CRISPR RNA

CRISPR RNA or crRNA is a RNA transcript from the CRISPR locus.^[1] CRISPR-Cas (clustered, regularly interspaced short palindromic repeats - CRISPR associated systems) is an adaptive immune system found in bacteria and archaea to protect against mobile genetic elements, like viruses, plasmids, and transposons.^[2] The CRISPR locus contains a series of repeats interspaced with unique spacers. These unique spacers can be acquired from MGEs.^[2]

Pre-crRNA is formed after the transcription of the CRISPR locus and before being processed by Cas proteins. Mature crRNA transcripts contain a partial conserved section of repeat and a sequence of spacer that is complementary to the target DNA.^[3] crRNA forms an effector complex with a single nuclease or multiple Cas proteins called a Cascade (CRISPR-associated complex for antiviral defense).^[3]^[1] Once the effector complex is formed a Cas nuclease or single effector protein will cause interference guided by the crRNA match.^[4]

Type-I

Type-I CRISPR systems are characterized by Cas3, a nuclease-helicase protein, and the multi-subunit Cascade (CRISPR-associated complex for antiviral defense). The crRNA can form a complex with the Cas proteins in the Cascade and guide the complex to the target DNA sequence. Cas3 is recruited for the nuclease-helicase activity.^[5]

Typically in the Cascade, Cas6 generates the mature crRNAs while Cas5 and Cas7 process and stabilize the crRNA.^[6]

Type-II

Type-II CRISPR systems^[7] are characterized by the single signature nuclease Cas9.^[8] In type-II CRISPR systems crRNA and tracrRNA (trans-activating CRISPR RNA) can form a complex known as the guide RNA or gRNA.^[9] The crRNA within the gRNA is what matches up with the target sequence or protospacer after the PAM is found. Once the match is made Cas9 will make a double-stranded break.

Type-III

Type-III CRISPR systems are characterized by Cas10, an RNA cleaving protein.^[10] Similar to type-I, a large subunit effector complex is formed and crRNA guides the complex to the target sequence. Cas6 helps to generate the mature crRNA.^[10]

Type-IV

Type-IV CRISPR systems do not have an effector nuclease and are associated with plasmids and prophages. A Cas6-like enzyme is associated with the maturation of the crRNA. Not all type-IV systems have a CRISPR locus and therefore do not have crRNA.^[11]

Type-V

Type-V CRISPR systems are characterized by Cas12, a nuclease that can cleave ssDNA, dsDNA, and RNA.^[7] Like Cas9, Cas12 is the single effector nuclease. Type-V systems process pre-crRNA without tracrRNA. The mature crRNA in complex with Cas12 target the DNA sequence of interest and cleave the DNA.^[12]

Type-VI

Type-VI CRISPR systems are characterized by Cas13, a single effector protein that targets RNA. Like the type-V system, Cas13 can process the pre-crRNA without tracrRNA. The mature crRNA in complex with Cas13 guides the complex to the target RNA and degrades it.^[13]

Function