A Multiplex MoClo Toolkit for Extensive and Flexible Engineering of Saccharomyces cerevisiae
William M. Shaw, Ahmad S. Khalil and Tom Ellis
ACS Synthetic Biology, 12: 3393-3405 (2023)
Nanoscale Structural Mapping of Protein Aggregates in Live Cells Modeling Huntington’s Disease
Zhongyue Guo*, Giulio Chiesa*, Jiaze Yin, Adam Sanford, Stefan Meier, Ahmad S. Khalil and Ji-Xin Cheng
bioRxiv, doi: 10.1101/2023.10.09.561223 (2023)
Cooperative Assembly Confers Regulatory Specificity and Long-Term Genetic Circuit Stability
Meghan D. J. Bragdon*, Nikit Patel*, James Chuang, Ethan Levien, Caleb J. Bashor* and Ahmad S. Khalil*
Cell, 186: 3810-3825 (2023)
Mapping the Dynamics of Epigenetic Adaptation
Ajay Larkin*, Colin Kunze*, Melissa Seman, Alexander Levashkevich, Justin Curran, Ahmad S. Khalil and Kaushik Ragunathan
bioRxiv, doi: 10.1101/2023.07.10.548368 (2023)
Deep Neural Networks for Predicting Single-Cell Responses and Probability Landscapes
Heidi E. Klumpe*, Jean-Baptiste Lugagne*, Ahmad S. Khalil and Mary J. Dunlop
ACS Synthetic Biology, 12: 2367-2381 (2023)
Long-Term Evolution of Proliferating Yeast Cells Using the eVOLVER Platform
Daniel Garcia-Ruano, Akanksha Jain, Zachary J. Heins, Brandon G. Wong, Ezira Yimer Wolle, Ahmad S. Khalil and Damien Coudreuse
Open Biology, 13: 230118 (2023)
Human Herpesvirus 8 ORF57 Protein is Able to Reduce TDP-43 Pathology: Network Analysis Identifies Interacting Pathways
Chelsea J. Webber, Caroline N. Murphy, Alejandro N. Rondón-Ortiz, Sophie J. F. van der Spek, Elena X. Kelly, Noah M. Lampl, Giulio Chiesa, Ahmad S. Khalil, Andrew Emili and Benjamin Wolozin
Human Molecular Genetics, 32: 2966-2980 (2023)
Transcriptional Kinetic Synergy: A Complex Landscape Revealed by Integrating Modelling and Synthetic Biology
Rosa Martinez-Corral, Minhee Park, Kelly Biette, Dhana Friedrich, Clarrissa Scholes, Ahmad S. Khalil, Jeremy Gunawardena and Angela H. DePace
Cell Systems, 14: 324-339 (2023)
Effects of Antibiotic Interaction on Antimicrobial Resistance Development in Wastewater
Indorica Sutradhar, Carly Ching, Darash Desai, Zachary Heins, Ahmad S. Khalil and Muhammad H. Zaman
Scientific Reports, 13: 7801 (2023)
Unlocking the Magic in Mycelium: Using Synthetic Biology to Optimize Filamentous Fungi for Biomanufacturing and Sustainability
Charles Jo, Jing Zhang, Jenny M. Tam, George M. Church, Ahmad S. Khalil, Daniel Segrè and Tzu-Chieh Tang
Materials Today Bio, 19: 100560 (2023)
High-Throughput Continuous Evolution of Compact Cas9 Variants Targeting Single-Nucleotide-Pyrimidine PAMs
Tony P. Huang*, Zachary J. Heins*, Shannon M. Miller, Brandon G. Wong, Pallavi A. Balivada, Tina Wang, Ahmad S. Khalil and David R. Liu
Nature Biotechnology, 41: 96-107 (2023)
The Sound of Silence: Transgene Silencing in Mammalian Cell Engineering (Perspective)
Alan Cabrera, Hailey I. Edelstein, Fokion Glykofrydis, Kasey S. Love, Sebastian Palacios, Josh Tycko, Meng Zhang, Sarah Lensch, Cara E. Shields, Mark Livingston, Ron Weiss, Huimin Zhao, Karmella A. Haynes, Leonardo Morsut, Yvonne Y. Chen, Ahmad S. Khalil, Wilson W. Wong, James J. Collins, Susan J. Rosser, Karen Polizzi, Michael B. Elowitz, Martin Fussenegger, Isaac B. Hilton, Joshua N. Leonard, Lacramioara Bintu, Kate E. Galloway and Tara L. Deans
Cell Systems, 13: 950-973 (2022)
Multidimensional Control of Therapeutic Human Cell Function with Synthetic Gene Circuits
Hui-Shan Li*, Divya V. Israni*, Keith A. Gagnon, Kok Ann Gan, Michael H. Raymond, Jeffry D. Sander, Kole T. Roybal, J. Keith Joung, Wilson W. Wong and Ahmad S. Khalil
Science, 378: 1227-1234 (2022)

  Commentary in Science by Emanuel Salazar-Cavazos & Grégoire Altan-Bonnet (link); in Nature by Heidi Ledford (link); in The Scientist by Natalia Mesa (link); in El Pais by Daniel Mediavilla (link); in Nature by Heidi Ledford (link)
A Toolkit for Precise, Multi-Gene Control in Saccharomyces cerevisiae
Adam Sanford, Szilvia Kiriakov and Ahmad S. Khalil
ACS Synthetic Biology, 11: 3912-3920 (2022)
Enhancing Nutritional Niche and Host Defenses by Modifying the Gut Microbiome
Qing Sun*, Nic M. Vega*, Bernardo Cervantes, Christopher P. Mancuso, Ning Mao, Megan N. Taylor, James J. Collins, Ahmad S. Khalil, Jeff Gore and Timothy K. Lu
Molecular Systems Biology, 18: e9933 (2022)
Recent Progress of Gene Circuit Designs in Immune Cell Therapies
Seunghee Lee, Ahmad S. Khalil and Wilson W. Wong
Cell Systems, 13: 864-873 (2022)
High-Performance Multiplex Drug-Gated CAR Circuits
Hui-Shan Li*, Nicole M. Wong*, Elliot Tague, John T. Ngo, Ahmad S. Khalil and Wilson W. Wong
Cancer Cell, 40: 1-12 (2022)
Screening Microbially Produced Δ9-tetrahydrocannabinol Using a Yeast Biosensor Workflow
William M. Shaw, Yunfeng Zhang, Xinyu Lu, Ahmad S. Khalil, Graham Ladds, Xiaozhou Luo and Tom Ellis
Nature Communications, 13: 5509 (2022)
In Vivo Hypermutation and Continuous Evolution
Rosana S. Molina, Gordon Rix, Amanuella A. Mengiste, Beatriz Alvarez, Daeje Seo, Haiqi Chen, Juan E. Hurtado, Qiong Zhang, Jorge Donato Garcia-Garcia, Zachary J. Heins, Patrick J. Almhjell, Frances H. Arnold, Ahmad S. Khalil, Andrew D. Hanson, John E. Dueber, David V. Schaffer, Fei Chen, Seokhee Kim, Luis Angel Fernandez, Matthew D. Shoulders, and Chang C. Liu
Nature Reviews Methods Primer, 2: 37 (2022)
Modular Design of Synthetic Receptors for Programmed Gene Regulation in Cell Therapies
Iowis Zhu*, Raymond Liu*, Julie M. Garcia, Axel Hyrenius-Wittsten, Dan I. Piraner, Josef Alavi, Divya V. Israni, Bin Liu, Ahmad S. Khalil and Kole T. Roybal
Cell, 185: 1431-1443 (2022)

  Commentary in Nature by Mohamad Hamieh & Maria Themeli (PDF)
Once Cell, Many Fates (Perspective)
Colin Kunze and Ahmad S. Khalil
Science, 375: 262-263 (2022)
Here to Stay: Writing Lasting Epigenetic Memories (Preview)
Hagar F. Moussa*, James F. Angstman* and Ahmad S. Khalil
Cell, 184: 2281-2283 (2021)
Environmental Fluctuations Reshape an Unexpected Diversity-Disturbance Relationship in a Microbial Community
Christopher P. Mancuso, Hyunseok Lee, Clare I. Abreu, Jeff Gore and Ahmad S. Khalil
eLife, 10: e67175 (2021)
Computational Model To Quantify the Growth of Antibiotic-Resistant Bacteria in Wastewater
Indorica Sutradhar, Carly Ching, Darash Desai, Mark Suprenant, Emma Briars, Zachary Heins, Ahmad S. Khalil and Muhammad H. Zaman
mSystems, 6: e0036021 (2021)
A Code of Ethics for Gene Drive Research
George J. Annas, Chase L. Beisel, Kendell Clement, Andrea Crisanti, Stacy Francis, Marco Galardini, Roberto Galizi, Julian Grünewald, Greta Immobile, Ahmad S. Khalil, Ruth Müller, Vikram Pattanayak, Karl Petri, Ligi Paul, Luca Pinello, Alekos Simoni, Chrysanthi Taxiarchi and J. Keith Joung
CRISPR J, 4: 19-24 (2021)
Automated Continuous Evolution of Proteins In Vivo
Ziwei Zhong*, Brandon G. Wong*, Arjun Ravikumar, Garri A. Arzumanyan, Ahmad S. Khalil and Chang C. Liu
ACS Synthetic Biology, 9: 1270-1276 (2020)

  Commentary in preLights by Pavithran Ravindran (link)
Barcoded Microbial System for High-Resolution Object Provenance
Jason Qian*, Zhi-xiang Lu*, Christopher P. Mancuso*, Han-Ying Jhuang*, Rocío del Carmen Barajas-Ornelas*, Sarah A. Boswell*, Fernando H. Ramírez-Guadiana, Victoria Jones, Akhila Sonti, Kole Sedlack, Lior Artzi, Giyoung Jung, Mohammad Arammash, Mary E. Pettit, Michael Melfi, Lorena Lyon, Siân V. Owen, Michael Baym, Ahmad S. Khalil, Pamela A. Silver, David Z. Rudner and Michael Springer
Science, 368: 1135-1140 (2020)

  Commentary in Science by Jeff Nivala (PDF)
Protein Assembly Systems in Natural and Synthetic Biology
Giulio Chiesa*, Szilvia Kiriakov and Ahmad S. Khalil
BMC Biology, 28: 35 (2020)
Sphingomonas solaris sp. nov., Isolated From a Solar Panel in Boston, Massachusetts
Kristie Tanner, Christopher P. Mancuso, Juli Peretó, Ahmad S. Khalil, Cristina Vilanova and Javier Pascual
International Journal of Systematic and Evolutionary Microbiology, 70: 1814-1821 (2020)
Designing Automated, High-Throughput Continuous Cell Growth Experiments Using eVOLVER
Zachary J. Heins, Christopher P. Mancuso, Szilvia Kiriakov, Brandon G. Wong, Caleb J. Bashor and Ahmad S. Khalil
Journal of Visualized Experiments, 147: e59652 (2019)
Complex Signal Processing in Synthetic Gene Circuits Using Cooperative Regulatory Assemblies
Caleb J. Bashor*, Nikit Patel*, Sandeep Choubey, Ali Beyzavi, Jane Kondev, James J. Collins and Ahmad S. Khalil
Science, 364: 593-597 (2019)

  Commentary in Science by Andrew H. Ng & Hana El-Samad (PDF); in Nature Biotechnology by Ross Cloney (PDF)
Functional Genomics of the Rapidly Replicating Bacterium Vibrio natriegens by CRISPRi
Henry H. Lee, Nili Ostrov, Brandon G. Wong, Michaela A. Gold, Ahmad S. Khalil and George M. Church
Nature Microbiology, 4: 1105-1113 (2019)

  Commentary in Nature Microbiology by Jonathan D'Gama & Matthew Waldor (PDF)
Engineering Epigenetic Regulation Using Synthetic Read-Write Modules
Minhee Park, Nikit Patel, Albert J. Keung and Ahmad S. Khalil
Cell, 176: 227-238 (2019)

  Recommended for Faculty of 1000 (link)
  Commentary in Nature Reviews Genetics by Michelle Trenkmann (PDF); in Nature Methods by Nicole Rusk (PDF); in Biochemistry by Theresa Loveless & Chang Liu (PDF); in Molecular Cell by Timothy Chang, James Kuo & Pamela Silver (PDF); in Nature Chemical Biology by Yiyun Song (link)
Modeling the Impact of Drug Interactions on Therapeutic Selectivity
Zohar B. Weinstein, Nurdan Kuru, Szilvia Kiriakov, Adam C. Palmer, Ahmad S. Khalil, Paul A. Clemons, Muhammad H. Zaman, Frederick P. Roth and Murat Cokol
Nature Communications, 9: 3452 (2018)
Precise, Automated Control of Conditions for High-Throughput Growth of Yeast and Bacteria with eVOLVER
Brandon G. Wong*, Christopher P. Mancuso*, Szilvia Kiriakov, Caleb J. Bashor and Ahmad S. Khalil
Nature Biotechnology, 36: 614-623 (2018)

  Commentary in Nature by Mike May (PDF)
Hsf1 Phosphorylation Generates Cell-to-Cell Variation in Hsp90 Levels and Promotes Phenotypic Plasticity
Xu Zheng*, Ali Beyzavi*, Joanna Krakowiak, Nikit Patel, Ahmad S. Khalil and David Pincus
Cell Reports, 22: 3099-3106 (2018)
Hsf1 and Hsp70 Constitute a Two-Component Feedback Loop that Regulates the Yeast Heat Shock Response
Joanna Krakowiak*, Xu Zheng*, Nikit Patel*, Zoe A. Feder, Jayamani Anandhakumar, Kendra Valerius, David S. Gross, Ahmad S. Khalil and David Pincus
eLife, 7: e31668 (2018)
Uncovering and Engineering the Protein Aggregome
Gregory A. Newby*, Erinc Hallacli*, Szilvia Kiriakov* and Ahmad S. Khalil
In: Principles of Systems Biology, No. 23
Cell Systems, 5: 428-430 (2017)
A Genetic Tool to Track Protein Aggregates and Control Prion Inheritance
Gregory A. Newby*, Szilvia Kiriakov*, Erinc Hallacli*, Can Kayatekin, Peter Tsvetkov, Christopher P. Mancuso, J. Maeve Bonner, William R. Hesse, Sohini Chakrabortee, Anita L. Manogaran, Susan W. Liebman, Susan Lindquist and Ahmad S. Khalil
Cell, 171: 966-979 (2017)

  Recommended for Faculty of 1000 (link); highlighted in The Scientist by Ruth Williams (link)
Prospective Isolation of NKX2-1–Expressing Human Lung Progenitors Derived From Pluripotent Stem Cells
Finn Hawkins, Philipp Kramer, Anjali Jacob, Ian Driver, Dylan C. Thomas, Katherine B. McCauley, Nicholas Skvir, Ana M. Crane, Anita A. Kurmann, Anthony N. Hollenberg, Sinead Nguyen, Brandon G. Wong, Ahmad S. Khalil, Sarah X.L. Huang, Susan Guttentag, Jason R. Rock, John M. Shannon, Brian R. Davis and Darrell N. Kotton
The Journal of Clinical Investigation, 127: 2277-2294 (2016)
Heat Shock Signaling, Fast and Slow
David Pincus and Ahmad S. Khalil
In: Principles of Systems Biology, No. 12
Cell Systems, 3: 504-506 (2016)
Dynamic Control of Hsf1 During Heat Shock By a Chaperone Switch and Phosphorylation
Xu Zheng, Joanna Krakowiak, Nikit Patel, Ali Beyzavi, Jideofor Ezike, Ahmad S. Khalil and David Pincus
eLife, 5: e18638 (2016)

  Commentary in eLife by Laura Le Breton & Matthias P. Mayer (PDF)
The Epigenome: The Next Substrate for Engineering
Minhee Park, Albert J. Keung and Ahmad S. Khalil
Genome Biology, 17: 183 (2016)
Cellular Advantages to Signaling in a Digital World (Preview)
Christopher P. Mancuso, Szilvia Kiriakov and Ahmad S. Khalil
Cell Systems, 3: 114-115 (2016)
Biological Insights From Synthetic Biology (Editorial)
Jordi Garcia-Ojalvo, Ahmad S. Khalil and John McCarthy
Integrative Biology, 8: 380-382 (2016)
A Unifying Model of Epigenetic Regulation (Perspective)
Albert J. Keung and Ahmad S. Khalil
Science, 351: 661-662 (2016)
Antibiotic Efficacy is Linked to Bacterial Cellular Respiration
Michael A. Lobritz, Peter Belenky, Caroline B.M. Porter, Arnaud Gutierrez, Jason H. Yang, Eric G. Schwarz, Daniel J. Dwyer, Ahmad S. Khalil and James J. Collins
Proc. Natl. Acad. Sci. USA, 112: 8173-8180 (2015)
Chromatin Regulation at the Frontier of Synthetic Biology
Albert J. Keung, J. Keith Joung, Ahmad S. Khalil and James J. Collins
Nature Reviews Genetics, 16: 159-171 (2015)
Using Targeted Chromatin Regulators to Engineer Combinatorial and Spatial Transcriptional Regulation
Albert J. Keung, Caleb J. Bashor, Szilvia Kiriakov, James J. Collins and Ahmad S. Khalil
Cell, 158: 110-120 (2014)

  Commentary in Nature Reviews Genetics by Darren J. Burgess (PDF)
Antibiotics Induce Redox-Related Physiological Alterations as Part of Their Lethality
Daniel J. Dwyer, Peter A. Belenky, Jason H. Yang, I. Cody MacDonald, Jeffrey D. Martell, Noriko Takahashi, Clement T. Y. Chan, Michael A. Lobritz, Dana Braff, Eric G. Schwarz, Jonathan D. Ye, Mekhala Pati, Maarten Vercruysse, Paul S. Ralifo, Kyle R. Allison, Ahmad S. Khalil, Alice Y. Ting, Graham C. Walker and James J. Collins
Proc. Natl. Acad. Sci. USA, 111: E2100-E2109 (2014)
Engineering Life
Ahmad S. Khalil, Caleb J. Bashor, and Timothy K. Lu
The Scientist, August 1, 2013

Iterative Plug-and-Play Methodology for Constructing and Modifying Synthetic Gene Networks
Kevin D. Litcofsky, Raffi B. Afeyan, Russell J. Krom, Ahmad S. Khalil and James J. Collins
Nature Methods, 9: 1077-80 (2012)

  Commentary in Nature Methods by Jeffrey J. Tabor (PDF)
A Synthetic Biology Framework for Programming Eukaryotic Transcription Functions
Ahmad S. Khalil*, Timothy K. Lu*, Caleb J. Bashor*, Cherie L. Ramirez, Nora C. Pyenson, J. Keith Joung and James J. Collins
Cell, 150: 647-658 (2012)

  Commentary in Nature Biotechnology by Susan Jones (PDF); in Nature Methods (PDF)
Signaling-Mediated Bacterial Persister Formation
Nicole M. Vega, Kyle R. Allison, Ahmad S. Khalil and James J. Collins
Nature Chemical Biology, 8: 431-433 (2012)

  Commentary in Nature (PDF); in Science (PDF); in Nature Medicine (PDF); in Nat. Rev. Microbiol. by Andrew Jermy (PDF)
Synthetic Biology: Applications Come of Age
Ahmad S. Khalil and James J. Collins
Nature Reviews Genetics, 11: 367-370 (2010)

  Highlighted in The New York Times by Olivia Judson (link); in Folha de S. Paulo (PDF)
Functionalized Endothelialized Microvascular Networks with Circular Cross Sections in a Tissue Culture Substrate
Jeffrey T. Borenstein, Malinda M. Tupper, Peter J. Mack, Eli J. Weinberg, Ahmad S. Khalil, James Hsiao and Guillermo Garcia-Cardena
Biomedical Microdevices, 12: 71-79 (2010)
Next-Generation Synthetic Gene Networks
Timothy K. Lu, Ahmad S. Khalil and James J. Collins
Nature Biotechnology, 27: 1139-1150 (2009)
Kinesin's Cover-Neck Bundle Folds Forward to Generate Force
Ahmad S Khalil, David C Appleyard, Anna K. Labno, Adrien Georges, Martin Karplus, Angela M. Belcher, Wonmuk Hwang and Matthew J. Lang
Proc. Natl. Acad. Sci. USA, 105: 19247-19252 (2008)

Single M13 Bacteriophage Tethering and Stretching
Ahmad S. Khalil, Jorge M. Ferrer, Ricardo R. Brau, Stephen T. Kottmann, Christopher J. Noren, Matthew J. Lang and Angela M. Belcher
Proc. Natl. Acad. Sci. USA, 104: 4892-4897 (2007)

A Combined FEM/Genetic Algorithm for Vascular Soft Tissue Elasticity Estimation
Ahmad S. Khalil, Brett E. Bouma and Mohammad R. Kaazempur-Mofrad
Cardiovascular Engineering, 6: 93-102 (2006)
Tissue Elasticity Estimation with Optical Coherence Elastography: Toward Complete Mechanical Characterization of In Vivo Soft Tissue
Ahmad S. Khalil, Raymond C. Chan, Alexandra H. Chau, Brett E. Bouma and Mohammad R. Kaazempur-Mofrad
Annals of Biomedical Engineering, 33: 1631-1639 (2005)
OCT-Based Arterial Elastography: Robust Estimation Exploiting Tissue Biomechanics
Raymond C. Chan, Alexandra H. Chau, W. Clem Karl, Seemantini Nadkarni, Ahmad S. Khalil, Nicusor Iftimia, Milen Shishkov, Guillermo J. Tearney, Mohammad R. Kaazempur-Mofrad and Brett E. Bouma
Optics Express, 12: 4532-4572 (2004)

Synthetic Reconstitution of Complex Cellular Behavior

Can we build biological systems that recapitulate complex cellular functions like those seen in nature? Answering this question is the central goal of our research.

This question also forms the basis of reconstitution, an established experimental approach that reimagines how a biological process can be recapitulated outside of its natural context (e.g. outside of the cell and in a test tube) using a reduced set of molecular components. Biochemical reconstitution has been successfully applied to recreate many processes, enabling precise control over molecular parameters and a powerful way to test mechanistic models and establish sufficiency. Our vision is to implement the power and precision afforded by biochemical reconstitution within the complex environment of a living cell. If we can achieve this, then we can understand and predictably control complex cellular functions that have eluded our understanding, such as those that regulate how cells make decisions, establish memories, actuate responses, and develop into multicellular organisms.

To do this, our laboratory is developing novel tools at the intersection of synthetic & systems biology, protein & cell engineering, laboratory evolution, genomics, and computation that enable us to recapitulate and control cellular behavior with synthetic circuitry. This enables us to effectively replace biochemistry with genetics. Taking this leap forward is fundamentally important for basic biology, to discover how cellular behaviors and diseases arise from complex networks of interacting molecules. It is also important for engineering and medicine, offering the potential to precisely control cellular function for next-generation therapies and to “teach” cells and organisms to solve the greatest health, climate, and engineering challenges of today.

Specifically, our laboratory develops tools of synthetic biology that allow us to construct regulatory circuits inside living cells. We are using our tools to dissect the molecular circuits that control gene regulation in eukaryotes, toward addressing the grand challenge of understanding their organization across scale and how they function to generate diverse cellular phenotypes. The basic insights we generate inform the development of platforms to program therapeutically-useful cellular functions for emerging gene and cell-based therapies, such as CAR-T cells for cancer. In addition, our team is developing novel continuous evolution technologies that are automated and scalable, and applying these to generate biomolecules with radically altered or new functions to address unmet needs in biology, medicine, and biotechnology. To broaden the impact of our basic science and medical discoveries, we make the technologies we develop widely usable and accessible to the scientific community. Overall, by learning how to build biological systems from scratch, our broad goal is to connect the molecular building blocks of life to a comprehensive understanding of cellular behavior and ultimately to clinical and other applications.

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