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2015 ; 479-480
(ä): 213-20
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Bacterial CRISPR/Cas DNA endonucleases: A revolutionary technology that could
dramatically impact viral research and treatment
#MMPMID25759096
Kennedy EM
; Cullen BR
Virology
2015[May]; 479-480
(ä): 213-20
PMID25759096
show ga
CRISPR/Cas systems mediate bacterial adaptive immune responses that evolved to
protect bacteria from bacteriophage and other horizontally transmitted genetic
elements. Several CRISPR/Cas systems exist but the simplest variant, referred to
as Type II, has a single effector DNA endonuclease, called Cas9, which is guided
to its viral DNA target by two small RNAs, the crRNA and the tracrRNA. Initial
efforts to adapt the CRISPR/Cas system for DNA editing in mammalian cells, which
focused on the Cas9 protein from Streptococcus pyogenes (Spy), demonstrated that
Spy Cas9 can be directed to DNA targets in mammalian cells by tracrRNA:crRNA
fusion transcripts called single guide RNAs (sgRNA). Upon binding, Cas9 induces
DNA cleavage leading to mutagenesis as a result of error prone non-homologous end
joining (NHEJ). Recently, the Spy Cas9 system has been adapted for high
throughput screening of genes in human cells for their relevance to a particular
phenotype and, more generally, for the targeted inactivation of specific genes,
in cell lines and in vivo in a number of model organisms. The latter aim seems
likely to be greatly enhanced by the recent development of Cas9 proteins from
bacterial species such as Neisseria meningitidis and Staphyloccus aureus that are
small enough to be expressed using adeno-associated (AAV)-based vectors that can
be readily prepared at very high titers. The evolving Cas9-based DNA editing
systems therefore appear likely to not only impact virology by allowing
researchers to screen for human genes that affect the replication of pathogenic
human viruses of all types but also to derive clonal human cell lines that lack
individual gene products that either facilitate or restrict viral replication.
Moreover, high titer AAV-based vectors offer the possibility of directly
targeting DNA viruses that infect discrete sites in the human body, such as
herpes simplex virus and hepatitis B virus, with the hope that the entire
population of viral DNA genomes might be destroyed. In conclusion, we believe
that the continued rapid evolution of CRISPR/Cas technology will soon have a
major, possibly revolutionary, impact on the field of virology.
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