Abstract

Among the >100 anti-CRISPRs (Acrs) identified to date, the vast majority inhibit CRISPR-Cas immunity on its own. Here we report a multilayered interaction between CRISPR-Cas subtype I-A immunity and two Acrs encoded adjacently in the genome of Saccharolobus virus SIRV2, gp47 (AcrIA3) and gp48 (previously known as AcrIIIB1, hence termed AcrIIIB1/AcrIA4). The host subtype I-A CRISPR-Cas interference module was shown previously to be up-regulated upon SIRV2 infection, through the release of transcriptional repressor Csa3b from the promoter. We demonstrate that AcrIIIB1/AcrIA4 on its own increases viral infectivity 4-5 orders of magnitude in the presence of the host subtype I-A CRISPR-Cas immunity. This Acr is able to completely inhibit the subtype I-A CRISPR-Cas immunity when the transcriptional activation of the latter is artificially removed, suggesting that Acrs might be one of the driving forces for the evolution of CRISPR-Cas up-regulation. Interestingly, AcrIA3 cooperates with AcrIIIB1/AcrIA4 by inhibiting transcriptional activation of the host subtype I-A CRISPR-Cas interference module through interaction with the promoter of the latter. Taken together, our data shed light on how virus-host arms race shaped the evolution of CRISPR-Cas and Acrs.