4 research outputs found
Discovery and Functional Characterization of Diverse Class 2 CRISPR-Cas Systems
Microbial CRISPR-Cas systems are divided into Class 1, with multisubunit effector complexes, and Class 2, with single protein effectors. Currently, only two Class 2 effectors, Cas9 and Cpf1, are known. We describe here three distinct Class 2 CRISPR-Cas systems. The effectors of two of the identified systems, C2c1 and C2c3, contain RuvC-like endonuclease domains distantly related to Cpf1. The third system, C2c2, contains an effector with two predicted HEPN RNase domains. Whereas production of mature CRISPR RNA (crRNA) by C2c1 depends on tracrRNA, C2c2 crRNA maturation is tracrRNA independent. We found that C2c1 systems can mediate DNA interference in a 5'-PAM-dependent fashion analogous to Cpf1. However, unlike Cpf1, which is a single-RNA-guided nuclease, C2c1 depends on both crRNA and tracrRNA for DNA cleavage. Finally, comparative analysis indicates that Class 2 CRISPR-Cas systems evolved on multiple occasions through recombination of Class 1 adaptation modules with effector proteins acquired from distinct mobile elements.National Institute of Mental Health (U.S.) (Grant 5DP1-MH100706)National Institute of Mental Health (U.S.) (Grant 1R01-MH110049)National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (Grant 5R01DK097768-03)National Institutes of Health (U.S.) (Grant GM10407
Mol Cell
Microbial CRISPR-Cas systems are divided into Class 1, with multisubunit effector complexes, and Class 2, with single protein effectors. Currently, only two Class 2 effectors, Cas9 and Cpf1, are known. We describe here three distinct Class 2 CRISPR-Cas systems. The effectors of two of the identified systems, C2c1 and C2c3, contain RuvC-like endonuclease domains distantly related to Cpf1. The third system, C2c2, contains an effector with two predicted HEPN RNase domains. Whereas production of mature CRISPR RNA (crRNA) by C2c1 depends on tracrRNA, C2c2 crRNA maturation is tracrRNA independent. We found that C2c1 systems can mediate DNA interference in a 5'-PAM-dependent fashion analogous to Cpf1. However, unlike Cpf1, which is a single-RNA-guided nuclease, C2c1 depends on both crRNA and tracrRNA for DNA cleavage. Finally, comparative analysis indicates that Class 2 CRISPR-Cas systems evolved on multiple occasions through recombination of Class 1 adaptation modules with effector proteins acquired from distinct mobile elements.R01 MH110049/MH/NIMH NIH HHS/United States5DP1-MH100706/DP/NCCDPHP CDC HHS/United StatesR01 GM104071/GM/NIGMS NIH HHS/United StatesDP1 MH100706/MH/NIMH NIH HHS/United StatesR01 DK097768/DK/NIDDK NIH HHS/United States5R01DK097768-03/DK/NIDDK NIH HHS/United StatesGM10407/GM/NIGMS NIH HHS/United States2016-11-05T00:00:00Z26593719PMC466026
The physicist's guide to one of biotechnology's hottest new topics: CRISPR-Cas
Clustered regularly interspaced short palindromic repeats (CRISPR) and
CRISPR-associated proteins (Cas) constitute a multi-functional, constantly
evolving immune system in bacteria and archaea cells. A heritable, molecular
memory is generated of phage, plasmids, or other mobile genetic elements that
attempt to attack the cell. This memory is used to recognize and interfere with
subsequent invasions from the same genetic elements. This versatile prokaryotic
tool has also been used to advance applications in biotechnology. Here we
review a large body of CRISPR-Cas research to explore themes of evolution and
selection, population dynamics, horizontal gene transfer, specific and
cross-reactive interactions, cost and regulation, non-immunological CRISPR
functions that boost host cell robustness, as well as applicable mechanisms for
efficient and specific genetic engineering. We offer future directions that can
be addressed by the physics community. Physical understanding of the CRISPR-Cas
system will advance uses in biotechnology, such as developing cell lines and
animal models, cell labeling and information storage, combatting antibiotic
resistance, and human therapeutics.Comment: 75 pages, 15 figures, Physical Biology (2018