Skip to main content
2023 BMES Annual Meeting Main banner
Icon Legend
This session is not in your schedule.
This session is in your schedule. Click again to remove it.
Back

    Cellular and Molecular Bioengineering

    (H-300) Microbial teamwork: circuits for in situ differentiation from seed cells

    Saturday, October 14, 2023
    9:30 AM – 10:30 AM PDT
    Location: Exhibit Hall - Row H - Poster # 300

      Presenting Author(s)

    • Arash Farhadi, PhD (he/him/his)

      Postdoctoral Scholar
      Massachusetts Institute of Technology
      Cambridge, Massachusetts, United States

      • Co-Author(s)

    • HK

      Hyoujin Kim

      Postdoctoral Scholar
      Massachusetts Institute of Technology, United States

    • PC

      Phillip Clauer

      Graduate Student
      Massachusetts Institute of Technology, United States

    • JA

      Juliana Abramson

      Undergraduate Student
      Massachusetts Institute of Technology, United States

      • Last Author(s)

    • CV

      Christopher A. Voigt

      Professor
      Massachusetts Institute of Technology, United States

    Introduction:: Single-cell organisms can perform many interesting functions as a community by distributing their collective tasks among different members of their population. This division of labor allows cells to collectively perform many tasks without burdening each individual cell with all the functions. There is great interest in developing robust and efficient genetic circuits that can similarly distribute tasks across cells in a population as we engineer microbes to perform many functions for our benefit. To achieve this, we have developed a genetic circuit, inspired by the hierarchical lineage differentiation of stem cells, which allows engineered cells to differentiate from a monoclonal progenitor cell type into a multi-member community with distributed tasks across the population. The design of these differentiation circuits can be used in broad scenarios including engineering probiotic bacteria to treat human diseases as well as engineering plant-associated microbes that provide nutrients directly to plants.

    Materials and Methods:: The differentiation circuit uses the DNA rearrangement reactions of serine integrases to differentiate the state of cells. Initially, cells are engineered to be in a progenitor state, with all the promoters that drive the output functions turned off. Upon activation of the differentiation circuit, the encoded integrase-logic stochastically differentiates the circuit to activate only one promoter, allowing the cell to produce a single output function. Furthermore, the differentiation circuits are genomically integrated into insulated landing pads to allow stable and robust differentiation of cells in challenging environments.

    Results, Conclusions, and Discussions::

    Cells engineered with differentiation circuits can be maintained for many generations in the quiescent progenitor state, and once activated, in the differentiated community. The topology of the differentiation landscape can be finely tuned to achieve a desired relative ratio of states from 5-90%. In addition, we have developed up to four member communities, and show a roadmap to scaling up the number of possible states using a combination of orthogonal integrase attachment sequences and ECF sigma factors. To characterize the function of these circuits in vivo, we engineered Klebsiella variicola (a maize-associated root epiphyte) with two- and three-state differentiation circuits. The engineered K. variicola cells can colonize the root of maize and differentiate into multi-member communities in situ upon receiving a small molecule differentiation cue.


     


    The division of labor genetic circuits will enable engineered cells to produce multiple resource-intensive products more effectively in challenging environments such as the human and rhizosphere-associated microbiomes.



    Acknowledgements (Optional): :

    References (Optional): :