CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer
Summary: Dissecting mechanisms driving subclone expansion in primary cancers has been challenging. Here, we present a protocol to systematically disrupt entire gene networks and assess the functional impact of this perturbation on cancer cell fitness. By combining arrayed CRISPR libraries and high-c...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Elsevier
2023-03-01
|
| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166723000552 |
| _version_ | 1811165440301334528 |
|---|---|
| author | Fabrizio Simeoni Ioannis Loukas Thomas Stuart Wilson Paola Scaffidi |
| author_facet | Fabrizio Simeoni Ioannis Loukas Thomas Stuart Wilson Paola Scaffidi |
| author_sort | Fabrizio Simeoni |
| collection | DOAJ |
| description | Summary: Dissecting mechanisms driving subclone expansion in primary cancers has been challenging. Here, we present a protocol to systematically disrupt entire gene networks and assess the functional impact of this perturbation on cancer cell fitness. By combining arrayed CRISPR libraries and high-content microscopy, we describe steps to identify classes of genes whose inactivation promotes resistance to environmental challenges faced by cancer cells during tumor growth or upon therapy. A proof-of-principle interrogation of the epigenetic regulatory network is described.For complete details on the use and execution of this protocol, please refer to Loukas et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
| first_indexed | 2024-04-10T15:37:11Z |
| format | Article |
| id | doaj.art-e853ea83f60c4e0b99a1d782f32274a8 |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-04-10T15:37:11Z |
| publishDate | 2023-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-e853ea83f60c4e0b99a1d782f32274a82023-02-13T04:07:36ZengElsevierSTAR Protocols2666-16672023-03-0141102097CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancerFabrizio Simeoni0Ioannis Loukas1Thomas Stuart Wilson2Paola Scaffidi3Cancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UK; Corresponding authorCancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UKCancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UKCancer Epigenetics Laboratory, The Francis Crick Institute, NW1 1AT London, UK; Department of Experimental Oncology, IEO, European Institute of Oncology IRCCS, 20139 Milan, Italy; Corresponding authorSummary: Dissecting mechanisms driving subclone expansion in primary cancers has been challenging. Here, we present a protocol to systematically disrupt entire gene networks and assess the functional impact of this perturbation on cancer cell fitness. By combining arrayed CRISPR libraries and high-content microscopy, we describe steps to identify classes of genes whose inactivation promotes resistance to environmental challenges faced by cancer cells during tumor growth or upon therapy. A proof-of-principle interrogation of the epigenetic regulatory network is described.For complete details on the use and execution of this protocol, please refer to Loukas et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.http://www.sciencedirect.com/science/article/pii/S2666166723000552Cell BiologyCell-based AssaysCancerGeneticsHigh-throughput ScreeningMolecular Biology |
| spellingShingle | Fabrizio Simeoni Ioannis Loukas Thomas Stuart Wilson Paola Scaffidi CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer STAR Protocols Cell Biology Cell-based Assays Cancer Genetics High-throughput Screening Molecular Biology |
| title | CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer |
| title_full | CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer |
| title_fullStr | CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer |
| title_full_unstemmed | CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer |
| title_short | CRISPR-based large-scale modeling of loss-of-function mutations to investigate mechanisms of stress resistance in cancer |
| title_sort | crispr based large scale modeling of loss of function mutations to investigate mechanisms of stress resistance in cancer |
| topic | Cell Biology Cell-based Assays Cancer Genetics High-throughput Screening Molecular Biology |
| url | http://www.sciencedirect.com/science/article/pii/S2666166723000552 |
| work_keys_str_mv | AT fabriziosimeoni crisprbasedlargescalemodelingoflossoffunctionmutationstoinvestigatemechanismsofstressresistanceincancer AT ioannisloukas crisprbasedlargescalemodelingoflossoffunctionmutationstoinvestigatemechanismsofstressresistanceincancer AT thomasstuartwilson crisprbasedlargescalemodelingoflossoffunctionmutationstoinvestigatemechanismsofstressresistanceincancer AT paolascaffidi crisprbasedlargescalemodelingoflossoffunctionmutationstoinvestigatemechanismsofstressresistanceincancer |