First indication for a functional CRISPR/Cas system in Francisella tularensis

Abstract

F. tularensis is a zoonotic agent and the subspecies novicida is proposed to be a waterassociated bacterium. The intracellular pathogen Francisella tularensis causes tularemia in humans and is known for its potential to be used as biological threat. We analyzed the genome sequence of F. tularensis subsp. novicida U112 in silico for the presence of a putative functional CRISPR/Cas (Clustered Regularly Interspaced Short Palindromic

A CRISPR-Cas system enhances envelope integrity mediating antibiotic resistance and inflammasome evasion

“…We demonstrated a role for components of a type II-B CRISPR-Cas system, which are encoded predominantly in pathogens and commensals (8)(9)(10), in the regulation of a membrane lipoprotein produced by the intracellular pathogen Francisella novicida (11). Through the action of the RNA-directed endonuclease Cas9 and two small RNAs, tracrRNA and scaRNA, the transcript for a bacterial lipoprotein (BLP; FTN_1103) is targeted and its stability altered, resulting in a decrease in protein production (SI Appendix, Fig.…”

Degeneration of a CRISPR/Cas system and its regulatory target during the evolution of a pathogen

“…Further suggesting that the Type II system is active in the targeting of foreign nucleic acid, most F. novicida genomes encode spacers identical to sequences in a predicted prophage present in the genome of a single known isolate of F. novicida. 7 Interestingly, this isolate does not encode such spacers, potentially explaining why it harbors this prophage. 7 In addition, F. novicida genomes encode a second CRISPR/Cas locus that most closely resembles a Type II locus in its architecture, but rather than Cas9, it encodes a novel Cas protein with no homology to known proteins.…”

“…8 In addition to the role of components of the Type II CRISPR/ Cas system in F. novicida pathogenesis, this system is predicted to be functional in the canonical role of targeting foreign nucleic acid. 7 The Type II CRISPR/Cas locus in F. novicida genomes encodes full-length forms of all the necessary components for the adaptation (cas1, cas2, cas4) and effector phases (cas9, crRNA, tracrRNA-also required for crRNA processing and interaction of the crRNA with Cas9) of targeting foreign DNA, as compared with functional Type II systems in Streptococcus spp and other bacteria. Further suggesting that the Type II system is active in the targeting of foreign nucleic acid, most F. novicida genomes encode spacers identical to sequences in a predicted prophage present in the genome of a single known isolate of F. novicida.…”

“…7 In addition, F. novicida genomes encode a second CRISPR/Cas locus that most closely resembles a Type II locus in its architecture, but rather than Cas9, it encodes a novel Cas protein with no homology to known proteins. 7 In contrast, we and others observe that the CRISPR/Cas systems present in the more virulent F. holarctica, F. mediasiatica, and F. tularensis species, have degenerated and lack critical components for CRISPR/Cas functionality (Fig. 1A).…”

“…5,6 The Gram-negative intracellular pathogen, Francisella novicida, encodes a Type-II CRISPR/Cas system, which is characterized by the presence of the Cas9 endonuclease. 1,7 We recently established the importance of this system in the pathogenesis of F. novicida. 8 Like other Francisella species, F. novicida is capable of infecting and replicating within the cytosol of a variety of host cells, including phagocytic cells of the innate immune system.…”