthetic circuits with desired functions common in biology, for example, switch-like behavior or Boolean decision functions.

Science , this issue p. [593][1]; see also p. [531][2]

Eukaryotic genes are regulated by multivalent transcription factor complexes. Through cooperative self-assembly, these complexes perform nonlinear regulatory operations involved in cellular decision-making and signal processing. In this study, we apply this design principle to synthetic networks, testing whether engineered cooperative assemblies can program nonlinear gene circuit behavior in yeast. Using a model-guided approach, we show that specifying the strength and number of assembly subunits enables predictive tuning between linear and nonlinear regulatory responses for single- and multi-input circuits. We demonstrate that assemblies can be adjusted to control circuit dynamics. We harness this capability to engineer circuits that perform dynamic filtering, enabling frequency-dependent decoding in cell populations. Programmable cooperative assembly provides a versatile way to tune the nonlinearity of network connections, markedly expanding the engineerable behaviors available to synthetic circuits.

[1]: /lookup/doi/10.1126/science.aau8287
[2]: /lookup/doi/10.1126/science.aax4556