6nue

Structural highlights

Function

CAS10_STRTR CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA). The type III-A Csm effector complex binds crRNA and acts as a crRNA-guided RNase, DNase and cyclic oligoadenylate synthase; binding of target RNA cognate to the crRNA is required for all activities. In a heterologous host this Csm effector complex restricts ssRNA phage MS2, suggesting it may target RNA viruses in vivo.^[1] Csm functions as a non-specific ssDNase. Base-pairing between crRNA and target RNA to form a ternary Csm complex activates a ssDNase activity; target RNA cleavage suppresses the ssDNase, a temporal control that prevents uncontrolled DNA degradation. Viral RNA transcripts probably tether the Csm complex to the viral genome, recruiting Cas10 ssDNA activity which is able to degrade DNA in the transcription bubble, spatially controlling the DNase activity.^[2] This subunit has a weak ssDNase activity that is dramatically activated by the ternary Csm effector complex (the crRNA, Cas proteins and a cognate target ssRNA). Target RNA and ssDNA are cleaved simultaneously, although RNase activity (of Csm3) is much faster. RNA cleavage by Csm3 is not required for ssDNase activity as Csm complex with inactive Csm3 still has ssDNase activity; however as the cleaved target RNA products dissociate away ssDNase activity decreases. Self-recognition, with subsequent repression of the ssDNase activity, occurs when the 5' handle of the crRNA bases pairs with the 3' flanking sequence of the target RNA (which would occur if the CRISPR locus were transcribed as an anti-pre-crRNA). This protein has low activity on dsDNA which is not stimulated by the Csm complex.^[3] This subunit is a single-strand-specific deoxyribonuclease (ssDNase) which digests both linear and circular ssDNA; it has both exo- and endonuclease activity.[UniProtKB:B6YWB8] When associated with the ternary Csm effector complex (the crRNA, Cas proteins and a cognate target ssRNA) synthesizes cyclic oligoadenylates (cOA) from ATP, producing cyclic triadenylate (cA3) up to cyclic hexaadenylate (cA6), which is the active cOA. The enzyme is also able to cyclize pppA3 up to pppA6. cOAs are second messengers that induce an antiviral state important for defense against invading nucleic acids. Synthesis of cOA can occur with AMP plus ATP, 2'dATP or 3'dATP (but no other nucleotides), and requires a free 3'-OH ribose moiety.^[4]

References

1. ↑ Tamulaitis G, Kazlauskiene M, Manakova E, Venclovas C, Nwokeoji AO, Dickman MJ, Horvath P, Siksnys V. Programmable RNA shredding by the type III-A CRISPR-Cas system of Streptococcus thermophilus. Mol Cell. 2014 Nov 20;56(4):506-17. doi: 10.1016/j.molcel.2014.09.027. Epub 2014 , Nov 6. PMID:25458845 doi:http://dx.doi.org/10.1016/j.molcel.2014.09.027
2. ↑ Kazlauskiene M, Tamulaitis G, Kostiuk G, Venclovas C, Siksnys V. Spatiotemporal Control of Type III-A CRISPR-Cas Immunity: Coupling DNA Degradation with the Target RNA Recognition. Mol Cell. 2016 Apr 21;62(2):295-306. doi: 10.1016/j.molcel.2016.03.024. PMID:27105119 doi:http://dx.doi.org/10.1016/j.molcel.2016.03.024
3. ↑ Kazlauskiene M, Tamulaitis G, Kostiuk G, Venclovas C, Siksnys V. Spatiotemporal Control of Type III-A CRISPR-Cas Immunity: Coupling DNA Degradation with the Target RNA Recognition. Mol Cell. 2016 Apr 21;62(2):295-306. doi: 10.1016/j.molcel.2016.03.024. PMID:27105119 doi:http://dx.doi.org/10.1016/j.molcel.2016.03.024
4. ↑ Kazlauskiene M, Kostiuk G, Venclovas C, Tamulaitis G, Siksnys V. A cyclic oligonucleotide signaling pathway in type III CRISPR-Cas systems. Science. 2017 Aug 11;357(6351):605-609. doi: 10.1126/science.aao0100. Epub 2017, Jun 29. PMID:28663439 doi:http://dx.doi.org/10.1126/science.aao0100