Article
Biochemistry & Molecular Biology
Maximilian Billmann, Henry N. Ward, Michael Aregger, Michael Costanzo, Brenda J. Andrews, Charles Boone, Jason Moffat, Chad L. Myers
Summary: CRISPR screens are widely used to investigate the relationship between phenotype and genotype. Recent efforts focus on identifying specific phenotypes that are related to cell lines, genetic backgrounds, or specific conditions of interest. However, accurately measuring the reproducibility of context-specific hits is a challenge, and it is important to develop metrics that are sensitive to context-specific signals to assess screen quality.
Review
Agriculture, Multidisciplinary
Rui-Xiang Zhang, Bin-Bin Li, Zheng-Guang Yang, Jia-Qi Huang, Wei-Hang Sun, Nadeem Bhanbhro, Wen-Ting Liu, Kun-Ming Chen
Summary: This article provides a comprehensive overview of the CRISPR-based gene editing technology in agricultural breeding, highlighting the application and capacity of different toolsets in plant gene expression regulation. It also discusses the general problems in CRISPR application and optimal solutions for high-throughput gene function analysis.
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
(2022)
Article
Biotechnology & Applied Microbiology
Elizabeth R. Hutton, Christopher R. Vakoc, Adam Siepel
Summary: High-throughput CRISPR-Cas9 knockout screens are widely used in cancer research to evaluate gene essentiality. The probabilistic modeling framework ACE can predict absolute and differential essentiality, identify genotype-specific essentiality candidates, and provide a robust framework for identifying genes responsive to subtype-specific therapeutic targeting.
Article
Biotechnology & Applied Microbiology
Yihan Wang, Shiqi Xie, Daniel Armendariz, Gary C. Hon
Summary: This study reveals the presence of clonal cell expansion in single-cell CRISPR screens, where multiplexed sgRNAs are used as barcodes to identify clonal cells. Clonal cells show transcriptional similarities and segmental copy number changes, indicating their genetic distinctness. Clonal cell expansion in single-cell CRISPR screens could lead to false positives, emphasizing the importance of reducing clonal expansion and implementing computational filtering for improved reliability.
Article
Biochemistry & Molecular Biology
Ahyoung Choi, Insu Jang, Heewon Han, Min-Seo Kim, Jinhyuk Choi, Jieun Lee, Sung-Yup Cho, Yukyung Jun, Charles Lee, Jaesang Kim, Byungwook Lee, Sanghyuk Lee
Summary: iCSDB is an integrated database of CRISPR screening experiments using human cell lines, compiling data from DepMap and BioGRID ORCS, covering numerous human cell lines and cancer types. The database also integrates clinical and molecular information, and provides interactive tools and viewers to help users choose and compare screening results.
NUCLEIC ACIDS RESEARCH
(2021)
Article
Biotechnology & Applied Microbiology
Peter C. DeWeirdt, Kendall R. Sanson, Annabel K. Sangree, Mudra Hegde, Ruth E. Hanna, Marissa N. Feeley, Audrey L. Griffith, Teng Teng, Samantha M. Borys, Christine Strand, J. Keith Joung, Benjamin P. Kleinstiver, Xuewen Pan, Alan Huang, John G. Doench
Summary: In this study, an optimized Cas12a toolkit was developed for pooled, combinatorial genetic screens in human cells. By assaying the activity of thousands of guides, refined on-target design rules and comprehensive off-target rules were established. The toolkit was validated by screening for synthetic lethality in cancer cells and showed similar performance to Cas9 in genome-wide dropout screens with greatly reduced library size.
NATURE BIOTECHNOLOGY
(2021)
Article
Biology
Muhammad Salim Hakeemi, Salim Ansari, Matthias Teuscher, Matthias Weisskopf, Daniela Grossmann, Tobias Kessel, Juergen Doenitz, Janna Siemanowski, Xuebin Wan, Dorothea Schultheis, Manfred Frasch, Siegfried Roth, Michael Schoppmeier, Martin Klingler, Gregor Bucher
Summary: By systematically comparing the gene sets required for homologous but divergent developmental processes between fruit flies and beetles, the study found that the differences in these gene sets are greater than previously believed. Therefore, the insights gained from fruit flies may not be as representative for insects or protostomes as previously thought, and further research in complementary model systems is necessary to obtain a comprehensive understanding.
Article
Biochemical Research Methods
Rebecca L. Hutcheson, Mitchell Hayes, Bill Sugden
Summary: The development of genome-wide CRISPR screening has advanced cellular understanding of molecular systems. In this study, we demonstrate the utility of sequentially conducted CRISPR knockout screens using an existing library to explore a biological question in the human genome, followed by examining each gene candidate against a common gene of interest. This approach, called Many vs One CRISPR screening, is made possible by using a modified 7SK promoter instead of the U6 promoter. The insertion of this novel 7SK promoter into the widely used lentiCRISPRv2 backbone is crucial as it eliminates the need for a significant increase in CRISPR library coverage during screening and sequencing, while maintaining the same lentiviral titer, knockout efficiency, and ease of use as the original lentiCRISPRv2 vector.
ACS SYNTHETIC BIOLOGY
(2022)
Article
Biochemical Research Methods
Rebecca L. Hutcheson, Mitchell Hayes, Bill Sugden
Summary: The advent of genome-wide CRISPR screening has advanced the understanding of molecular systems within cells. In this study, we demonstrate the utility of sequential CRISPR knockout screens using an existing library to explore the human genome and examine the relationship between each gene and a common gene of interest.
ACS SYNTHETIC BIOLOGY
(2022)
Article
Multidisciplinary Sciences
Simon Wisnovsky, Leonhard Mockl, Stacy A. Malaker, Kayvon Pedram, Gaelen T. Hess, Nicholas M. Riley, Melissa A. Gray, Benjamin A. H. Smith, Michael C. Bassik, W. E. Moerner, Carolyn R. Bertozzi
Summary: The study identified key genes required for cell-surface presentation of glycan ligands on leukemia cells using a CRISPRi genomic screening strategy. It revealed a selective interaction between Siglec-7 and the mucin-type glycoprotein CD43, with a specific N-terminal glycopeptide region of CD43 containing clusters of disialylated O-glycan tetrasaccharides forming specific Siglec-7 binding motifs. Knockout or blockade of CD43 in leukemia cells relieves Siglec-7-mediated inhibition of immune killing activity.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
(2021)
Review
Genetics & Heredity
Laralynne Przybyla, Luke A. Gilbert
Summary: The latest approaches in CRISPR-based functional genomics screens provide a scalable and multiplexable way to characterize genes and gene regulatory elements encoded by the human genome. These tools promise to reveal mechanisms of gene function and regulation, as well as enable exploration of how genes work together to modulate complex phenotypes.
NATURE REVIEWS GENETICS
(2022)
Review
Immunology
David W. Vredevoogd, Daniel S. Peeper
Summary: Functional genetic screens have identified tumor-intrinsic nodes of immune resistance, revealing various mechanisms by which tumors evade the immune system. However, limitations in capturing tumor heterogeneity have impacted these analyses. This article provides an overview of the nature and sources of heterogeneity that impact tumor-immune interactions and argues that considering tumor heterogeneity is crucial for discovering novel immune evasion mechanisms.
FRONTIERS IN IMMUNOLOGY
(2023)
Review
Biochemistry & Molecular Biology
Nicholas Z. Lue, Brian B. Liau
Summary: A fundamental challenge in biology is understanding the molecular details of protein function. How mutations alter protein activity, regulation, and response to drugs is of critical importance to human health. Recent years have seen the emergence of pooled base editor screens for in situ mutational scanning, which revolutionize the investigation of proteins in their native contexts.
Article
Biochemistry & Molecular Biology
Mahfuzur Rahman, Maximilian Billmann, Michael Costanzo, Michael Aregger, Amy H. Y. Tong, Katherine Chan, Henry N. Ward, Kevin R. Brown, Brenda J. Andrews, Charles Boone, Jason Moffat, Chad L. Myers
Summary: FLEX is a pipeline that establishes reference standards for benchmarking human genome-wide CRISPR screen data, identifying a predominant mitochondria-associated signal within co-essentiality networks. By characterizing functional bias associated with mitochondria genes, it provides insights into interpreting differential hits in CRISPR screening. The utility of the FLEX pipeline for performing robust comparative evaluations of CRISPR screens or methods for processing them is demonstrated.
MOLECULAR SYSTEMS BIOLOGY
(2021)
Article
Multidisciplinary Sciences
Kendall J. Condon, Jose M. Orozco, Charles H. Adelmann, Jessica B. Spinelli, Pim W. van der Helm, Justin M. Roberts, Tenzin Kunchok, David M. Sabatini
Summary: Our study identified a catalog of genes that impact the mTORC1 pathway and clarified the multifaceted ways in which mTORC1 senses mitochondria dysfunction.AMPK and HRI are two of the key kinases that signal mitochondria distress to mTORC1, and their loss renders mTORC1 signaling largely resistant to mitochondria dysfunction induced by various inhibitors.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
(2021)
