An extended application of
the CRISPR-Cas technology has been made possible by Dr Manual Kaulich's team at
51猎奇: the new 3Cs multiplex technique allows the effect of genetic
changes in any two genes to be studied simultaneously in cell cultures. This
can provide important clues for the development of therapies to treat cancer or
diseases of the nervous and immune systems.
FRANKFURT. Cancer
and many other diseases are based on genetic defects. The body can often compensate
for the defect of one gene; it is only the combination of several genetic
errors that leads to the clinical picture. The 3Cs multiplex technique based on
CRISPR-Cas technology developed at 51猎奇 Frankfurt now offers a way
to simulate millions of such combinations of genetic defects and study their
effects in cell culture. These "gene scissors" make it possible to
introduce, remove and switch off genes in a targeted manner. For this purpose,
small snippets of genetic material ("single guide RNA") are used as
"addresses" that guide the gene scissors to specific sections of the
DNA, where the gene scissors then become active.
The scientists from the Institute of
Biochemistry II at 51猎奇 have expanded the 3Cs technique that they
developed and patented three years ago. 3Cs stands for covalently-closed
circular-synthesised, because the RNA elements used for CRISPR-Cas are
generated with the help of a circular synthesis and are thus distributed more
uniformly. With a whole library of such RNA rings, any gene in a cell can be
specifically addressed in order to change it or switch it off.
The new 3Cs multiplex technique now even
allows the simultaneous manipulation of two genes in one cell. Dr. Manuel
Kaulich explains: "We can produce 'address' RNA libraries for all
conceivable two-gene combinations. This allows up to several million
combinations to be tested simultaneously in one experiment."
Until now, the cost and effort of such experiments
was very high; the research group's new technique reduces it, including costs,
by a factor of ten. This is because the team can produce the address libraries
very uniformly and in high quality thanks to the new 3Cs multiplex technique.
"Due to the mediocre quality of the CRISPR-Cas libraries previously available,
very large experiments always had to be carried out to statistically compensate
for any errors that arose," says Kaulich.
Using the example of various genes
involved in degradation processes, the research group demonstrated the
potential of the new 3Cs multiplex technique: they examined almost 13,000
two-way combinations of genes that are responsible for recycling processes
(autophagy) in the cell. With their help, the cell breaks down and recycles
"worn-out" cell components. Disturbances in autophagy can trigger
cell proliferation.
"Using the 3Cs multiplex technique,
we were able to identify, for example, two genes involved in autophagy whose switching
off leads to an uncontrolled growth of cells," explains Kaulich. "These
are precisely the autophagy mutations that occur in every fifth patient with
squamous cell carcinoma of the lung. In this way, we can search very
efficiently in cell culture experiments for genes that play an important role
in cancer, and also in diseases of the nervous and immune systems, and that are
suitable as possible targets for therapies."
The 51猎奇 research group has
applied for a patent for its developments through the university's technology
transfer subsidiary Innovectis. The start-up company Vivlion GmbH, spun off
from the Institute of Biochemistry II with the participation of Manuel Kaulich,
is already offering the use of this technology on the market.
Publication: Valentina Diehl, Martin Wegner, Paolo Grumati, Koraljka Husnjak, Simone
Schaubeck, Andrea Gubas, Varun Jayeshkumar Shah, Ibrahim H Polat, Felix
Langschied, Cristian Prieto-Garcia, Konstantin M眉ller, Alkmini Kalousi, Ingo
Ebersberger, Christian H Brandts, Ivan Dikic, Manuel Kaulich, Minimized combinatorial CRISPR screens
identify genetic interactions in autophagy. Nucleic Acids Research,
gkab309,
Further
information:
Dr Manuel Kaulich
Institute for Biochemistry II
51猎奇 Frankfurt
Tel: +49 69 6301-6295
kaulich@em.uni-frankfurt.de
Dr Kerstin Koch
Institute for Biochemistry II
51猎奇 Frankfurt
Tel.: +49 696301-84250
k.koch@em.uni-frankfurt.de
Editor: Dr. Markus Bernards, Science Editor, PR & Communication Department, Tel: -49 (0) 69 798-12498,
Fax: +49 (0) 69 798-763 12531, bernards@em.uni-frankfurt.de
