Structural insights into the inactivation of CRISPR-Cas systems by diverse anti-CRISPR proteins

Abstract

A molecular arms race is progressively being unveiled between prokaryotes and viruses. Prokaryotes utilize CRISPR-mediated adaptive immune systems to kill the invading phages and mobile genetic elements, and in turn, the viruses evolve diverse anti-CRISPR proteins to fight back. The structures of several anti-CRISPR proteins have now been reported, and here we discuss their structural features, with a particular emphasis on topology, to discover their similarities and differences. We summarize the CRISPR-Cas inhibition mechanisms of these anti-CRISPR proteins in their structural context. Considering anti-CRISPRs in this way will provide important clues for studying their origin and evolution.

Keywords: Adaptive immune; Anti-CRISPR; CRISPR-Cas system; Structure; Viral infection.

Fig. 1

A cartoon depicting the architecture of the type I-F CRISPR-Cas system and inhibition mechanisms of three type I-F anti-CRISPRs. a The type I-F Csy complex is a 350-kDa crRNA-guided surveillance complex composed of a 60-nucleotide crRNA and nine Cas proteins, which recruits a nuclease-helicase protein Cas3 for target degradation. b AcrF1 interacts with Cas7f, sterically hindering target DNA access to the crRNA guide. c AcrF2 interacts with Cas8f and Cas7f, resembling a DNA duplex and sterically hindering target dsDNA access to the binding pocket. d AcrF3 forms a homodimer, interacting with Cas3 and impeding the recruitment of Cas3 to Cascade

Fig. 2

Cartoon view of the structure of the type I-F Csy complex. a Structure of the type I-F Csy complex. The cas5f, cas6f, cas7f, and cas8f subunits of the type I-F Csy complex are colored cyan, yellow, green, and magenta, respectively. The crRNA is colored blue. b An enlarged view of the structure of crRNA, which resembles a string

Fig. 3

Cartoon view of the structure of the type I-F Csy complex bound to AcrF1/2. a Structure of the type I-F Csy complex bound to two anti-CRISPR proteins, AcrF1 and AcrF2. The cas5f, cas6f, cas7f, and cas8f subunits of the type I-F Csy complex are colored as Fig. 2a. AcrF1 and AcrF2 are colored red and orange, respectively. 3D structure (b) and topological view (c) of AcrF1. 3D structure (d) and topological view (e) of AcrF2. Helices and strands are shown as green cylinders and arrows, respectively

Fig. 4

Cartoon view of the structure of PaCas3 in complex with AcrF3. a Structure of the PaCas3–AcrF3 complex. PaCas3 is colored green. The AcrF3 dimer is colored cyan and yellow, respectively. The Cas2, RecA1/2, HD, and CTD domains of PaCas3 are labeled. b Topological view of AcrF3

Fig. 5

Cartoon view of the structure of the type I-F Csy complex bound to AcrF10. a Structure of the type I-F Csy complex bound to AcrF10. The cas5f, cas6f, cas7f, and cas8f subunits of the type I-F Csy complex are colored as Fig. 2a. AcrF10 is colored black. 3D structure (b) and topological view (c) of AcrF10

Fig. 6.

Cartoon and topological views of the structures of type II anti-CRISPR proteins. a Structure of the AcrIIA4–SpyCas9–sgRNA complex. SpyCas9 and AcrIIA4 are colored magenta and cyan, respectively. The crRNA is colored blue. The RuvC, CTD, and TOPO domains of SpyCas9 are labeled. b Topological view of AcrIIA4. c Structure of the AcrIIC1–HNH_NmeCas9 domain complex. AcrIIC1 and the HNH_NmeCas9 domain are colored cyan and orange, respectively. d Topological view of AcrIIC1. e Structure of the AcrIIA1 dimer. The AcrIIA1 dimer is colored cyan and green. f Topological view of AcrIIA1