Genetic determinants of PAM-dependent DNA targeting and pre-crRNA processing inSulfolobus islandicus

• DOI: 10.4161/rna.23798
• Publication Date: 2013-02-07T15:53:55Z
• Authors: Wenfang Peng; Huan Li; Søren Hallstrøm; Nan Peng; Yun Xiang Liang; Qunxin She
• Keywords: Trans-activating crRNA; Sulfolobus solfataricus; CRISPR interference; Nuclease

Bacteria and Archaea encode clustered, regularly interspaced, short palindromic repeat (CRISPR) systems to confer adaptive immunity invasive viruses plasmids. Recent studies of CRISPR revealed that diverse CRISPR-associated (Cas) interference modules often coexist in different organisms but functions cas genes have not been dissected any these systems. The crenarchaeon Sulfolobus islandicus encodes three distinct modules, including a type IA system two IIIB systems: Cmr-α Cmr-β. To study the genetic determinants protospacer-adjacent motif (PAM)-dependent DNA targeting activity mature RNA (crRNA) production this organism, mutants deleting individual or removing each other Cas were constructed. Characterization Cas7, Cas5, Cas6, Cas3′ Cas3" are essential for PAM-dependent activity, whereas Csa5, along with all is dispensable crenarchaeon. Cas6 implicated as only enzyme pre-crRNA processing crRNA maturation independent activity. Importantly, we show Cas7 Cas5 stabilizing intermediates crRNAs, respectively, depleting helicase nuclease domain Cas3 leads accumulation intermediates. This demonstrates addition proteins an archaeal also contribute processing.