Title of Presentation

“The transformative CRISPR-Cas9 genome engineering technology: lessons learned from bacteria”

The RNA-programmable CRISPR-Cas9 system has recently emerged as a transformative technology in biological sciences, allowing rapid and efficient targeted genome editing, chromosomal marking and gene regulation. In this system, the endonuclease Cas9 or catalytically inactive Cas9 variants are programmed with single guide RNAs (sgRNAs) to target site-specifically any DNA sequence of interest given the presence of a short sequence (Protospacer Adjacent Motif, PAM) juxtaposed to the complementary region between the sgRNA and target DNA. The system is efficient, versatile and easily programmable.

Originally, CRISPR-Cas is an RNA-mediated adaptive immune system that protects bacteria and archaea from invading mobile genetic elements (phages, plasmids). Short crRNA (CRISPR RNA) molecules containing unique genome-targeting spacers commonly guide Cas protein(s) to invading cognate nucleic acids to affect their maintenance. CRISPR-Cas has been classified into three main types and further subtypes. CRISPR-Cas9 originates from the type II CRISPR-Cas system that has evolved unique molecular mechanisms for maturation of crRNAs and targeting of invading DNA, which my laboratory has identified in the human pathogen Streptococcus pyogenes. During the step of crRNA biogenesis, a unique CRISPR-associated RNA, tracrRNA, base pairs with the repeats of precursor-crRNA to form anti-repeat-repeat dual-RNAs that are cleaved by RNase III in the presence of Cas9 (formerly Csn1), generating mature tracrRNA and intermediate forms of crRNAs. Following a second maturation event, the mature dual-tracrRNA-crRNAs guide the endonuclease Cas9 to cleave cognate target DNA and thereby affect the maintenance of invading genomes. We have shown that the endonuclease Cas9 can be programmed with sgRNAs mimicking the natural dual-tracrRNA-crRNAs to target site-specifically any DNA sequence of interest. Based on this harnessing principle, we proposed that RNA-programmable Cas9 could be useful as a versatile system for genome editing in cells of all three kingdoms of life for biotechnological, biomedical and gene-therapeutic purposes. As demonstrated by a large number of studies published over the last 2 years, DNA targeting by CRISPR-Cas9 has been quickly and broadly adopted by the scientific community to edit and silence genomes in a large variety of cells and organisms, including human cells, plants and mice. I will discuss the biological roles of CRISPR-Cas9, the mechanisms involved, the evolution of type II CRISPR-Cas components in bacteria and the applications of CRISPR-Cas9 as a novel genome engineering technology.

Profile

Web Site URL

http://www.mpiib-berlin.mpg.de/research/regulation_in_infection_biology

A brief Biography

Emmanuelle Charpentier studied biochemistry, microbiology and genetics at the University Pierre and Marie Curie, Paris, France and obtained her PhD in Microbiology for her research performed at the Pasteur Institute. She then continued her work in the United States, at The Rockefeller University, New York University Langone Medical Center and the Skirball Institute of Biomolecular Medicine (all in New York, NY) and at St Jude Children’s Research Hospital (in Memphis, TN). E. Charpentier returned to Europe to establish her own research group at the Max F. Perutz Laboratories of the University of Vienna in Austria where she habilitated in the field of Microbiology. She was then appointed Associate Professor at the Laboratory for Molecular Infection Medicine Sweden (MIMS, part of Nordic European Molecular Biology Laboratory (EMBL) Partnership for Molecular Medicine) at Umeå University in Sweden where she habilitated in the field of Medical Microbiology and is still active as a Visiting Professor. Between 2013 and 2015, E. Charpentier was Head of the Department of Regulation in Infection Biology at the Helmholtz Centre for Infection Research, Braunschweig, and Professor at the Medical School of Hannover in Germany. In 2013, she was awarded an Alexander von Humboldt Professorship, which she has held since 2014. In 2015, E. Charpentier was appointed Scientific Member of the Max Planck Society in Germany and Director at the Max Planck Institute for Infection Biology in Berlin, Germany.

E. Charpentier is recognized as a world-leading expert in regulatory mechanisms underlying processes of infection and immunity in bacterial pathogens. Her work has led to a number of seminal discoveries and insights into pathways governing antibiotic resistance and virulence of bacterial pathogens. With her recent groundbreaking findings in the field of RNA-mediated regulation based on the CRISPR-Cas9 system, E. Charpentier has laid the foundation for the development of a novel, highly versatile and specific genome editing technology that is revolutionizing life sciences research and could open up whole new opportunities in biomedical gene therapies. The resulting field of research is now developing at dazzling speed, with exciting new aspects emerging almost weekly. For this research, E. Charpentier has been awarded a number of prestigious honors.

E. Charpentier is inventor and co-owner of seminal intellectual property comprising the CRISPR-Cas9 technology, and co-founder of CRISPR Therapeutics and ERS Genomics, created to facilitate the development of the CRISPR-Cas9 genome engineering technology for biotechnological and biomedical purposes.

Details of selected Awards and Honors

Leibniz Prize 2016

Otto Warburg Medal 2016

Carus-Medal of the German National Academy of Sciences Leopoldina 2015

Gruber Prize in Genetics 2015

Princess of Asturias Award for Technical and Scientific Research 2015

Louis Jeantet Prize for Medicine 2015

Ernst Jung Prize for Medicine 2015

Breakthrough Prize in Life Sciences 2015

Grand Prix Jean-Pierre LeCocq of the French Academy of Sciences 2014

Göran Gustafsson Prize of the Royal Swedish Academy of Sciences 2014

A list of selected Publications

Fonfara I, Richter H, Bratovic M. Le Rhun A and Charpentier E. 2016. The CRISPR-associated DNA-cleaving enzyme Cpf1 also processes precursor CRISPR RNA. Nature. In Press.

Charpentier E, Richter H, van der Oost J and White MF. 2015. Biogenesis pathways of RNA guides in archaeal and bacterial CRISPR-Cas adaptive immunity. FEMS Microbiol. Rev. 39:428-441.

Plagens A, Richter H, Charpentier E and Randau L. 2015. DNA and RNA interference mechanisms by CRISPR-Cas surveillance complexes. FEMS Microbiol. Rev. 39:442-463.

Charpentier E. 2015. CRISPR-Cas9: how research on a bacterial RNA-guided mechanism opened new perspectives in biotechnology and biomedicine. EMBO Mol. Med. 7:363-365.

Chylinski K, Makarova KS, Charpentier E and Koonin EV. 2014. Classification and evolution of type II CRISPR-Cas systems. Nucleic Acids Res. 42:6091-105.

Jinek M, Jiang F, Taylor DW, Stenberg SH, Kaya E, Ma E, Anders C, Hauer M, Zhou K, Lin S, Kaplan M, Iavarone AT, Charpentier E, Nogales E and Doudna JA. 2014. Structures of Cas9 endonucleases reveal RNA-mediated conformational activation. Science 343:1247997.

Fonfara I, Le Rhun A, Chylinski K, Makarova KS, Lécrivain AL, Bzdrenga J, Koonin EV and Charpentier E. 2014. Phylogeny of Cas9 determines functional exchangeability of dual-RNA and Cas9 among orthologous type II CRISPR-Cas systems. Nucleic Acids Res. 42:2577-2590.

Chylinski K, Le Rhun A and Charpentier E. 2013. The tracrRNA and Cas9 families of type II CRISPR-Cas immunity systems. RNA Biol. 10:726-737.

Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA and Charpentier E. 2012. A programmable dual-RNA guided DNA endonuclease in adaptive bacterial immunity. Science 337:816-821.

Deltcheva E, Chylinski K, Sharma C, Gonzales K, Chao Y, Pirzada ZA, Eckert M, Vogel J and Charpentier E. 2011. CRISPR RNA maturation by trans-encoded small RNA and host factor RNAse III. Nature 471:602-607.