A CRISPR/Cas system mediates bacterial innate immune evasion and virulence

Sampson TR, Saroj SD, Llewellyn AC, Tzeng YL, Weiss DS.

Speaker: Bo-yang Tsai (蔡博仰)                Time: 14:10~15:00, Apr.30, 2014

Commentator: Dr. I-hsiu Huang (黃一修老師)          Place: Room 601

Abstract:

F. novicida is a model intracellular pathogen that evades host defenses as it traffics through the phagosome to replicate within the cytosol.¹ Previous studies by the authors demonstrated that the F. novicida FTN_1103, encoding a bacterial lipoprotein (BLP), is overproduced in the FTN_0757 mutant and increased TLR2-dependent IL-6 production, although the mechanism remain unclear. It appeared that F. novicida FTN_0757 can facilitate the evasion of TLR2 recognition through repression of FTN_1103 expression.² In this study, the authors wanted to figure out the mechanism of how F. noviceda subverts the host defense via FTN_0757 regulation. Unexpectedly, the bioinformatic analysis revealed that protein sequence of FTN_0757 is similar to Cas9, typically known to degrade foreign DNA targeted by crRNAs and it is present in a complete type II CRISPR/Cas locus containing a unique small CRISPR/Cas-associated RNA (scaRNA) not previously described in the locus. To test whether FTN_0757 (Cas9) repress FTN_1103 (BLP) required the CRISPR/Cas system, the authors deleted the Cas1, Cas2, Cas4, Cas9 genes,crRNA, tracrRNA and scRNA, as well as make point mutations in the RuvC/HNH endonuclease domain of Cas9. However the results showed that the Cas9, tracrRNA, scaRNA mediated the repression of FTN_1103 without the canonical crRNA involved and none of the RuvC or HNH engonuclease were required for this activity. Sequence hybridization prediction andimmunoprecipitation results also showed that Cas9 interacts with the tracrRNA-scaRNA-FTN_1103 mRNA via a arginine-rich motif (ARM) and any of three mutations in the complex resulted in the inability to repress FTN_1103. Finally, the authors analyzed the temporal expression of these components during the infection to determine whether repression of FTN_1103 was an active evasion process. Indeed these data indicated that Cas9, tracrRNA, scrRNA are induced during in vitro and intracellular infection resulted in mice death in vivo. In conclusion, the authors proposed a model whereby the bacterial CRISPR/Cas system can regulate the endogenous gene, ultimately promoting pathogensis.

References:

1.         Jones, C.L., et al. Subversion of host recognition and defense systems by Francisella spp. Microbiology and molecular biology reviews : MMBR 76, 383-404 (2012).

2.         Jones, C.L., Sampson, T.R., Nakaya, H.I., Pulendran, B. & Weiss, D.S. Repression of bacterial lipoprotein production by Francisella novicida facilitates evasion of innate immune recognition. Cellular microbiology 14, 1531-1543 (2012).