CRISPRRNA maturation by trans-encoded small RNA and host factor RNase III
Elitza Deltcheva, Krzysztof Chylinski, Cynthia M. Sharma, Karine Gonzales, Yanjie Chao,
Zaid A. Pirzada, Maria R. Eckert, Jörg Vogel & Emmanuelle Charpentier
Nature. 2011 Mar 31;471(7340):602-7.
Presenter: Kun-Ta Chung
Date/Time: 2011/10/06 17:10~18:00
Commentator: Ching-Hao Teng Ph.D. Location: Room 601, Med College Building
Background
Clustered regularly interspaced short palindromic repeats (CRISPR) are
a mechanism for bacteria and archaea to against the foreign genetic elements.
Once the foreign genetic element invades into host cell, Cas protein complex
may recognize and cut it into fragment. This fragment would be integrated into
CRISPR array and it would be a CRISPR RNA (crRNA) as a guide when the
same invasion comes next time. The endonuclease is important in crRNA
maturation; however it does not appear in many CRISPR/Cas systems.
Objective
To understand the mechanism of crRNA is maturation in Streptococcus
pyogenes.
Material & Method
The clinical strain SF370 was used in this article. To detect RNA signal,
the probes were designed at lepA, csn2 and repeat region.
Results
The abundant RNA sequences that contain sequence which is similar
with the CRISPR repeat was detected by differential RNA library. These RNA
that locate at the upstream of CRISPR array and are on the opposite strand
were named trans-activating CRISPR RNA (tracrRNA). The pre-crRNA
maturation was inhibited when tracrRNA was deleted, and the tracrRNA
modification was also affected when pre-crRNA deletion. The authors
combined these data and then guest that pre-crRNA and tracrRNA is
co-processed by RNase III. tracrRNA directed pre-crRNA cleavage by RNase
III and protected pre-crRNA against RNase T1 and lead (II) digestion in
footprinting assay. Furthermore, crRNA could not be maturation in Csn1, one
of cas protein, deletion mutant. The CRISPR01 system lost function when
Csn1, RNase III, tracrRNA or pre-crRNA was deleted in vivo.
Conclusion
Pre-crRNA requires tracrRNA, RNase III and Csn1 to be maturation.
Reference
Horvath, P. & Barrangou, R. CRISPR/Cas, the immune system of bacteria and
archaea. Science 327, 167–170 (2010).