CONICET | Buscador de Institutos y Recursos Humanos

Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas) areconsidered the adaptive immune system of bacteria and archaea. They play a fundamental role in defending thehost against plasmid and phage invasion. CRISPR-Cas unique defense mechanism involves acquisition of foreignDNA as spacers in order to gain immunity, target recognition and cleavage of exogenous DNA or RNA. There isstill much to be revealed of the external factors that activate these systems. The aim of this work was tounderstand the impact that these factors play in the expression of the type I-F CRISPR-Cas system fromShewanella sp. Sh95. This strain is a gram-negative rod isolated from a patient with an ocular infection from apublic hospital of Argentina. Its CRISPR-Cas type I-F system comprises the cas operon (cas1, cas2/3, csy1, csy2,csy3 and csy4), a leader region and CRISPR array with 152 elements. Under standard growth conditions weobserved by RT-PCR that the cas operon and the whole CRISPR array were transcribed suggesting that thiselement is active. We then assessed the effect of sucrose (20%) and NaCl (3.5 and 10%) on CRISPR-Cas systemactivity. qRT-PCR assays showed that NaCl did not have significant impact on the system activity, howeversucrose stress resulted in a 4-fold increase of csy1 and csy4 after 1h exposure and no significant fold-variationsfor cas2/3. Our results confirm that Shewanella sp. Sh95 has an active CRISPR-Cas system. Furthermore, in thepresence of high sucrose concentrations the expression of the Cascade complex was increased, which could bethe result of cell membrane integrity weakening. This work provides new insights on the external factors thatactivate CRISPR-Cas systems, which could be used as a tool to control immunity against mobile genetic elements