The rapid advances in human genome sequencing has generated a large amount of data and consequently a large number of hypotheses. For psychiatric disease, a number of landmark studies have led to lists of genetic variations that are linked to specific diseases. However, functional testing of these hypotheses has remained challenging due to a lack of appropriate model systems and high-throughput functional assays. Here we propose to develop and apply a series of high-throughput genome perturbation methodologies to enable the rapid identification of causal genetic variants from large-scale genetics data. The technology we propose to develop will be very broadly applicable and has the potential to radically transform the scale and rate of discovery across different biomedical fields. Three complementary technologies will be developed as a part of this proposal: 1. Functional screening methods to enable rapid identification of reprogramming factors for specific cell types. By using genome-scale transcription activation or epigenetic reprogramming libraries, we will establish a systematic approach to reverse-engineer the sufficient combinations of reprogramming factors for neuron subtypes that are relevant for different neuropsychiatric diseases, such as parvalbumin positive interneurons in schizophrenia. 2. Large-scale and systematic genome perturbation tools to enable massively-parallel functional testing of genetic variants, to enable narrowing of the long lists of correlated genetic variants to a short list of causal variants. We will establish genetic perturbation technologies to enable multiplex and systematic deletion or mutational analysis of coding and non-coding genomic regions, as well as probing of their epigenetic states, to identify causal genetic variants. 3. Efficient and precise genome modification technologies to enable rapid introduction of causal variants into cellular or animal models to enable elucidation of their disease-causing mechanisms. Whereas nuclease-based genome editing systems enables efficient gene knockout, gene insertion or gene correction remains inefficient. We will characterize a novel programmed genome rearrangement mechanism and develop it into a technology for efficient and precise mammalian genome editing. This will significantly accelerate the generation of cellular and animal models Given the broad applicability of this technology, the impact of this proposed work will be far reaching and will radically transform existing experimental approaches for studying gene interactions in all fields of life science and medicine.

Public Health Relevance

A major challenge facing the study of the genetic underpinning of psychiatric diseases is the lack of appropriate high-throughput models and functional testing methods to identify disease-causing genetic variants. Here we propose ways to develop high-throughput assays that will enable the derivation of specific neuronal subtypes and rapid and systematic functional genetic perturbations in cellular and animal models. The combination of technologies proposed here will establish a new and much-needed approach for the functional study of the genetic determinants of psychiatric disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Research Project (R01)
Project #
1R01MH110049-01
Application #
8948623
Study Section
Special Emphasis Panel (ZRG1-BCMB-A (51))
Program Officer
Beckel-Mitchener, Andrea C
Project Start
2015-09-23
Project End
2020-06-30
Budget Start
2015-09-23
Budget End
2016-06-30
Support Year
1
Fiscal Year
2015
Total Cost
$1,186,474
Indirect Cost
$480,681
Name
Broad Institute, Inc.
Department
Type
DUNS #
623544785
City
Cambridge
State
MA
Country
United States
Zip Code
02142
Nishimasu, Hiroshi; Shi, Xi; Ishiguro, Soh et al. (2018) Engineered CRISPR-Cas9 nuclease with expanded targeting space. Science 361:1259-1262
Myhrvold, Cameron; Freije, Catherine A; Gootenberg, Jonathan S et al. (2018) Field-deployable viral diagnostics using CRISPR-Cas13. Science 360:444-448
Gootenberg, Jonathan S; Abudayyeh, Omar O; Kellner, Max J et al. (2018) Multiplexed and portable nucleic acid detection platform with Cas13, Cas12a, and Csm6. Science 360:439-444
Tekin, Halil; Simmons, Sean; Cummings, Beryl et al. (2018) Effects of 3D culturing conditions on the transcriptomic profile of stem-cell-derived neurons. Nat Biomed Eng 2:540-554
Zetsche, Bernd; Heidenreich, Matthias; Mohanraju, Prarthana et al. (2017) Multiplex gene editing by CRISPR-Cpf1 using a single crRNA array. Nat Biotechnol 35:31-34
Meier, Joshua A; Zhang, Feng; Sanjana, Neville E (2017) GUIDES: sgRNA design for loss-of-function screens. Nat Methods 14:831-832
Joung, Julia; Engreitz, Jesse M; Konermann, Silvana et al. (2017) Genome-scale activation screen identifies a lncRNA locus regulating a gene neighbourhood. Nature 548:343-346
Cox, David B T; Gootenberg, Jonathan S; Abudayyeh, Omar O et al. (2017) RNA editing with CRISPR-Cas13. Science 358:1019-1027
Chow, Ryan D; Guzman, Christopher D; Wang, Guangchuan et al. (2017) AAV-mediated direct in vivo CRISPR screen identifies functional suppressors in glioblastoma. Nat Neurosci 20:1329-1341
Gootenberg, Jonathan S; Abudayyeh, Omar O; Lee, Jeong Wook et al. (2017) Nucleic acid detection with CRISPR-Cas13a/C2c2. Science 356:438-442

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