Genome Project-write (GP-write) is an open international, multi-disciplinary, research project lead by scientific leaders that plans to reduce costs of engineering and testing large genomes in cell lines. GP-write includes whole genome engineering of human cell lines and other organisms with relevance to agriculture and public health.
The main purpose of GP-write is to better understand the human genome and other genomes in order to benefit from that knowledge. Despite large-scale studies to interpret the human genome, such as HapMap, Encyclopedia of DNA Elements (ENCODE) and genome-wide association studies (GWAS), the understanding of the human genome is considered far from complete. Many scientists believe that by building genomes, they will be better understood because building genomes allows genomes to be tested. The manifesto for the synthetic biology field has been borrowed from physicist Richard Feynman who said, “What I cannot create, I cannot understand”.
Potential benefits to humanity which could come out of GP-write projects on human cells include, growing transplantable human organs, engineering cell lines immune to viruses, engineering cancer resistance into therapeutic cell lines, improving productivity and cost of producing vaccines and pharmaceuticals with human cells.
GP-write will enable better engineering of biology, which could have applications to combat a wide range of challenges including energy production and climate change. GP-write is expected to accelerate research and development in areas like production of food and materials. June Medford from Colorado State University has ideas to engineer plants to filter water or detect chemicals. Harris Wang at Columbia University is investigating the manipulation of genomes of the human microbiome which could potentially lead to modified gut microbes that produce essential amino acids, influence metabolism to treat obesity or secrete compounds to treat irritable bowel syndrome.
The Synthetic Yeast Genome project (Yeast 2.0 or Sc2.0) will be the first eukaryotic genome to be synthesized, and as human are eukaryotic as well, it provides groundwork for synthetic human genome projects. Genome writing in yeast could also lead to new uses for yeast, such as production of biofuel or other useful compounds.
Jef Boeke (New York University), George Church (Harvard) and Andrew Hessel (Autodesk Research) pubished a proposal in Science in 2016 to synthesize the entire genomes from scratch. Initially the term the Human Genome Project-Write was used to refer what they consider the next step after the sequencing of the human genome, which they refer to as the Human Genome Project-Read (HGP-write). HGP-write will be a critical core activity within GP-write and will focus on synthesizing human genomes in whole or in part.
A few weeks prior to the Science publication, a meeting that was closed to the press was held in May 2016 by George Church and collaborators along with 130 invited scientists, lawyers, entrepreneurs and government officials which discussed the feasibility and implementation of a project to synthesize large genomes in vitro. The private nature of the meeting was met with some criticism, with Laurie Zoloth of Northwestern University an Drew Endy of Stanford University publishing an article in Cosmos documenting their disapproval.
GP-write is being implemented through an independent non-profit organization called the Center of Excellence for Engineering Biology (CEEB). The non-profit is managing the initial planning and coordination which involves supporting multi-institutional and interdisciplinary research teams working in an integrated fashion. The organization is also engaged in public outreach.
Scientific priorities will be set by a scientific executive committee which will supervise peer-reviewed research projects. The Center will enter into affiliation agreements with major universities and philanthropies participating in GP-write. There will be a training and citizen science outreach component.
- Jef Boeke, Director at the Institute for Systems Genetics at NYU Langone Medica Center
- George Church, Founding Core Faculty at the Wyss Institute at Harvard University
- Andrew Hessel, CEO at Humane Genomics Inc.
- Nancy J. Kelley, President and CEO of Nancy J Kelly & Associates, former founding Executive Director at the New York Genome Center
The IT infrastructure for GP-write is being designed for large-scale international collaboration. An open sourced, fully automated design, test, build platform will be used to share knowledge and pool data.
Ethical, policy and pubic education components of GP-write are being considered by experts in collaboration with the Center. HGP-write projects will be explicitly limited to cell culture and organoids derived from cells. For HGP-write there will be an expanded examination of ethical, legal and social implications.
Funding may come from public, private, philanthropic and academic sectors and may be international. A gift of $250,000 from Autodesk to seed planning and launch of GP-write was made available to Andrew Hessel, a research scientist at Autodesk and one of the GP-write project leaders. As of May 2017 approximately $200 million in funding for GP-write has been made available across multiple institutions. GP-write partners include Autodesk, Nancy J Kelley + Associates, Feinstein Kean Healthcare, WilmerHale and Labcyte.
Labcyte, a biotechnology tools company, has an agreement with the CEEB which includes providing preferred pricing and support to researchers at qualified laboratories affiliated with and contributing to the GP-write mission and goals. Labcyte will also provide selected researchers with early access to certain newly developed products.
Pilot projects are evaluated with the Scientific Executive Committee. GP-write supports scientists submitting pilot projects with letters of recommendation until the project is fully funded. In the future GP-write has the goal of directly financially support pilot projects. Scientists can apply for funding through their university with GP-write providing a letter of recommendation or scientists can partner with the Center of Excellence to apply for funding.
HGP-write has potential applications in growing transplantable human organs, engineering immunity to viruses in cell lines, engineering cancer resistance into therapeutic cell lines and vaccine and pharmaceutical development using human cells and organoids. Sometimes drug companies are forced to halt production when the cells that are used to produce therapeutic proteins get contaminated with a virus.
The main portion of the details of the construction of five out of 16 chromosomes that make up the yeast (Saccharomyces cerevisiae) genome, lead by Jef Boeke and his Sc2.0 team was published in Science in 2017. To build their first chromosome, which is the shortest, took nearly 10 years. It took less than three years to generate the next five chromosomes, which includes the longest one. The blueprints for the chromosomes are written using a computer program and analyzed by biologists and then they are broken into shorter manageable segments. Segments are synthesized in DNA synthesis labs and then they are joined together using a technique developed by Boeke and undergraduates who took his “build-a-genome” course at Johns Hopkins. The new synthetic genomic material is injected into yeast cells and swaps in for the native DNA by homologous recombination.
The principles for Sc2.0 genome design are to maintain a “wild-type” phenotype while introducing genetic flexibility and reducing sources of genomic instability. The genetic code for Sc2.0 chromosomes have TAG stop codons changed to TAA stop codons. There are loxPsym sites as part of the inducible evolution system called SCRaMbLE. Repeat elements are removed as well as many introns. Transfer RNA (tRNA) genes are relocated to a “neochromosome”. Wild-type DNA and synthetic DNA are distinguished by PCRTags, which are short recoded sequences within the open reading frames (ORF) that can be assayed by PCR reactions. DNA sequence base substitutions within ORFs introduce or remove recognition sites for enzymes in order to facilitate assembly of synthetic chromosomes. For Sc2.0 scientists used BioStudio, an open-source design platform developed for eukaryotic genome design. BioStudio coordinates design modifications at both nucleotide and genome scales and enforces version control to track edits systematically.
A community-wide project to produce “ultra-safe” versions of human cells for cell therapy or drug production is underway. The idea is to produce versions of human cells that are recoded so that they resist viruses, radiation, freezing, aging or cancer. Synthetic genome recoding is the process of changing the three-letter DNA sequences called codons which encode amino acids, building blocks of proteins. This is possible because there are multiple codons for the same amino acid and redundant codons can be swapped while preserving vital function. If a redundant codon is eliminated along with the transfer RNA (tRNA) that translates the deleted codon, the cell will not be able to translate DNA sequences with the eliminated codon. The cell will still produce proteins with the same amino acid sequence but viral DNA entering the cell could not be translated since the virus contains the eliminated codon(s). It is estimated that at least 400,000 changes to the genome would be required to make human cells resistant to viruses. The project is lead by Jef Boeke and George Church.
Jeffrey Way and Pamela Silver, both of Harvard, are leading a project proposing to build two regions of the human genome each of about 1 megabase in size as a human artificial chromosome (HAC). The genomic sequences would be assembled in yeast and technology would be developed to move them into human cells. The system would be used to study mechanisms of long-range gene expression and for engineering humans for an optimized antiviral immune response.
Jasper Rine at University of California has a pilot project with one aspect investigating methods for safety and containment of synthetic genomes or prevent synthetic organisms from hybridizing with natural organisms if they were to be used in agriculture. Methods will be developed and investigated that would prevent survival of recombinants from such hybrids. The project would involve scrambling chromosomes and the utilization of balancer chromosomes similar to those used in the breeding of Drosophila (fruit flies) for genetics research but done in yeast. Synthetic yeast genome Sc2.0 technology would be used. Toxin-antidote combinations inserted into chromosomes that cause the cell to die when separated would be another route of investigation.
The other aspect of the project aims to introduce synthetic sequences into Drosophila in order to use polytene chromosomes to understand the principles behind how certain genomic sequences create particular chromatin structures. Polytene chromosomes are found in salivary glands of Drosophila. The chromosomes undergo multiple rounds of replication without cell division resulting in enlarged chromosomes where the DNA remains aligned. Upon staining an imaging, banding patterns correlate to more or less condensed chromatin structure. Polytene chromosomes are an important model system for studying chromatin architecture changes that are related to gene activity.
Liam Holt at University of California, Berkeley, leads a pilot project to use synthetic biology to map the cell signaling pace of the seven core developmental pathways (Hedgehog, Wnt, TGF-beta, receptor tyrosine kinase, Notch, JAK/STAT and nuclear hormone pathways) to differentiation space which is the range of possible cell states in the progression of stem cells to differentiated cells. The goal is to develop a genetic toolbox for precise temporal modulation of the seven pathways using optiogenetic or chemical control, along with a complimentary toolbox of reporters to profile cell phenotypes. It is expected that the tools will be applicable to controlling the use HGP2 (Human Genome Project 2) cell lines in cell therapies, tissue replacement or organ transplants.
Neville Sanjana at the Broad Institute at MIT has a pilot project to develop a pipeline for rapid engineering of disease-specific variants into human cells with high efficiency. Using CRISPR/Cas technology to knock out a gene with non-homologous end-joining repair is much more efficient than making precise single nucleotide changes using homology-directed repair, which has an efficiency of about 100-fold lower. The project aims to develop selection techniques to enrich for homologous recombination repair and develop bioinformatic tools to assist in choosing the best CRISPR system for a particular mutation or edit.
Harris Wang at Columbia University plans to produce human cell lines that have biosynthetic pathways for production of essential amino acids and vitamins that humans must otherwise obtain from diet. One goal of the project is to use the cells to combat malnutritional conditions, with the cells also proposed to have utility in studying biochemistry in mammalian development and relationships between nutrition and aging processes. Another application of biosynthetic pathway enhanced human cell lines is as an alternative to mammalian cell lines for production of biomolecules for research and as therapeutics. The modified prototrophic human cells could potentially be grown with less expensive growth media . Boeke and Wang received $500,00 from Defense Advanced Research Projects Agency (DARPA) for this project.
Todd Kuiken of North Carolina State University and Gigi Gronvall of Johns Hopkins Center for Health and Security are leading a pilot project that aims to gain understanding and anticipate the governance issues around GP-write. The project will include local, state and national oversight regimes for the research stage and for approval of products or applications that come out of GP-write. Since public support is needed to enable and sustain funding, public dialogue and outreach programs will be established that inform and incorporate the views of the public into the research program.
Yasunori Aizawa of the Tokyo Institute of Technology, Japan, has a pilot project intended to screen for essential introns and retroelements in human cells, mouse and fruit fly in a systematic manner. One and then both alleles of each gene will be replaced with synthetic versions that lack introns and upstream and downstream intergenic regions. Transcriptional, epigenetic status will be compared between native and synthetic versions. Functional analysis of the genes and pathways will be performed when alleles are replaced with the synthetic version. To understand any changes that occur, introns and retroelements will be systematically placed back into the synthetic gene to identify which DNA elements are significant. Their technology includes the development are a “scar-free” marker, useful for others in the GP-write community and development of a protocol for CRISPR/Cas9 usage for promoting the native and synthetic swapping. It is expected that knowing which introns and retroelements are dispensable will allow the synthesis of more compact and simple gene structures, reducing cost and labor in future genome synthesis projects.
Matthew Maurano of NYU Langone Health has a pilot project to use synthetic approach to study regulatory genomics. The project aims to address the gap in the understanding of the gene regulatory role of genetic variants that lie in non-coding regions of the human genome.
A Grand Challenge for Genome Engineering: GP-write | Columbia University Department of Systems Biology
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